[ { "id": "https://authors.library.caltech.edu/records/npa54-0gt95", "eprint_status": "archive", "datestamp": "2024-01-09 21:42:08", "lastmod": "2024-01-09 21:42:08", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Cho-Hyunju", "name": { "family": "Cho", "given": "Hyunju" }, "orcid": "0000-0001-7393-657X" }, { "id": "Liu-Yumeng", "name": { "family": "Liu", "given": "Yumeng" }, "orcid": "0000-0001-6581-0184" }, { "id": "Chung-SangYoon", "name": { "family": "Chung", "given": "SangYoon" }, "orcid": "0000-0002-0592-4099" }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Weiss-Shimon", "name": { "family": "Weiss", "given": "Shimon" }, "orcid": "0000-0002-0720-5426" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Dynamic stability of Sgt2 enables selective and privileged client handover in a chaperone triad", "ispublished": "pub", "full_text_status": "public", "keywords": "General Physics and Astronomy; General Biochemistry, Genetics and Molecular Biology; General Chemistry; Multidisciplinary", "note": "
© The Author(s) 2024. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
\n\nWe thank members of the Shan laboratory for comments on the manuscript and A. Siegel for assistance with the quantification of Bpa crosslinking data. This work was supported by National Institutes of Health grants R35 GM136321 to S.-o. Shan and by Dean Willard Chair funds to S.W.
\n\nH.C., Y.L., S.S. designed research; H.C., Y.L. and S. Chandrasekar performed biochemical experiments and analyzed data; Y.L. and S. Chung performed µs-ALEX experiments and analyzed data; S.W. provided guidance for µs-ALEX analysis; H.C., Y.L. and S.S. wrote the manuscript with input from S. Chandrasekar, S. Chung, and S.W.
\n\nAll data generated in this study are provided in the manuscript’s main text, Supplemental information, Supplementary Data 1, and Source Data file. A previously published structure of E.coli DnaK used to design fluorescent dye labeling positions is available under PDB code 4EZO. Source data are provided with this paper.
\nAll mpH2MM analyses in this study were performed using the Python packages, burstH2MM (Available at https://github.com/harripd/burstH2MM) and H2MM_C (Available at https://github.com/harripd/H2MMpythonlib).
\nThe authors declare no competing interests.
\nPublished - sciadv.adf5336.pdf
Supplemental Material - sciadv.adf5336_movie_s1.zip
Supplemental Material - sciadv.adf5336_sm.pdf
", "abstract": "Ring-forming AAA\u207a chaperones solubilize protein aggregates and protect organisms from proteostatic stress. In metazoans, the AAA\u207a chaperone Skd3 in the mitochondrial intermembrane space (IMS) is critical for human health and efficiently refolds aggregated proteins, but its underlying mechanism is poorly understood. Here, we show that Skd3 harbors both disaggregase and protein refolding activities enabled by distinct assembly states. High-resolution structures of Skd3 hexamers in distinct conformations capture ratchet-like motions that mediate substrate extraction. Unlike previously described disaggregases, Skd3 hexamers further assemble into dodecameric cages in which solubilized substrate proteins can attain near-native states. Skd3 mutants defective in dodecamer assembly retain disaggregase activity but are impaired in client refolding, linking the disaggregase and refolding activities to the hexameric and dodecameric states of Skd3, respectively. We suggest that Skd3 is a combined disaggregase and foldase, and this property is particularly suited to meet the complex proteostatic demands in the mitochondrial IMS.", "date": "2023-05-10", "date_type": "published", "publication": "Science Advances", "volume": "9", "number": "19", "publisher": "American Association for the Advancement of Science", "pagerange": "Art. No. eadf5336", "id_number": "CaltechAUTHORS:20230519-1772000.24", "issn": "2375-2548", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20230519-1772000.24", "funders": { "items": [ { "agency": "NIH", "grant_number": "R35 GM136321" }, { "agency": "Burroughs Wellcome Fund" }, { "agency": "NIH Postdoctoral Fellowship", "grant_number": "F32 GM137463" }, { "agency": "NIH", "grant_number": "R35 GM140847" }, { "agency": "Howard Hughes Medical Institute (HHMI)" }, { "agency": "NIH", "grant_number": "S10OD021741" }, { "agency": "NIH", "grant_number": "1S10OD026881" }, { "agency": "NIH", "grant_number": "U24 GM129541" } ] }, "doi": "10.1126/sciadv.adf5336", "pmcid": "PMC10171807", "primary_object": { "basename": "sciadv.adf5336.pdf", "url": "https://authors.library.caltech.edu/records/aeex6-fvw30/files/sciadv.adf5336.pdf" }, "related_objects": [ { "basename": "sciadv.adf5336_sm.pdf", "url": "https://authors.library.caltech.edu/records/aeex6-fvw30/files/sciadv.adf5336_sm.pdf" }, { "basename": "sciadv.adf5336_movie_s1.zip", "url": "https://authors.library.caltech.edu/records/aeex6-fvw30/files/sciadv.adf5336_movie_s1.zip" } ], "resource_type": "article", "pub_year": "2023", "author_list": "Gupta, Arpit; Lentzsch, Alfred M.; et el." }, { "id": "https://authors.library.caltech.edu/records/d4cyd-wtr51", "eprint_id": 119071, "eprint_status": "archive", "datestamp": "2023-08-22 18:42:14", "lastmod": "2023-10-24 23:49:19", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Role of Hsp70 in Post-Translational Protein Targeting: Tail-Anchored Membrane Proteins and Beyond", "ispublished": "pub", "full_text_status": "public", "keywords": "Inorganic Chemistry; Organic Chemistry; Physical and Theoretical Chemistry; Computer Science Applications; Spectroscopy; Molecular Biology; General Medicine; Catalysis", "note": "\u00a9 2023 by the author. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). \n\nS.o.S. is supported by grant R35 GM135321 from NIGMS.\n\nPublished - ijms-24-01170.pdf
", "abstract": "The Hsp70 family of molecular chaperones acts as a central 'hub' in the cell that interacts with numerous newly synthesized proteins to assist in their biogenesis. Apart from its central and well-established role in facilitating protein folding, Hsp70s also act as key decision points in the cellular chaperone network that direct client proteins to distinct biogenesis and quality control pathways. In this paper, we review accumulating data that illustrate a new branch in the Hsp70 network: the post-translational targeting of nascent membrane and organellar proteins to diverse cellular organelles. Work in multiple pathways suggests that Hsp70, via its ability to interact with components of protein targeting and translocation machineries, can initiate elaborate substrate relays in a sophisticated cascade of chaperones, cochaperones, and receptor proteins, and thus provide a mechanism to safeguard and deliver nascent membrane proteins to the correct cellular membrane. We discuss the mechanistic principles gleaned from better-studied Hsp70-dependent targeting pathways and outline the observations and outstanding questions in less well-studied systems.", "date": "2023-01-09", "date_type": "published", "publication": "International Journal of Molecular Sciences", "volume": "24", "number": "2", "publisher": "MDPI", "pagerange": "Art. No. 1170", "id_number": "CaltechAUTHORS:20230206-9587900.32", "issn": "1422-0067", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20230206-9587900.32", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "R35 GM135321" } ] }, "doi": "10.3390/ijms24021170", "pmcid": "PMC9866221", "primary_object": { "basename": "ijms-24-01170.pdf", "url": "https://authors.library.caltech.edu/records/d4cyd-wtr51/files/ijms-24-01170.pdf" }, "resource_type": "article", "pub_year": "2023", "author_list": "Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/p1mbn-5f986", "eprint_id": 116303, "eprint_status": "archive", "datestamp": "2023-08-22 17:25:06", "lastmod": "2023-12-22 23:41:32", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Yang-Chien-I", "name": { "family": "Yang", "given": "Chien-I" }, "orcid": "0000-0001-8606-5013" }, { "id": "Zhu-Zikun", "name": { "family": "Zhu", "given": "Zikun" }, "orcid": "0000-0001-5934-8368" }, { "id": "Jones-Jeffrey-J", "name": { "family": "Jones", "given": "Jeffrey J." } }, { "id": "Lomenick-Brett", "name": { "family": "Lomenick", "given": "Brett" }, "orcid": "0000-0002-5023-9998" }, { "id": "Chou-Tsui-Fen", "name": { "family": "Chou", "given": "Tsui-Fen" }, "orcid": "0000-0003-2410-2186" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "System-wide analyses reveal essential roles of N-terminal protein modification in bacterial membrane integrity", "ispublished": "pub", "full_text_status": "public", "keywords": "Molecular biology; Bacteriology; Omics; Multidisciplinary", "note": "\u00a9 2022 The Authors. Under a Creative Commons license - Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0).\n\nReceived 7 April 2022, Revised 20 June 2022, Accepted 7 July 2022, Available online 15 July 2022, Version of Record 31 July 2022. \n\nWe thank D. Newman for critical discussions and members of the Shan lab for comments on the manuscript. The E. coli strains CAG12184 and KPS73 (\u0394fmt) are generous gifts from A. Varshavsky. We thank L. Shan, F. Wang, and C. Sanfiorenzo for the assistance in MS sample preparation, data analysis, and microscopy. Sequencing was performed at the Millard and Muriel Jacobs Genetics and Genomics Laboratory at California Institute of Technology. This work was supported by NIH grant R35 GM136321 to S.S. and Think Global Education Trust Fellowship to C.-I.Y. \n\nAuthor contributions. Conceptualization: C.-I.Y. and S.S.; Investigation: C.-I.Y., Z.Z., and B.L.; Formal analysis: C.-I.Y, Z.Z., and J.J.; Software: Z.Z. and J.J.; Writing \u2013 original draft: C.-I.Y; Writing \u2013 review & editing: C. -I.Y, Z.Z., J.J., B.L., T.-F.C, and S.S.; Supervision: T.-F.C, and S.S. \n\nData and code availability: \nThe mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE (Perez-Riverol et al., 2022) partner, and the ribosome profiling data is available at GEO. The data are publicly available as of the date of publication. Accession numbers are listed in the key resources table. \n\nAll original code has been deposited to the ProteomeXchange Consortium and is publicly available as of the date of publication. The accession number is listed in the key resources table. \n\nAny additional information required to reanalyze the data reported in this paper is available from the lead contact upon request. \n\nThe authors declare no competing interests.\n\nPublished - 1-s2.0-S2589004222010288-main.pdf
Supplemental Material - 1-s2.0-S2589004222010288-mmc1.pdf
Supplemental Material - 1-s2.0-S2589004222010288-mmc2.xlsx
Supplemental Material - 1-s2.0-S2589004222010288-mmc3.xlsx
Supplemental Material - 1-s2.0-S2589004222010288-mmc4.xlsx
Supplemental Material - 1-s2.0-S2589004222010288-mmc5.xlsx
", "abstract": "The removal of the N-terminal formyl group on nascent proteins by peptide deformylase (PDF) is the most prevalent protein modification in bacteria. PDF is a critical target of antibiotic development; however, its role in bacterial physiology remains a long-standing question. This work used the time-resolved analyses of the Escherichia coli translatome and proteome to investigate the consequences of PDF inhibition. Loss of PDF activity rapidly induces cellular stress responses, especially those associated with protein misfolding and membrane defects, followed by a global down-regulation of metabolic pathways. Rapid membrane hyperpolarization and impaired membrane integrity were observed shortly after PDF inhibition, suggesting that the plasma membrane disruption is the most immediate and primary consequence of formyl group retention on nascent proteins. This work resolves the physiological function of a ubiquitous protein modification and uncovers its crucial role in maintaining the structure and function of the bacterial membrane.", "date": "2022-08-19", "date_type": "published", "publication": "iScience", "volume": "25", "number": "8", "publisher": "Cell Press", "pagerange": "Art. No. 104756", "id_number": "CaltechAUTHORS:20220816-373597000", "issn": "2589-0042", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20220816-373597000", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "R35 GM136321" }, { "agency": "Think Global Education Trust" } ] }, "local_group": { "items": [ { "id": "Millard-and-Muriel-Jacobs-Genetics-and-Genomics-Laboratory" }, { "id": "Division-of-Biology-and-Biological-Engineering" } ] }, "doi": "10.1016/j.isci.2022.104756", "pmcid": "PMC9356101", "primary_object": { "basename": "1-s2.0-S2589004222010288-main.pdf", "url": "https://authors.library.caltech.edu/records/p1mbn-5f986/files/1-s2.0-S2589004222010288-main.pdf" }, "related_objects": [ { "basename": "1-s2.0-S2589004222010288-mmc1.pdf", "url": "https://authors.library.caltech.edu/records/p1mbn-5f986/files/1-s2.0-S2589004222010288-mmc1.pdf" }, { "basename": "1-s2.0-S2589004222010288-mmc2.xlsx", "url": "https://authors.library.caltech.edu/records/p1mbn-5f986/files/1-s2.0-S2589004222010288-mmc2.xlsx" }, { "basename": "1-s2.0-S2589004222010288-mmc3.xlsx", "url": "https://authors.library.caltech.edu/records/p1mbn-5f986/files/1-s2.0-S2589004222010288-mmc3.xlsx" }, { "basename": "1-s2.0-S2589004222010288-mmc4.xlsx", "url": "https://authors.library.caltech.edu/records/p1mbn-5f986/files/1-s2.0-S2589004222010288-mmc4.xlsx" }, { "basename": "1-s2.0-S2589004222010288-mmc5.xlsx", "url": "https://authors.library.caltech.edu/records/p1mbn-5f986/files/1-s2.0-S2589004222010288-mmc5.xlsx" } ], "resource_type": "article", "pub_year": "2022", "author_list": "Yang, Chien-I; Zhu, Zikun; et el." }, { "id": "https://authors.library.caltech.edu/records/1bpmj-5ry74", "eprint_id": 115142, "eprint_status": "archive", "datestamp": "2023-08-22 16:17:01", "lastmod": "2023-10-24 15:25:19", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Zhu-Zikun", "name": { "family": "Zhu", "given": "Zikun" }, "orcid": "0000-0001-5934-8368" }, { "id": "Wang-Shuai", "name": { "family": "Wang", "given": "Shuai" }, "orcid": "0000-0002-8920-969X" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Ribosome profiling reveals multiple roles of SecA in cotranslational protein export", "ispublished": "pub", "full_text_status": "public", "keywords": "General Physics and Astronomy; General Biochemistry, Genetics and Molecular Biology; General Chemistry; Multidisciplinary", "note": "\u00a9 The Author(s) 2022. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. \n\nReceived 13 January 2022. Accepted 26 May 2022. Published 13 June 2022. \n\nWe thank A. Buskirk, J. Chartron and J. Chen for advice on the ribosome profiling protocol, P. He for advice on data analysis, and members of the Shan lab for discussions and advice. Sequencing was performed at the Millard and Muriel Jacobs Genetics and Genomics Laboratory at California Institute of Technology. This work was supported by NIH grant R35 GM136321 to S.-o.S. \n\nThese authors contributed equally: Zikun Zhu, Shuai Wang. \n\nContributions. Conceptualization, Z.Z., S.W., and S.-o.S.; Methodology, Z.Z. and S.W.; Investigation, Z.Z. and S.W.; Formal Analysis, Z.Z.; Data curation, Z.Z.; Visualization, Z.Z.; Software, Z.Z. and S.W.; Resources, Z.Z. and S.W.; Writing \u2013 Original Draft, Z.Z. and S.-o.S.; Writing \u2013 Review & Editing, Z.Z., S.W., and S.-o.S.; Supervision, S.-o.S.; Funding Acquisition, S.-o.S. \n\nReporting summary. Further information on research design is available in the Nature Research Reporting Summary linked to this article. \n\nData availability. The data supporting the findings of this study are available from the corresponding authors upon reasonable request. The accession number for the data reported in this paper is GSE185572. The protein structures used to calculate absolute contact order were downloaded from AlphaFold Protein Structure Database (https://alphafold.ebi.ac.uk/). Source data for the figures and supplementary figures are provided as a Source Data file. Source data are provided with this paper. \n\nThe authors declare no competing interests. \n\nPeer review information. Nature Communications thanks the anonymous reviewers for their contribution to the peer review of this work.\n\nPublished - s41467-022-31061-5.pdf
Supplemental Material - 41467_2022_31061_MOESM1_ESM.pdf
Supplemental Material - 41467_2022_31061_MOESM2_ESM.pdf
Supplemental Material - 41467_2022_31061_MOESM3_ESM.xlsx
Supplemental Material - 41467_2022_31061_MOESM4_ESM.pdf
Supplemental Material - 41467_2022_31061_MOESM5_ESM.xlsx
", "abstract": "SecA, an ATPase known to posttranslationally translocate secretory proteins across the bacterial plasma membrane, also binds ribosomes, but the role of SecA's ribosome interaction has been unclear. Here, we used a combination of ribosome profiling methods to investigate the cotranslational actions of SecA. Our data reveal the widespread accumulation of large periplasmic loops of inner membrane proteins in the cytoplasm during their cotranslational translocation, which are specifically recognized and resolved by SecA in coordination with the proton motive force (PMF). Furthermore, SecA associates with 25% of secretory proteins with highly hydrophobic signal sequences at an early stage of translation and mediates their cotranslational transport. In contrast, the chaperone trigger factor (TF) delays SecA engagement on secretory proteins with weakly hydrophobic signal sequences, thus enforcing a posttranslational mode of their translocation. Our results elucidate the principles of SecA-driven cotranslational protein translocation and reveal a hierarchical network of protein export pathways in bacteria.", "date": "2022-06-13", "date_type": "published", "publication": "Nature Communications", "volume": "13", "publisher": "Nature Publishing Group", "pagerange": "Art. No. 3393", "id_number": "CaltechAUTHORS:20220614-222089000", "issn": "2041-1723", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20220614-222089000", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM136321" } ] }, "local_group": { "items": [ { "id": "Millard-and-Muriel-Jacobs-Genetics-and-Genomics-Laboratory" } ] }, "doi": "10.1038/s41467-022-31061-5", "pmcid": "PMC9192764", "primary_object": { "basename": "s41467-022-31061-5.pdf", "url": "https://authors.library.caltech.edu/records/1bpmj-5ry74/files/s41467-022-31061-5.pdf" }, "related_objects": [ { "basename": "41467_2022_31061_MOESM1_ESM.pdf", "url": "https://authors.library.caltech.edu/records/1bpmj-5ry74/files/41467_2022_31061_MOESM1_ESM.pdf" }, { "basename": "41467_2022_31061_MOESM2_ESM.pdf", "url": "https://authors.library.caltech.edu/records/1bpmj-5ry74/files/41467_2022_31061_MOESM2_ESM.pdf" }, { "basename": "41467_2022_31061_MOESM3_ESM.xlsx", "url": "https://authors.library.caltech.edu/records/1bpmj-5ry74/files/41467_2022_31061_MOESM3_ESM.xlsx" }, { "basename": "41467_2022_31061_MOESM4_ESM.pdf", "url": "https://authors.library.caltech.edu/records/1bpmj-5ry74/files/41467_2022_31061_MOESM4_ESM.pdf" }, { "basename": "41467_2022_31061_MOESM5_ESM.xlsx", "url": "https://authors.library.caltech.edu/records/1bpmj-5ry74/files/41467_2022_31061_MOESM5_ESM.xlsx" } ], "resource_type": "article", "pub_year": "2022", "author_list": "Zhu, Zikun; Wang, Shuai; et el." }, { "id": "https://authors.library.caltech.edu/records/77bq6-apk91", "eprint_id": 113868, "eprint_status": "archive", "datestamp": "2023-10-19 17:14:26", "lastmod": "2023-10-19 17:14:26", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Yang-Chien-I", "name": { "family": "Yang", "given": "Chien-I" } }, { "id": "Kim-Jiwoo", "name": { "family": "Kim", "given": "Jiwoo" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Ribosome-nascent Chain Interaction Regulates N-terminal Protein Modification", "ispublished": "pub", "full_text_status": "public", "keywords": "N-terminal protein modification; ribosome; methionine aminopeptidase; peptide deformylase; cotranslational protein biogenesis; Molecular Biology; Structural Biology", "note": "\u00a9 2022 Published by Elsevier. \n\nReceived 28 December 2021, Revised 1 March 2022, Accepted 4 March 2022, Available online 10 March 2022, Version of Record 17 March 2022. \n\nWe thank members of the Shan lab for discussions and comments on the manuscript. The E. coli strains CAG12184 and KPS73, and the plasmids pHUE and pHUsp2-cc are generous gifts from A. Varshavsky. This work was supported by NIH grant R35 GM136321 to S.S. and Think Global Education Trust Fellowship to C.I.Y. \n\nCRediT authorship contribution statement: Chien-I Yang: Conceptualization, Investigation, Formal analysis, Writing \u2013 original draft, Writing \u2013 review & editing. Jiwoo Kim: Investigation. Shu-ou Shan: Supervision, Writing \u2013 review & editing. \n\nThe authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.\n\nSupplemental Material - 1-s2.0-S0022283622001097-mmc1.pdf
", "abstract": "Numerous proteins initiate their folding, localization, and modifications early during translation, and emerging data show that the ribosome actively participates in diverse protein biogenesis pathways. Here we show that the ribosome imposes an additional layer of substrate selection during N-terminal methionine excision (NME), an essential protein modification in bacteria. Biochemical analyses show that cotranslational NME is exquisitely sensitive to a hydrophobic signal sequence or transmembrane domain near the N terminus of the nascent polypeptide. The ability of the nascent chain to access the active site of NME enzymes dictates NME efficiency, which is inhibited by confinement of the nascent chain on the ribosome surface and exacerbated by signal recognition particle. In vivo measurements corroborate the inhibition of NME by an N-terminal hydrophobic sequence, suggesting the retention of formylmethionine on a substantial fraction of the secretory and membrane proteome. Our work demonstrates how molecular features of a protein regulate its cotranslational modification and highlights the active participation of the ribosome in protein biogenesis pathways via interactions of the ribosome surface with the nascent protein.", "date": "2022-05-15", "date_type": "published", "publication": "Journal of Molecular Biology", "volume": "434", "number": "9", "publisher": "Elsevier", "pagerange": "Art. No. 167535", "id_number": "CaltechAUTHORS:20220310-121624000", "issn": "0022-2836", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20220310-121624000", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM136321" }, { "agency": "Think Global Education Trust" } ] }, "doi": "10.1016/j.jmb.2022.167535", "pmcid": "PMC9126151", "primary_object": { "basename": "nihms-1792381.pdf", "url": "https://authors.library.caltech.edu/records/77bq6-apk91/files/nihms-1792381.pdf" }, "related_objects": [ { "basename": "1-s2.0-S0022283622001097-mmc1.pdf", "url": "https://authors.library.caltech.edu/records/77bq6-apk91/files/1-s2.0-S0022283622001097-mmc1.pdf" } ], "resource_type": "article", "pub_year": "2022", "author_list": "Yang, Chien-I; Kim, Jiwoo; et el." }, { "id": "https://authors.library.caltech.edu/records/bq01n-xax54", "eprint_id": 113613, "eprint_status": "archive", "datestamp": "2023-08-22 14:08:54", "lastmod": "2023-10-23 23:08:27", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Jomaa-Ahmad", "name": { "family": "Jomaa", "given": "Ahmad" }, "orcid": "0000-0002-5543-7942" }, { "id": "Gamerdinger-Martin", "name": { "family": "Gamerdinger", "given": "Martin" }, "orcid": "0000-0003-1483-8181" }, { "id": "Hsieh-Hao-Hsuan", "name": { "family": "Hsieh", "given": "Hao-Hsuan" }, "orcid": "0000-0001-9629-5832" }, { "id": "Wallisch-Annalena", "name": { "family": "Wallisch", "given": "Annalena" }, "orcid": "0000-0002-4582-9619" }, { "id": "Chandrasekaran-Viswanathan", "name": { "family": "Chandrasekaran", "given": "Viswanathan" }, "orcid": "0000-0002-0871-4740" }, { "id": "Ulusoy-Zeynel", "name": { "family": "Ulusoy", "given": "Zeynel" }, "orcid": "0000-0002-5476-3455" }, { "id": "Scaiola-Alain", "name": { "family": "Scaiola", "given": "Alain" }, "orcid": "0000-0003-3233-3910" }, { "id": "Hedge-Ramanujan-S", "name": { "family": "Hegde", "given": "Ramanujan S." }, "orcid": "0000-0001-8338-852X" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Ban-Nenad", "name": { "family": "Ban", "given": "Nenad" }, "orcid": "0000-0002-9527-210X" }, { "id": "Deuerling-Elke", "name": { "family": "Deuerling", "given": "Elke" }, "orcid": "0000-0002-8905-0260" } ] }, "title": "Mechanism of signal sequence handover from NAC to SRP on ribosomes during ER-protein targeting", "ispublished": "pub", "full_text_status": "public", "keywords": "Multidisciplinary", "note": "\u00a9 2022 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. This is an article distributed under the terms of the Science Journals Default License. \n\nReceived: 27 July 2021. Accepted: 27 January 2022. Published in print: 25 February 2022. \n\nWe thank M. Leibundgut, T. Lenarcic, and M. Jaskolowski for discussions; R. Schloemer and E. Coellen for technical assistance; and S. Kreft for help with in vitro cysteine cross-linking experiments. Cryo-EM was collected at ScopeM at the ETH Zurich. We acknowledge the MRC - LMB Electron Microscopy Facility for access and support of electron microscopy sample preparation and data collection for NAC-TTC5-RNC and the Caenorhabditis Genetics Center for strains. \n\nThis work was supported by the Swiss National Science Foundation (grant no. 310030B_163478); the National Center of Excellence in Research RNA & Disease Program of the SNSF (grant no. 51NF40_141735); a Roessler Prize, Ernst Jung Prize, and Otto Naegeli Prize for Medical Research (to N.B.); the German Science Foundation (grant nos. SFB969/A01 and A07 to E.D. and M.G.); the National Institutes of Health (grant no. R35 GM136321 to S.S.); the National Science Foundation (grant no. MCB-1929452 to S.-o.S); and the UK Medical Research Council (MRC grant MC_UP_A022_1007 to R.S.H.). V.C. was supported by V. Ramakrishnan, whose funding was from the MRC (grant no. MC_U105184332), the Wellcome Trust (grant no. WT096570), the Agouron Institute, and the Louis-Jeantet Foundation. We also acknowledge the support of the NVIDIA Corporation for the Titan Xp GPU through a grant awarded to A.J. \n\nAuthor contributions: A.J., M.G., H.-H.H., R.S.H., S.S., N.B., and E.D. conceived the project. A.J. and A.S. performed cryo-EM data collection for ER-targeting complexes containing NAC and SRP. A.J. determined the cryo-EM structures of NAC-RNC and NAC-SRP-RNC. M.G. and A.W. performed C. elegans in vivo and A.W. cross-linking experiments. H.-H.H. performed FRET titrations and single-molecule experiments. V.C. performed structural analysis of the NAC-TTC5-RNC. Z.U. characterized NAC cysteine variants. A.J., M.G., H.-H.H., S.S., E.D., and N.B. wrote the manuscript. All authors contributed to data analysis and the final version of the manuscript. \n\nData and materials availability: Cryo-EM maps and model coordinates are deposited in the EMDB as EMD-14191, EMD-14192, and EMD-14193 and in the PDB as PDB ID 7QWQ, 7QWR, and 7QWS for the NAC-SRP-RNC_(SS), NAC-RNC_(SS), and NAC-TTC5-RNC_(TUBB), respectively. All other data are available in the main text or the supplementary materials. \n\nThe authors declare no competing interests.\n\nAccepted Version - EMS142053.pdf
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", "abstract": "The nascent polypeptide\u2013associated complex (NAC) interacts with newly synthesized proteins at the ribosomal tunnel exit and competes with the signal recognition particle (SRP) to prevent mistargeting of cytosolic and mitochondrial polypeptides to the endoplasmic reticulum (ER). How NAC antagonizes SRP and how this is overcome by ER targeting signals are unknown. Here, we found that NAC uses two domains with opposing effects to control SRP access. The core globular domain prevented SRP from binding to signal-less ribosomes, whereas a flexibly attached domain transiently captured SRP to permit scanning of nascent chains. The emergence of an ER-targeting signal destabilized NAC's globular domain and facilitated SRP access to the nascent chain. These findings elucidate how NAC hands over the signal sequence to SRP and imparts specificity of protein localization.", "date": "2022-02-25", "date_type": "published", "publication": "Science", "volume": "375", "number": "6583", "publisher": "American Association for the Advancement of Science", "pagerange": "839-844", "id_number": "CaltechAUTHORS:20220225-724347000", "issn": "0036-8075", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20220225-724347000", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Swiss National Science Foundation (SNSF)", "grant_number": "310030B_163478" }, { "agency": "Swiss National Science Foundation (SNSF)", "grant_number": "51NF40_141735" }, { "agency": "ETH Z\u00fcrich Foundation" }, { "agency": "Ernst Jung Foundation" }, { "agency": "Bonizzi-Theler-Foundation" }, { "agency": "Deutsche Forschungsgemeinschaft (DFG)", "grant_number": "SFB969/A01" }, { "agency": "Deutsche Forschungsgemeinschaft (DFG)", "grant_number": "SFB969/A07" }, { "agency": "NIH", "grant_number": "R35 GM136321" }, { "agency": "NSF", "grant_number": "MCB-1929452" }, { "agency": "Medical Research Council (UK)", "grant_number": "MC_UP_A022_1007" }, { "agency": "Medical Research Council (UK)", "grant_number": "MC_U105184332" }, { "agency": "Wellcome Trust", "grant_number": "WT096570" }, { "agency": "Agouron Institute" }, { "agency": "Louis-Jeantet Foundation" }, { "agency": "NVIDIA Corporation" } ] }, "doi": "10.1126/science.abl6459", "pmcid": "PMC7612438", "primary_object": { "basename": "EMS142053.pdf", "url": "https://authors.library.caltech.edu/records/bq01n-xax54/files/EMS142053.pdf" }, "related_objects": [ { "basename": "science.abl6459_mdar_reproducibility_checklist.pdf", "url": "https://authors.library.caltech.edu/records/bq01n-xax54/files/science.abl6459_mdar_reproducibility_checklist.pdf" }, { "basename": "science.abl6459_sm.pdf", "url": "https://authors.library.caltech.edu/records/bq01n-xax54/files/science.abl6459_sm.pdf" } ], "resource_type": "article", "pub_year": "2022", "author_list": "Jomaa, Ahmad; Gamerdinger, Martin; et el." }, { "id": "https://authors.library.caltech.edu/records/5hk2p-kj860", "eprint_id": 112835, "eprint_status": "archive", "datestamp": "2023-08-22 13:06:52", "lastmod": "2023-10-23 22:48:20", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Hsieh-Hao-Hsuan", "name": { "family": "Hsieh", "given": "Hao-Hsuan" }, "orcid": "0000-0001-9629-5832" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Fidelity of Cotranslational Protein Targeting to the Endoplasmic Reticulum", "ispublished": "pub", "full_text_status": "public", "keywords": "protein targeting; signal recognition particle; nascent polypeptide-associated complex; ribosome; endoplasmic reticulum; membrane proteins; fidelity; Inorganic Chemistry; Organic Chemistry; Physical and Theoretical Chemistry; Computer Science Applications; Spectroscopy; Molecular Biology; General Medicine; Catalysis", "note": "\u00a9 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). \n\nReceived: 4 December 2021; Revised: 17 December 2021; Accepted: 18 December 2021; Published: 28 December 2021. \n\nThis work is funded by grants NSF-1929452 from the National Science Foundation and R01 GM078024 and R35 GM136321 from the National Institute of Health to S.S. \n\nAuthor Contributions: Writing\u2014original draft preparation, S.-o.S.; writing\u2014review and editing, H.-H.H.; visualization, H.-H.H. and S.-o.S. All authors have read and agreed to the published version of the manuscript. \n\nInstitutional Review Board Statement: Not applicable. \n\nInformed Consent Statement: Not applicable. \n\nThe authors declare no conflict of interest.\n\nPublished - ijms-23-00281.pdf
", "abstract": "Fidelity of protein targeting is essential for the proper biogenesis and functioning of organelles. Unlike replication, transcription and translation processes, in which multiple mechanisms to recognize and reject noncognate substrates are established in energetic and molecular detail, the mechanisms by which cells achieve a high fidelity in protein localization remain incompletely understood. Signal recognition particle (SRP), a conserved pathway to mediate the localization of membrane and secretory proteins to the appropriate cellular membrane, provides a paradigm to understand the molecular basis of protein localization in the cell. In this chapter, we review recent progress in deciphering the molecular mechanisms and substrate selection of the mammalian SRP pathway, with an emphasis on the key role of the cotranslational chaperone NAC in preventing protein mistargeting to the ER and in ensuring the organelle specificity of protein localization.", "date": "2022-01", "date_type": "published", "publication": "International Journal of Molecular Sciences", "volume": "23", "number": "1", "publisher": "MDPI", "pagerange": "Art. No. 281", "id_number": "CaltechAUTHORS:20220112-394479800", "issn": "1422-0067", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20220112-394479800", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NSF", "grant_number": "MCB-1929452" }, { "agency": "NIH", "grant_number": "R01 GM078024" }, { "agency": "NIH", "grant_number": "R35 GM136321" } ] }, "doi": "10.3390/ijms23010281", "pmcid": "PMC8745203", "primary_object": { "basename": "ijms-23-00281.pdf", "url": "https://authors.library.caltech.edu/records/5hk2p-kj860/files/ijms-23-00281.pdf" }, "resource_type": "article", "pub_year": "2022", "author_list": "Hsieh, Hao-Hsuan and Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/an1c4-dyw75", "eprint_id": 111604, "eprint_status": "archive", "datestamp": "2023-08-20 05:48:17", "lastmod": "2023-10-23 20:39:07", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Chio-Un-Seng", "name": { "family": "Chio", "given": "Un Seng" }, "orcid": "0000-0002-5295-2690" }, { "id": "Liu-Yumeng", "name": { "family": "Liu", "given": "Yumeng" } }, { "id": "Chung-SangYoon", "name": { "family": "Chung", "given": "SangYoon" }, "orcid": "0000-0002-0592-4099" }, { "id": "Shim-Woo-Jun", "name": { "family": "Shim", "given": "Woo Jun" }, "orcid": "0000-0001-9596-7393" }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Weiss-Shimon", "name": { "family": "Weiss", "given": "Shimon" }, "orcid": "0000-0002-0720-5426" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Subunit cooperation in the Get1/2 receptor promotes tail-anchored membrane protein insertion", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2021 Chio et al. This article is distributed under the terms of an Attribution\u2013Noncommercial\u2013Share Alike\u2013No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms/). After six months it is available under a Creative Commons License (Attribution\u2013Noncommercial\u2013Share Alike 4.0 International license, as described at https://creativecommons.org/licenses/by-nc-sa/4.0/). \n\nSubmitted: 12 March 2021; Revised: 3 August 2021; Accepted: 19 August 2021. \n\nWe thank Vladimir Denic for the mini-Get1/2 construct, Jennifer Keefe for help with SEC-MALS, and members of the Shan laboratory for general discussion and comments on the manuscript. \n\nThis work was supported by Dean Willard Chair funds to S. Weiss and National Institutes of Health grants R01 GM107368 and R35 GM136321 and the Gordon and Betty Moore Foundation grant GBMF2939 to S.-o. Shan. \n\nThe authors declare no competing financial interests.\n\nAuthor contributions: U.S. Chio, Y. Liu, and S.-o. Shan designed research; U.S. Chio, Y. Liu, W.J. Shim, and S. Chandrasekar performed biochemical experiments; U.S. Chio and Y. Liu analyzed biochemical data with input from S.-o. Shan; U.S. Chio, Y. Liu, and SY. Chung performed sm fluorescence experiments; U.S. Chio, Y. Liu, and SY. Chung analyzed biophysics data with input from S. Weiss and S.-o. Shan; U.S. Chio, Y. Liu, and S.-o. Shan wrote the manuscript; SY. Chung and S. Weiss revised the manuscript. All authors approved the final manuscript.\nU.S. Chio and Y. Liu contributed equally to this paper.\n\nPublished - jcb_202103079.pdf
", "abstract": "The guided entry of tail-anchored protein (GET) pathway, in which the Get3 ATPase delivers an essential class of tail-anchored membrane proteins (TAs) to the Get1/2 receptor at the endoplasmic reticulum, provides a conserved mechanism for TA biogenesis in eukaryotic cells. The membrane-associated events of this pathway remain poorly understood. Here we show that complex assembly between the cytosolic domains (CDs) of Get1 and Get2 strongly enhances the affinity of the individual subunits for Get3\u2022TA, thus enabling efficient capture of the targeting complex. In addition to the known role of Get1CD in remodeling Get3 conformation, two molecular recognition features (MoRFs) in Get2CD induce Get3 opening, and both subunits are required for optimal TA release from Get3. Mutation of the MoRFs attenuates TA insertion into the ER in vivo. Our results demonstrate extensive cooperation between the Get1/2 receptor subunits in the capture and remodeling of the targeting complex, and emphasize the role of MoRFs in receptor function during membrane protein biogenesis.", "date": "2021-11", "date_type": "published", "publication": "Journal of Cell Biology", "volume": "220", "number": "11", "publisher": "Rockefeller University Press", "pagerange": "Art. No. e202103079", "id_number": "CaltechAUTHORS:20211022-170110641", "issn": "0021-9525", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20211022-170110641", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Dean Willard Chair" }, { "agency": "NIH", "grant_number": "R01 GM107368" }, { "agency": "NIH", "grant_number": "R35 GM136321" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" } ] }, "doi": "10.1083/jcb.202103079", "primary_object": { "basename": "jcb_202103079.pdf", "url": "https://authors.library.caltech.edu/records/an1c4-dyw75/files/jcb_202103079.pdf" }, "resource_type": "article", "pub_year": "2021", "author_list": "Chio, Un Seng; Liu, Yumeng; et el." }, { "id": "https://authors.library.caltech.edu/records/sev89-fcy55", "eprint_id": 109790, "eprint_status": "archive", "datestamp": "2023-08-22 11:22:00", "lastmod": "2023-10-23 18:10:23", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Ji-Shuiling", "name": { "family": "Ji", "given": "Shuiling" } }, { "id": "Siegel-Alex", "name": { "family": "Siegel", "given": "Alex" }, "orcid": "0000-0003-3601-5178" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Grimm-Bernhard", "name": { "family": "Grimm", "given": "Bernhard" }, "orcid": "0000-0002-9730-1074" }, { "id": "Wang-Peng", "name": { "family": "Wang", "given": "Peng" }, "orcid": "0000-0001-8420-5326" } ] }, "title": "Chloroplast SRP43 autonomously protects chlorophyll biosynthesis proteins against heat shock", "ispublished": "pub", "full_text_status": "public", "keywords": "Chloroplasts; Heat; Plant physiology", "note": "\u00a9 2021 Nature Publishing Group. \n\nReceived 03 February 2021; Accepted 28 July 2021; Published 02 September 2021. \n\nWe thank D. Sch\u00fcnemann (Ruhr-Universit\u00e4t Bochum) for providing us the chaos and ffc mutants, the GST\u2013cpSRP43 expression vector, and antibodies against cpSRP43 and cpSRP54, and P. Hardy for critical reading of the manuscript. This work was supported by a grant from the Chinese Scholarship Council to S.J., grant nos R35 GM136321 and DOE.DE-SC0020661 to A.S. and S.S., and grants from the Deutsche Forschungsgemeinschaft to B.G. (nos FOR2092, GR 936/18-1 and SFB TRR175, subproject C04) and to P.W. (no. WA 4599/2-1). \n\nData availability: The data supporting the findings of this study are available within the article and its Supplementary Information. Source data are provided with this paper. \n\nThese authors contributed equally: Shuiling Ji, Alex Siegel. \n\nAuthor Contributions: B.G. and P.W. designed the experiments. S.J. and A.S. performed the experiments. S.J., A.S., S.S., B.G. and P.W. analysed the data. B.G. and P.W. wrote the manuscript. S.J., A.S. and S.S. gave critical comments and revisions to the manuscript. \n\nThe authors declare no competing interests. \n\nPeer review information: Nature Plants thanks Briardo Llorente, Tatsuru Masuda and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.\n\nAccepted Version - nihms-1780143.pdf
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", "abstract": "The assembly of light-harvesting chlorophyll-binding proteins (LHCPs) is coordinated with chlorophyll biosynthesis during chloroplast development. The ATP-independent chaperone known as chloroplast signal recognition particle 43 (cpSRP43) mediates post-translational LHCP targeting to the thylakoid membrane and also participates in tetrapyrrole biosynthesis (TBS). How these distinct actions of cpSRP43 are controlled has remained unclear. Here, we demonstrate that cpSRP43 effectively protects several TBS proteins from heat-induced aggregation and enhances their stability during leaf greening and heat shock. While the substrate-binding domain of cpSRP43 is sufficient for chaperoning LHCPs, the stabilization of TBS clients requires the chromodomain 2 of the protein. Strikingly, cpSRP54\u2014which activates cpSRP43's LHCP-targeted function\u2014inhibits the chaperone activity of cpSRP43 towards TBS proteins. High temperature weakens the interaction of cpSRP54 with cpSRP43, thus freeing cpSRP43 to interact with and protect the integrity of TBS proteins. Our data indicate that the temperature sensitivity of the cpSRP43\u2013cpSRP54 complex enables cpSRP43 to serve as an autonomous chaperone for the thermoprotection of TBS proteins.", "date": "2021-10", "date_type": "published", "publication": "Nature Plants", "volume": "7", "number": "10", "publisher": "Nature Publishing Group", "pagerange": "1420-1432", "id_number": "CaltechAUTHORS:20210713-201044719", "issn": "2055-026X", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20210713-201044719", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Chinese Scholarship Council" }, { "agency": "NIH", "grant_number": "R35 GM136321" }, { "agency": "Department of Energy (DOE)", "grant_number": "DE-SC0020661" }, { "agency": "Deutsche Forschungsgemeinschaft (DFG)", "grant_number": "FOR2092" }, { "agency": "Deutsche Forschungsgemeinschaft (DFG)", "grant_number": "GR 936/18-1" }, { "agency": "Deutsche Forschungsgemeinschaft (DFG)", "grant_number": "SFB TRR175" }, { "agency": "Deutsche Forschungsgemeinschaft (DFG)", "grant_number": "WA 4599/2-1" } ] }, "doi": "10.1038/s41477-021-00994-y", "pmcid": "PMC8879858", "primary_object": { "basename": "41477_2021_994_Fig11_ESM.webp", "url": "https://authors.library.caltech.edu/records/sev89-fcy55/files/41477_2021_994_Fig11_ESM.webp" }, "related_objects": [ { "basename": "41477_2021_994_Fig13_ESM.webp", "url": "https://authors.library.caltech.edu/records/sev89-fcy55/files/41477_2021_994_Fig13_ESM.webp" }, { "basename": "41477_2021_994_MOESM11_ESM.xlsx", "url": "https://authors.library.caltech.edu/records/sev89-fcy55/files/41477_2021_994_MOESM11_ESM.xlsx" }, { "basename": "41477_2021_994_MOESM9_ESM.xlsx", "url": "https://authors.library.caltech.edu/records/sev89-fcy55/files/41477_2021_994_MOESM9_ESM.xlsx" }, { "basename": "nihms-1780143.pdf", "url": "https://authors.library.caltech.edu/records/sev89-fcy55/files/nihms-1780143.pdf" }, { 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"https://authors.library.caltech.edu/records/sev89-fcy55/files/41477_2021_994_MOESM16_ESM.xlsx" }, { "basename": "41477_2021_994_MOESM5_ESM.xlsx", "url": "https://authors.library.caltech.edu/records/sev89-fcy55/files/41477_2021_994_MOESM5_ESM.xlsx" }, { "basename": "41477_2021_994_Fig15_ESM.webp", "url": "https://authors.library.caltech.edu/records/sev89-fcy55/files/41477_2021_994_Fig15_ESM.webp" }, { "basename": "41477_2021_994_MOESM17_ESM.xlsx", "url": "https://authors.library.caltech.edu/records/sev89-fcy55/files/41477_2021_994_MOESM17_ESM.xlsx" }, { "basename": "41477_2021_994_MOESM18_ESM.xlsx", "url": "https://authors.library.caltech.edu/records/sev89-fcy55/files/41477_2021_994_MOESM18_ESM.xlsx" }, { "basename": "41477_2021_994_MOESM4_ESM.xlsx", "url": "https://authors.library.caltech.edu/records/sev89-fcy55/files/41477_2021_994_MOESM4_ESM.xlsx" } ], "resource_type": "article", "pub_year": "2021", "author_list": "Ji, Shuiling; Siegel, Alex; et el." }, { "id": "https://authors.library.caltech.edu/records/1pcc0-ymq78", "eprint_id": 110312, "eprint_status": "archive", "datestamp": "2023-08-22 10:31:10", "lastmod": "2023-10-23 19:33:59", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Jomaa-Ahmad", "name": { "family": "Jomaa", "given": "Ahmad" } }, { "id": "Eitzinger-Simon", "name": { "family": "Eitzinger", "given": "Simon" } }, { "id": "Zhu-Zikun", "name": { "family": "Zhu", "given": "Zikun" } }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Kobayashi-Kan", "name": { "family": "Kobayashi", "given": "Kan" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Ban-Nenad", "name": { "family": "Ban", "given": "Nenad" }, "orcid": "0000-0002-9527-210X" } ] }, "title": "Molecular mechanism of cargo recognition and handover by the mammalian signal recognition particle", "ispublished": "pub", "full_text_status": "public", "keywords": "signal recognition particle; cryoelectron microscopy; co-translational protein targeting; GTPases; ribosome nascent chain complex; transmembrane domain; signal peptide; endoplasmic reticulum", "note": "\u00a9 2021 The Authors. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). \n\nReceived 21 December 2020, Revised 23 April 2021, Accepted 15 June 2021, Available online 13 July 2021. \n\nWe thank A. Scaiola and D. Boehringer for the support with EM data processing, M. Leibundgut for the support with model building, and Jae Ho Lee and members of the Ban and Shan groups for discussions and comments on the manuscript. Cryo-EM data were collected at the Scientific Center for Optical and Electron Microscopy at the ETH Zurich (ScopeM). We gratefully acknowledge the support of NVIDIA Corporation for the Titan Xp GPU used in this research through a GPU Grant program awarded to A.J. This work was supported by the Swiss National Science Foundation (SNSF) (grant no. 310030B_163478); the National Center of Excellence in Research (NCCR) RNA & Disease Program of the SNSF (grant no. 51NF40_141735), to N.B.; and National Institutes of Health grant GM107368, National Science Foundation grant MCB-1929452, and the Gordon and Betty Moore Foundation through grant GBMF2939 to S.S. \n\nAuthor contributions: A.J., N.B., and S.S. conceived the project. K.K. and A.J. prepared samples for the cryo-EM analysis. A.J. collected the cryo-EM data. A.J. and S.E. performed the reconstruction and model building. S.C. and Z.Z. purified the recombinant human SRP, SR, and performed the biochemical experiments. N.B., S.S., and A.J. supervised the research. A.J., N.B., and S.S. wrote the manuscript. All of the authors interpreted the data and contributed to the final versions of the manuscript. \n\nThe authors declare no competing interests. N.B. is a member of the Cell Reports advisory board.\n\nPublished - 1-s2.0-S2211124721007269-main.pdf
Supplemental Material - 1-s2.0-S2211124721007269-mmc1.pdf
", "abstract": "Co-translational protein targeting to membranes by the signal recognition particle (SRP) is a universally conserved pathway from bacteria to humans. In mammals, SRP and its receptor (SR) have many additional RNA features and protein components compared to the bacterial system, which were recently shown to play regulatory roles. Due to its complexity, the mammalian SRP targeting process is mechanistically not well understood. In particular, it is not clear how SRP recognizes translating ribosomes with exposed signal sequences and how the GTPase activity of SRP and SR is regulated. Here, we present electron cryo-microscopy structures of SRP and SRP\u00b7SR in complex with the translating ribosome. The structures reveal the specific molecular interactions between SRP and the emerging signal sequence and the elements that regulate GTPase activity of SRP\u00b7SR. Our results suggest the molecular mechanism of how eukaryote-specific elements regulate the early and late stages of SRP-dependent protein targeting.", "date": "2021-07-13", "date_type": "published", "publication": "Cell Reports", "volume": "36", "number": "2", "publisher": "Cell Press", "pagerange": "Art. No. 109350", "id_number": "CaltechAUTHORS:20210819-225523366", "issn": "2211-1247", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20210819-225523366", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NVIDIA Corporation" }, { "agency": "Swiss National Science Foundation (SNSF)", "grant_number": "310030B_163478" }, { "agency": "Swiss National Science Foundation (SNSF)", "grant_number": "51NF40_141735" }, { "agency": "NIH", "grant_number": "GM107368" }, { "agency": "NSF", "grant_number": "MCB-1929452" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" } ] }, "doi": "10.1016/j.celrep.2021.109350", "pmcid": "PMC8330425", "primary_object": { "basename": "1-s2.0-S2211124721007269-main.pdf", "url": "https://authors.library.caltech.edu/records/1pcc0-ymq78/files/1-s2.0-S2211124721007269-main.pdf" }, "related_objects": [ { "basename": "1-s2.0-S2211124721007269-mmc1.pdf", "url": "https://authors.library.caltech.edu/records/1pcc0-ymq78/files/1-s2.0-S2211124721007269-mmc1.pdf" } ], "resource_type": "article", "pub_year": "2021", "author_list": "Jomaa, Ahmad; Eitzinger, Simon; et el." }, { "id": "https://authors.library.caltech.edu/records/j2qf0-g5h52", "eprint_id": 100603, "eprint_status": "archive", "datestamp": "2023-08-20 03:18:44", "lastmod": "2023-10-20 23:15:49", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Lee-Jae-Ho", "name": { "family": "Lee", "given": "Jae Ho" }, "orcid": "0000-0002-8663-3209" }, { "id": "Jomaa-Ahmad", "name": { "family": "Jomaa", "given": "Ahmad" }, "orcid": "0000-0002-5543-7942" }, { "id": "Chung-SangYoon", "name": { "family": "Chung", "given": "SangYoon" }, "orcid": "0000-0002-0592-4099" }, { "id": "Hwang-Fu-Yu-Hsien", "name": { "family": "Hwang Fu", "given": "Yu-Hsien" }, "orcid": "0000-0002-2861-4843" }, { "id": "Qian-Ruilin", "name": { "family": "Qian", "given": "Ruilin" } }, { "id": "Sun-Xuemeng", "name": { "family": "Sun", "given": "Xuemeng" } }, { "id": "Hsieh-Hao-Hsuan", "name": { "family": "Hsieh", "given": "Hao-Hsuan" }, "orcid": "0000-0001-9629-5832" }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Bi-Xiaotian", "name": { "family": "Bi", "given": "Xiaotian" } }, { "id": "Mattei-Simone", "name": { "family": "Mattei", "given": "Simone" }, "orcid": "0000-0002-2005-8977" }, { "id": "Boehringer-Daniel", "name": { "family": "Boehringer", "given": "Daniel" }, "orcid": "0000-0002-6666-8447" }, { "id": "Weiss-Shimon", "name": { "family": "Weiss", "given": "Shimon" }, "orcid": "0000-0002-0720-5426" }, { "id": "Ban-Nenad", "name": { "family": "Ban", "given": "Nenad" }, "orcid": "0000-0002-9527-210X" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Receptor compaction and GTPase rearrangement drive SRP-mediated cotranslational protein translocation into the ER", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC). \n\nSubmitted 10 December 2020; Accepted 1 April 2021; Published 21 May 2021. \n\nWe thank the members of the Shan and Ban laboratory for comments on the manuscript. We thank A. Scaiola for the support with EM data processing and M. Leibundgut for the support with model building. Cryo-EM data were collected at the Scientific Center for Optical and Electron Microscopy at the ETH Zurich (ScopeM). \n\nThis work was supported by National Institutes of Health grants GM078024 and R35 GM136321, National Science Foundation grant MCB-1929452, and the Gordon and Betty Moore Foundation grant GBMF2939 to S.S.; by the Swiss National Science Foundation (SNSF) (grant number 310030B_163478) and National Center of Excellence in Research (NCCR) RNA and Disease Program of the SNSF (grant number 51NF40_141735) to N.B.; and by National Institutes of Health grant GM130942 and Dean Willard Chair funds to S.W. We gratefully acknowledge the support of NVIDIA Corporation for the Titan Xp GPU used in this research through a GPU Grant program awarded to A.J. \n\nAuthor contributions: J.H.L., A.J., Y.-H.H.F., N.B., and S.S. designed research. J.H.L., Y.-H.H.F., R.Q., X.S., H.-H.H., X.B., and S.C. performed biochemical experiments and analyzed data. A.J. and S.M. purified RNCs for cryo-EM data collection. A.J., S.M., and D.B. collected EM data. A.J. processed cryo-EM data and built atomic models. J.H.L., R.Q., and S.Y.C. performed \u03bcs-ALEX experiments and analyzed data. S.W. provided guidance for \u03bcs-ALEX analysis. J.H.L., S.S., A.J., and N.B. wrote the manuscript with input from S.C., H.-H.H., S.Y.C. and S.W. \n\nCompeting interests: S.W. is a consultant to Bio-Rad. The authors declare that they have no other competing interests. \n\nData and materials availability: All the data and associated procedures are described in the manuscript and/or in Supplementary Materials. Cryo-EM maps and model coordinates are deposited in the EMDB as EMD-12303, EMD-12304, and EMD-12305 and in the PDB as PDB ID 7NFX.\n\nPublished - eabg0942.full.pdf
Submitted - 2020.01.07.897827v1.full.pdf
Supplemental Material - abg0942_SM.pdf
", "abstract": "The conserved signal recognition particle (SRP) cotranslationally delivers ~30% of the proteome to the eukaryotic endoplasmic reticulum (ER). The molecular mechanism by which eukaryotic SRP transitions from cargo recognition in the cytosol to protein translocation at the ER is not understood. Here, structural, biochemical, and single-molecule studies show that this transition requires multiple sequential conformational rearrangements in the targeting complex initiated by guanosine triphosphatase (GTPase)\u2013driven compaction of the SRP receptor (SR). Disruption of these rearrangements, particularly in mutant SRP54G226E linked to severe congenital neutropenia, uncouples the SRP/SR GTPase cycle from protein translocation. Structures of targeting intermediates reveal the molecular basis of early SRP-SR recognition and emphasize the role of eukaryote-specific elements in regulating targeting. Our results provide a molecular model for the structural and functional transitions of SRP throughout the targeting cycle and show that these transitions provide important points for biological regulation that can be perturbed in genetic diseases.", "date": "2021-05-21", "date_type": "published", "publication": "Science Advances", "volume": "7", "number": "21", "publisher": "American Association for the Advancement of Science", "pagerange": "Art. No. eabg0942", "id_number": "CaltechAUTHORS:20200109-143242552", "issn": "2375-2548", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200109-143242552", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "NIH", "grant_number": "R35 GM136321" }, { "agency": "NSF", "grant_number": "MCB-1929452" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" }, { "agency": "Swiss National Science Foundation (SNSF)", "grant_number": "310030B_163478" }, { "agency": "Swiss National Science Foundation (SNSF)", "grant_number": "51NF40_141735" }, { "agency": "NIH", "grant_number": "GM130942" }, { "agency": "Dean Willard Chair" }, { "agency": "NVIDIA Corporation" } ] }, "doi": "10.1126/sciadv.abg0942", "pmcid": "PMC8139590", "primary_object": { "basename": "2020.01.07.897827v1.full.pdf", "url": "https://authors.library.caltech.edu/records/j2qf0-g5h52/files/2020.01.07.897827v1.full.pdf" }, "related_objects": [ { "basename": "abg0942_SM.pdf", "url": "https://authors.library.caltech.edu/records/j2qf0-g5h52/files/abg0942_SM.pdf" }, { "basename": "eabg0942.full.pdf", "url": "https://authors.library.caltech.edu/records/j2qf0-g5h52/files/eabg0942.full.pdf" } ], "resource_type": "article", "pub_year": "2021", "author_list": "Lee, Jae Ho; Jomaa, Ahmad; et el." }, { "id": "https://authors.library.caltech.edu/records/65zkb-mcf70", "eprint_id": 108470, "eprint_status": "archive", "datestamp": "2023-08-22 09:17:34", "lastmod": "2023-10-23 17:04:02", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Cho-Hyunju", "name": { "family": "Cho", "given": "Hyunju" }, "orcid": "0000-0001-7393-657X" }, { "id": "Shim-Woo-Jun", "name": { "family": "Shim", "given": "Woo Jun" }, "orcid": "0000-0001-9596-7393" }, { "id": "Liu-Yumeng", "name": { "family": "Liu", "given": "Yumeng" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "J-domain proteins promote client relay from Hsp70 during tail-anchored membrane protein targeting", "ispublished": "pub", "full_text_status": "public", "keywords": "molecular chaperone; Hsp70; Hsp40; tail-anchored protein; membrane proteins; protein targeting", "note": "\u00a9 2021 The Authors. Published by Elsevier Inc on behalf of American Society for Biochemistry and Molecular Biology.\nThis is an open access article under the CC\nBY license (http://creativecommons.org/licenses/by/4.0/).\n\nReceived 22 January 2021, Revised 22 February 2021, Accepted 15 March 2021, Available online 17 March 2021.\n\nWe thank D. Rapaport, T. Jores for the yeast strains, E. A. Craig for yeast strains and for antibodies against Ssa and Sis1, the Center for Molecular Medicine (California Institute of Technology) for use of the Li-Cor Odyssey imager, and members of the Shan laboratories for critical comments on the manuscript.\n\nData availability:\nAll data described are contained in the manuscript.\n\nAuthor contributions:\nH. C. and S. S. designed experiments; H. C., W. J. S., and Y. L. performed experiments and analyzed the data; H. C. and S. S. wrote the paper. All authors reviewed the results and approved the final version of the manuscript.\n\nFunding and additional information:\nThis work was supported by NIH grant R01 GM107368 and R35 GM136321 to S. S. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.\n\nConflict of interest:\nThe authors declare that they have no conflicts of interest with the contents of this article.\n\nPublished - 1-s2.0-S0021925821003240-main.pdf
Supplemental Material - 1-s2.0-S0021925821003240-mmc1.pdf
", "abstract": "J-domain proteins (JDPs) play essential roles in Hsp70 function by assisting Hsp70 in client trapping and regulating the Hsp70 ATPase cycle. Here, we report that JDPs can further enhance the targeting competence of Hsp70-bound client proteins during tail-anchored protein (TA) biogenesis. In the guided-entry-of-tail\u2013anchored protein pathway in yeast, nascent TAs are captured by cytosolic Hsp70 and sequentially relayed to downstream chaperones, Sgt2 and Get3, for delivery to the ER. We found that two JDPs, Ydj1 and Sis1, function in parallel to support TA targeting to the ER in vivo. Biochemical analyses showed that, while Ydj1 and Sis1 differ in their ability to assist Hsp70 in TA trapping, both JDPs enhance the transfer of Hsp70-bound TAs to Sgt2. The ability of the JDPs to regulate the ATPase cycle of Hsp70 is essential for enhancing the transfer competence of Hsp70-bound TAs in vitro and for supporting TA insertion in vivo. These results demonstrate a role of JDPs in regulating the conformation of Hsp70-bound clients during membrane protein biogenesis.", "date": "2021-03-17", "date_type": "published", "publication": "Journal of Biological Chemistry", "volume": "296", "publisher": "American Society for Biochemistry and Molecular Biology", "pagerange": "Art. No. 100546", "id_number": "CaltechAUTHORS:20210317-150527513", "issn": "0021-9258", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20210317-150527513", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "R01 GM107368" }, { "agency": "NIH", "grant_number": "R35 GM136321" } ] }, "doi": "10.1016/j.jbc.2021.100546", "primary_object": { "basename": "1-s2.0-S0021925821003240-main.pdf", "url": "https://authors.library.caltech.edu/records/65zkb-mcf70/files/1-s2.0-S0021925821003240-main.pdf" }, "related_objects": [ { "basename": "1-s2.0-S0021925821003240-mmc1.pdf", "url": "https://authors.library.caltech.edu/records/65zkb-mcf70/files/1-s2.0-S0021925821003240-mmc1.pdf" } ], "resource_type": "article", "pub_year": "2021", "author_list": "Cho, Hyunju; Shim, Woo Jun; et el." }, { "id": "https://authors.library.caltech.edu/records/4qreq-f7w72", "eprint_id": 106667, "eprint_status": "archive", "datestamp": "2023-08-22 07:57:40", "lastmod": "2023-10-20 23:41:01", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Siegel-Alex", "name": { "family": "Siegel", "given": "Alex" } }, { "id": "McAvoy-Camille-Z", "name": { "family": "McAvoy", "given": "Camille Z." } }, { "id": "Lam-Vinh-Q", "name": { "family": "Lam", "given": "Vinh" } }, { "id": "Liang-Fu-Cheng", "name": { "family": "Liang", "given": "Fu-Cheng" } }, { "id": "Kroon-Gerard-J", "name": { "family": "Kroon", "given": "Gerard" }, "orcid": "0000-0001-8038-7476" }, { "id": "Miaou-Emily", "name": { "family": "Miaou", "given": "Emily" } }, { "id": "Griffin-Patrick-R", "name": { "family": "Griffin", "given": "Patrick" }, "orcid": "0000-0002-3404-690X" }, { "id": "Wright-Peter-E", "name": { "family": "Wright", "given": "Peter E." }, "orcid": "0000-0002-1368-0223" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "A Disorder-to-Order Transition Activates an ATP-Independent Membrane Protein Chaperone", "ispublished": "pub", "full_text_status": "public", "keywords": "chaperone; membrane protein biogenesis; protein dynamics; ankyrin repeat proteins; NMR spectroscopy", "note": "\u00a9 2020 The Authors. Published by Elsevier Ltd.This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). \n\nReceived 6 August 2020, Revised 5 November 2020, Accepted 6 November 2020, Available online 12 November 2020. \n\nWe thank members of the Shan group for helpful comments on the manuscript and Dr. Peter Qin for helpful discussions in interpretation of the EPR data. This work was supported by NIH Training Grant 2T32 GM 7616-36 to Paul Sternberg, NIH R01 GM114390, NIH R35 GM136321, and DOE DE.SC0020661 to Shu-ou Shan, and the Skaggs Institute for Chemical Biology to Peter Wright. \n\nThe authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. \n\nCRediT authorship contribution statement: Alex Siegel: Conceptualization, Investigation, Formal analysis, Writing - original draft, Writing - review & editing. Camille Z. McAvoy: Conceptualization, Investigation, Formal analysis. Vinh Lam: Investigation, Formal analysis. Fu-Cheng Liang: Investigation, Formal analysis, Writing - original draft. Gerard Kroon: Methodology, Formal analysis. Emily Miaou: Investigation. Patrick Griffin: Methodology. Peter E. Wright: Supervision, Writing - review & editing. Shu-ou Shan: Supervision, Writing - review & editing.\n\nPublished - 1-s2.0-S0022283620306264-main.pdf
Accepted Version - nihms-1653549.pdf
Supplemental Material - 1-s2.0-S0022283620306264-mmc1.pdf
", "abstract": "The 43 kDa subunit of the chloroplast signal recognition particle, cpSRP43, is an ATP-independent chaperone essential for the biogenesis of the light harvesting chlorophyll-binding proteins (LHCP), the most abundant membrane protein family on earth. cpSRP43 is activated by a stromal factor, cpSRP54, to more effectively capture and solubilize LHCPs. The molecular mechanism underlying this chaperone activation is unclear. Here, a combination of hydrogen\u2013deuterium exchange, electron paramagnetic resonance, and NMR spectroscopy experiments reveal that a disorder-to-order transition of the ankyrin repeat motifs in the substrate binding domain of cpSRP43 drives its activation. An analogous coil-to-helix transition in the bridging helix, which connects the ankyrin repeat motifs to the cpSRP54 binding site in the second chromodomain, mediates long-range allosteric communication of cpSRP43 with its activating binding partner. Our results provide a molecular model to explain how the conformational dynamics of cpSRP43 enables regulation of its chaperone activity and suggest a general mechanism by which ATP-independent chaperones with cooperatively folding domains can be regulated.", "date": "2020-12-04", "date_type": "published", "publication": "Journal of Molecular Biology", "volume": "432", "number": "24", "publisher": "Elsevier", "pagerange": "Art. No. 166708", "id_number": "CaltechAUTHORS:20201113-100152867", "issn": "0022-2836", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20201113-100152867", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH Predoctoral Fellowship", "grant_number": "2T32 GM 7616-36" }, { "agency": "NIH", "grant_number": "R01 GM114390" }, { "agency": "NIH", "grant_number": "R35 GM136321" }, { "agency": "Department of Energy (DOE)", "grant_number": "DE-SC0020661" }, { "agency": "Skaggs Institute for Chemical Biology" } ] }, "doi": "10.1016/j.jmb.2020.11.007", "pmcid": "PMC7780713", "primary_object": { "basename": "1-s2.0-S0022283620306264-main.pdf", "url": "https://authors.library.caltech.edu/records/4qreq-f7w72/files/1-s2.0-S0022283620306264-main.pdf" }, "related_objects": [ { "basename": "1-s2.0-S0022283620306264-mmc1.pdf", "url": "https://authors.library.caltech.edu/records/4qreq-f7w72/files/1-s2.0-S0022283620306264-mmc1.pdf" }, { "basename": "nihms-1653549.pdf", "url": "https://authors.library.caltech.edu/records/4qreq-f7w72/files/nihms-1653549.pdf" } ], "resource_type": "article", "pub_year": "2020", "author_list": "Siegel, Alex; McAvoy, Camille Z.; et el." }, { "id": "https://authors.library.caltech.edu/records/tm8q3-wga42", "eprint_id": 106704, "eprint_status": "archive", "datestamp": "2023-08-22 07:37:50", "lastmod": "2023-10-20 23:43:00", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Hsieh-Hao-Hsuan", "name": { "family": "Hsieh", "given": "Hao-Hsuan" }, "orcid": "0000-0001-9629-5832" }, { "id": "Lee-Jae-Ho", "name": { "family": "Lee", "given": "Jae Ho" } }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "A ribosome-associated chaperone enables substrate triage in a cotranslational protein targeting complex", "ispublished": "pub", "full_text_status": "public", "keywords": "Biochemical reaction networks; Chaperones; Protein translocation; Single-molecule biophysics", "note": "\u00a9 The Author(s) 2020. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. \n\nReceived 06 May 2020; Accepted 20 October 2020; Published 17 November 2020. \n\nWe thank J. W. Chin for sharing PyltRNA/RS plasmid, D. Baltimore for sharing EMCV IRES sequence, A. Hoelz for sharing PreScission protease, R. M. Voorhees for advice on RRL in vitro translation and RNC purification, R. L. Gonzalez Jr for advice on the single-molecule TIRF setup, and members of the Shan lab for discussions and advice on this work. This work was supported by National Institutes of Health grant R01 GM078024, R35 GM136321, NSF grant MCB-1929452, and the Gordon and Betty Moore Foundation through grant GBMF2939 to S.-o.S. \n\nData availability: Data supporting the findings of this manuscript are available from the corresponding author upon reasonable request. Structural data associated with Fig. 1a are available in the Protein Data Bank under accession code 4UG0 (ribosome)78; and in Electron Microscopy Database under accession codes: EMD-3037 (SRP)6; EMD-4938 (NAC)23; EMD-6105 (RAC)79; and EMD-0202 (NatA/E)80. Source data are provided with this paper. \n\nCode availability: The scripts for modeling of cotranslational protein targeting are available at GitHub: https://github.com/emc2emc2/2020_cotranslational_targeting. \n\nAuthor Contributions: H.-H.H. and S.-o.S. conceived and designed the study. H.-H.H., J.H.L., and S.C. expressed and purified proteins and RNAs. H.-H.H. performed experiments, analyzed data, and performed simulations. H.-H.H. and S.-o.S. prepared the figures and wrote and edited the manuscript. S.-o.S. supervised the project and secured funding. \n\nCorresponding author: Correspondence to Shu-ou Shan. \n\nThe authors declare no competing interests. \n\nPeer review information: Nature Communications thanks all the anonymous reviewers for their contributions to the peer review of this work. Peer review reports are available.\n\nPublished - s41467-020-19548-5.pdf
Supplemental Material - 41467_2020_19548_MOESM1_ESM.pdf
Supplemental Material - 41467_2020_19548_MOESM2_ESM.pdf
Supplemental Material - 41467_2020_19548_MOESM3_ESM.pdf
Supplemental Material - 41467_2020_19548_MOESM4_ESM.xlsx
", "abstract": "Protein biogenesis is essential in all cells and initiates when a nascent polypeptide emerges from the ribosome exit tunnel, where multiple ribosome-associated protein biogenesis factors (RPBs) direct nascent proteins to distinct fates. How distinct RPBs spatiotemporally coordinate with one another to affect accurate protein biogenesis is an emerging question. Here, we address this question by studying the role of a cotranslational chaperone, nascent polypeptide-associated complex (NAC), in regulating substrate selection by signal recognition particle (SRP), a universally conserved protein targeting machine. We show that mammalian SRP and SRP receptors (SR) are insufficient to generate the biologically required specificity for protein targeting to the endoplasmic reticulum. NAC co-binds with and remodels the conformational landscape of SRP on the ribosome to regulate its interaction kinetics with SR, thereby reducing the nonspecific targeting of signalless ribosomes and pre-emptive targeting of ribosomes with short nascent chains. Mathematical modeling demonstrates that the NAC-induced regulations of SRP activity are essential for the fidelity of cotranslational protein targeting. Our work establishes a molecular model for how NAC acts as a triage factor to prevent protein mislocalization, and demonstrates how the macromolecular crowding of RPBs at the ribosome exit site enhances the fidelity of substrate selection into individual protein biogenesis pathways.", "date": "2020-11-17", "date_type": "published", "publication": "Nature Communications", "volume": "11", "publisher": "Nature Publishing Group", "pagerange": "Art. No. 5840", "id_number": "CaltechAUTHORS:20201117-112556236", "issn": "2041-1723", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20201117-112556236", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "R01 GM078024" }, { "agency": "NIH", "grant_number": "R35 GM136321" }, { "agency": "NSF", "grant_number": "MCB-1929452" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" } ] }, "doi": "10.1038/s41467-020-19548-5", "pmcid": "PMC7673040", "primary_object": { "basename": "41467_2020_19548_MOESM1_ESM.pdf", "url": "https://authors.library.caltech.edu/records/tm8q3-wga42/files/41467_2020_19548_MOESM1_ESM.pdf" }, "related_objects": [ { "basename": "41467_2020_19548_MOESM2_ESM.pdf", "url": "https://authors.library.caltech.edu/records/tm8q3-wga42/files/41467_2020_19548_MOESM2_ESM.pdf" }, { "basename": "41467_2020_19548_MOESM3_ESM.pdf", "url": "https://authors.library.caltech.edu/records/tm8q3-wga42/files/41467_2020_19548_MOESM3_ESM.pdf" }, { "basename": "41467_2020_19548_MOESM4_ESM.xlsx", "url": "https://authors.library.caltech.edu/records/tm8q3-wga42/files/41467_2020_19548_MOESM4_ESM.xlsx" }, { "basename": "s41467-020-19548-5.pdf", "url": "https://authors.library.caltech.edu/records/tm8q3-wga42/files/s41467-020-19548-5.pdf" } ], "resource_type": "article", "pub_year": "2020", "author_list": "Hsieh, Hao-Hsuan; Lee, Jae Ho; et el." }, { "id": "https://authors.library.caltech.edu/records/cnhtj-fr689", "eprint_id": 105969, "eprint_status": "archive", "datestamp": "2023-08-19 20:48:13", "lastmod": "2023-10-20 22:55:57", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Wang-Shuai", "name": { "family": "Wang", "given": "Shuai" }, "orcid": "0000-0002-8920-969X" }, { "id": "Yang-Chien-I", "name": { "family": "Yang", "given": "Chien-I" }, "orcid": "0000-0001-8606-5013" }, { "id": "Hsieh-Hao-Hsuan", "name": { "family": "Hsieh", "given": "Hao-Hsuan" } } ] }, "title": "Nascent Protein Selection and Triage at the Ribosome Exit Site", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2020 FASEB. \n\nIssue Online: 15 April 2020. \n\nSupport or Funding Information: R01 GM078024, R01 GM107368.", "abstract": "Proper protein biogenesis is a pre\u2010requisite for the generation and maintenance of a functional proteome. Accumulating data show that this process begins early, when nascent proteins begin to emerge from the ribosome. Indeed, the ribosome exit site is a crowded environment where a variety of ribosome\u2010associated protein biogenesis factors (RPBs) jostle for access to the nascent polypeptide. Within seconds, the nascent polypeptide must engage the correct set of factors and commit to the proper biogenesis pathway. These early decisions profoundly influence the folding, assembly, localization, maturation, and quality control of nascent proteins. This raises the question: How do these factors, which bind the ribosome at overlapping sites and prefers similar sequence features on the nascent polypeptide, compete or collaborate with one another to enable the efficient and accurate selection of nascent proteins into the proper biogenesis pathway? In this talk, I will describe recent works that begin to illustrate how the RPBs coordinate in space and time at the ribosome exit site and highlight the importance of this molecular crowding in enhancing the fidelity of individual protein biogenesis pathways.", "date": "2020-04", "date_type": "published", "publication": "FASEB Journal", "volume": "34", "number": "S1", "publisher": "Federation of American Societies for Experimental Biology", "pagerange": "1-1", "id_number": "CaltechAUTHORS:20201009-132404547", "issn": "0892-6638", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20201009-132404547", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "R01 GM078024" }, { "agency": "NIH", "grant_number": "R01 GM107368" } ] }, "doi": "10.1096/fasebj.2020.34.s1.00156", "resource_type": "article", "pub_year": "2020", "author_list": "Shan, Shu-ou; Wang, Shuai; et el." }, { "id": "https://authors.library.caltech.edu/records/4bb34-92z51", "eprint_id": 105942, "eprint_status": "archive", "datestamp": "2023-08-19 20:48:09", "lastmod": "2023-10-20 22:53:09", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Yang-Chien-I", "name": { "family": "Yang", "given": "Chien-I" }, "orcid": "0000-0001-8606-5013" }, { "id": "Hsieh-Hao-Hsuan", "name": { "family": "Hsieh", "given": "Hao-Hsuan" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Timing and Specificity of Cotranslational Nascent Protein Modification in Bacteria", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2020 FASEB. \n\nIssue Online: 17 April 2020. \n\nThis work was supported by National Institutes of Health grant GM078024 and a grant from the Weston Havens Foundation to S.\u2010o. Shan, and a Think Global Education Trust Fellowship from Taiwan to C.\u2010I. Yang.", "abstract": "Selection of newly\u2010synthesized proteins into correct protein biogenesis pathways is crucial for cellular homeostasis. The ubiquitous N\u2010terminal methionine excision (NME) process is mediated by peptide deformylase (PDF) and methionine aminopeptidase (MAP), two essential enzymes in bacteria. This reaction takes place near the nascent peptide exit site of the ribosome, where multiple ribosome\u2010associating protein biogenesis factors (RPBs) also compete for the access to the nascent chain. How NME achieves its efficiency and specificity at this crowded environment is unknown. Here, using kinetic measurements on purified ribosome\u2010nascent chain complexes, we show that the ribosome accelerates the MAP reaction for optimal substrates by 10\u00b2\u201310\u2074 folds. Kinetic competition with translation elongation and selective regulation from other RPBs enhance the specificity of NME by narrowing the processing time window for reactions on suboptimal substrates. With the kinetic data, we constructed a mathematical model and accurately predicted the cotranslational NME efficiency in cytosol. Our data demonstrate how a fundamental enzymatic activity is reshaped by its associated macromolecular environment to optimize both efficiency and selectivity. Moreover, the remodeled MAP activity prompted us to develop a cotranslational assay to screen for MAP inhibitors in the physiological context of translating ribosome. The results explain the discrepancy between traditional peptide\u2010based assays and cellular data, providing a powerful tool for the development of antibacterial agents targeting the NME machineries.", "date": "2020-04", "date_type": "published", "publication": "FASEB Journal", "volume": "34", "number": "S1", "publisher": "Federation of American Societies for Experimental Biology", "pagerange": "1", "id_number": "CaltechAUTHORS:20201008-143954486", "issn": "0892-6638", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20201008-143954486", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Weston Havens Foundation" }, { "agency": "Think Global Education Trust Fellowship" } ] }, "doi": "10.1096/fasebj.2020.34.s1.04322", "resource_type": "article", "pub_year": "2020", "author_list": "Yang, Chien-I; Hsieh, Hao-Hsuan; et el." }, { "id": "https://authors.library.caltech.edu/records/nc05b-qq381", "eprint_id": 99591, "eprint_status": "archive", "datestamp": "2023-08-19 18:42:52", "lastmod": "2023-10-18 18:34:48", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Yang-Chien-I", "name": { "family": "Yang", "given": "Chien-I" }, "orcid": "0000-0001-8606-5013" }, { "id": "Hsieh-Hao-Hsuan", "name": { "family": "Hsieh", "given": "Hao-Hsuan" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Timing and specificity of cotranslational nascent protein modification in bacteria", "ispublished": "pub", "full_text_status": "public", "keywords": "nascent protein modification | ribosome | protein biogenesis | peptide deformylase | methionine aminopeptidase", "note": "\u00a9 2019. Published under the PNAS license. \n\nEdited by Gisela Storz, National Institute of Child Health and Human Development, Bethesda, MD, and approved October 6, 2019 (received for review July 16, 2019) \n\nWe thank T. F. Miller, M. H. Zimmer, and Y. L. Ni for critical advice on the simulation and T. F. Miller, A. Varshavsky, and members of the S.-o.S. laboratory for discussions and comments on the manuscript. This work was supported by National Institutes of Health Grant GM078024 and a grant from the Weston Havens Foundation (to S.-o.S.) and a Think Global Education Trust Fellowship from Taiwan (to C.-I.Y.). \n\nData Availability. Materials are available on request. \n\nAuthor contributions: C.-I.Y. and S.-o.S. designed research; C.-I.Y. performed research; H.-H.H. contributed new reagents/analytic tools; C.-I.Y. and S.-o.S. analyzed data; and C.-I.Y. and S.-o.S. wrote the paper. \n\nThe authors declare no competing interest. \n\nThis article is a PNAS Direct Submission. \n\nData deposition: All data discussed in the paper will be made available to readers. \n\nThis article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1912264116/-/DCSupplemental.\n\nPublished - 23050.full.pdf
Supplemental Material - pnas.1912264116.sapp.pdf
", "abstract": "The nascent polypeptide exit site of the ribosome is a crowded environment where multiple ribosome-associated protein biogenesis factors (RPBs) compete for the nascent polypeptide to influence their localization, folding, or quality control. Here we address how N-terminal methionine excision (NME), a ubiquitous process crucial for the maturation of over 50% of the bacterial proteome, occurs in a timely and selective manner in this crowded environment. In bacteria, NME is mediated by 2 essential enzymes, peptide deformylase (PDF) and methionine aminopeptidase (MAP). We show that the reaction of MAP on ribosome-bound nascent chains approaches diffusion-limited rates, allowing immediate methionine excision of optimal substrates after deformylation. Specificity is achieved by kinetic competition of NME with translation elongation and by regulation from other RPBs, which selectively narrow the processing time window for suboptimal substrates. A mathematical model derived from the data accurately predicts cotranslational NME efficiency in the cytosol. Our results demonstrate how a fundamental enzymatic activity is reshaped by its associated macromolecular environment to optimize both efficiency and selectivity, and provides a platform to study other cotranslational protein biogenesis pathways.", "date": "2019-11-12", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "116", "number": "46", "publisher": "National Academy of Sciences", "pagerange": "23050-23060", "id_number": "CaltechAUTHORS:20191031-125530361", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20191031-125530361", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "R01 GM078024" }, { "agency": "Weston Havens Foundation" }, { "agency": "Think Global Education Trust" } ] }, "doi": "10.1073/pnas.1912264116", "pmcid": "PMC6859321", "primary_object": { "basename": "pnas.1912264116.sapp.pdf", "url": "https://authors.library.caltech.edu/records/nc05b-qq381/files/pnas.1912264116.sapp.pdf" }, "related_objects": [ { "basename": "23050.full.pdf", "url": "https://authors.library.caltech.edu/records/nc05b-qq381/files/23050.full.pdf" } ], "resource_type": "article", "pub_year": "2019", "author_list": "Yang, Chien-I; Hsieh, Hao-Hsuan; et el." }, { "id": "https://authors.library.caltech.edu/records/7e067-2g554", "eprint_id": 99013, "eprint_status": "archive", "datestamp": "2023-08-22 02:58:31", "lastmod": "2023-10-18 17:47:34", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Guiding tail-anchored membrane proteins to the endoplasmic reticulum in a chaperone cascade", "ispublished": "pub", "full_text_status": "public", "keywords": "chaperone; membrane protein; protein targeting; 70 kilodalton heat shock protein (Hsp70); ATPase; tail-anchored protein", "note": "\u00a9 2019 Shan. Published under exclusive license by The American Society for Biochemistry and Molecular Biology, Inc. \n\nFirst Published on October 1, 2019. \n\nThis work was supported by National Institutes of Health Grant GM107368, Gordon and Betty Moore Foundation Grant GBMF2939, and a fellowship from the Weston Havens Foundation (to S.-o. S.). The author declares that he has no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. \n\nI thank H. Cho and Y.lM. Liu for critical discussions and comments on the manuscript.\n\nPublished - J._Biol._Chem.-2019-Shan-16577-86.pdf
", "abstract": "Newly synthesized integral membrane proteins must traverse the aqueous cytosolic environment before arrival at their membrane destination and are prone to aggregation, misfolding, and mislocalization during this process. The biogenesis of integral membrane proteins therefore poses acute challenges to protein homeostasis within a cell and requires the action of effective molecular chaperones. Chaperones that mediate membrane protein targeting not only need to protect the nascent transmembrane domains from improper exposure in the cytosol, but also need to accurately select client proteins and actively guide their clients to the appropriate target membrane. The mechanisms by which cellular chaperones work together to coordinate this complex process are only beginning to be delineated. Here, we summarize recent advances in studies of the tail-anchored membrane protein targeting pathway, which revealed a network of chaperones, cochaperones, and targeting factors that together drive and regulate this essential process. This pathway is emerging as an excellent model system to decipher the mechanism by which molecular chaperones overcome the multiple challenges during post-translational membrane protein biogenesis and to gain insights into the functional organization of multicomponent chaperone networks.", "date": "2019-11-08", "date_type": "published", "publication": "Journal of Biological Chemistry", "volume": "294", "number": "45", "publisher": "American Society for Biochemistry and Molecular Biology", "pagerange": "16577-16586", "id_number": "CaltechAUTHORS:20191002-094951031", "issn": "0021-9258", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20191002-094951031", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM107368" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" }, { "agency": "Weston Havens Foundation" } ] }, "doi": "10.1074/jbc.rev119.006197", "pmcid": "PMC6851334", "primary_object": { "basename": "J._Biol._Chem.-2019-Shan-16577-86.pdf", "url": "https://authors.library.caltech.edu/records/7e067-2g554/files/J._Biol._Chem.-2019-Shan-16577-86.pdf" }, "resource_type": "article", "pub_year": "2019", "author_list": "Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/123jh-haf84", "eprint_id": 98761, "eprint_status": "archive", "datestamp": "2023-08-19 18:13:49", "lastmod": "2023-10-18 17:37:35", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Hwang-Fu-Yu-Hsien", "name": { "family": "Hwang Fu", "given": "Yu-Hsien" }, "orcid": "0000-0002-2861-4843" }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Lee-Jae-Ho", "name": { "family": "Lee", "given": "Jae Ho" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "A molecular recognition feature mediates ribosome-induced SRP-receptor assembly during protein targeting", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2019 Hwang Fu et al. This article is distributed under the terms of an Attribution\u2013Noncommercial\u2013Share Alike\u2013No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms/). After six months it is available under a Creative Commons License (Attribution\u2013Noncommercial\u2013Share Alike 4.0 International license, as described at https://creativecommons.org/licenses/by-nc-sa/4.0/). \n\nSubmitted: January 1, 2019. Revision received June 28, 2019. Accepted: August 26, 2019. Published September 19, 2019. \n\nWe thank the Shan laboratory members for valuable suggestions and H. Bernstein for sharing canine pancreatic microsomes. \n\nThis work was supported by National Institutes of Health grant GM078024 and Gordon and Betty Moore Foundation GBMF2939 to S.-o. Shan. \n\nThe authors declare no competing financial interests. \n\nAuthor contributions: Y.-H. Hwang Fu and S.-o. Shan designed research; Y.-H. Hwang Fu, S. Chandrasekar, and S.-o. Shan performed research; Y.-H. Hwang Fu, S. Chandrasekar, and J.H. Lee contributed new reagents/analytic tools; Y.-H. Hwang Fu, S. Chandrasekar, and S.-o. Shan analyzed data; Y.-H. Huang Fu and S.-o. Shan wrote the paper; S.-o. Shan supervised the project.\n\nPublished - 3307.full.pdf
Supplemental Material - JCB_201901001_sm.pdf
", "abstract": "Molecular recognition features (MoRFs) provide interaction motifs in intrinsically disordered protein regions to mediate diverse cellular functions. Here we report that a MoRF element, located in the disordered linker domain of the mammalian signal recognition particle (SRP) receptor and conserved among eukaryotes, plays an essential role in sensing the ribosome during cotranslational protein targeting to the endoplasmic reticulum. Loss of the MoRF in the SRP receptor (SR) largely abolishes the ability of the ribosome to activate SRP-SR assembly and impairs cotranslational protein targeting. These results demonstrate a novel role for MoRF elements and provide a mechanism for the ribosome-induced activation of the mammalian SRP pathway. Kinetic analyses and comparison with the bacterial SRP further suggest that the SR MoRF functionally replaces the essential GNRA tetraloop in the bacterial SRP RNA, providing an example for the replacement of RNA function by proteins during the evolution of ancient ribonucleoprotein particles.", "date": "2019-10-07", "date_type": "published", "publication": "Journal of Cell Biology", "volume": "218", "number": "10", "publisher": "Rockefeller University Press", "pagerange": "3307-3319", "id_number": "CaltechAUTHORS:20190919-142731254", "issn": "0021-9525", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190919-142731254", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" } ] }, "doi": "10.1083/jcb.201901001", "pmcid": "PMC6781444", "primary_object": { "basename": "3307.full.pdf", "url": "https://authors.library.caltech.edu/records/123jh-haf84/files/3307.full.pdf" }, "related_objects": [ { "basename": "JCB_201901001_sm.pdf", "url": "https://authors.library.caltech.edu/records/123jh-haf84/files/JCB_201901001_sm.pdf" } ], "resource_type": "article", "pub_year": "2019", "author_list": "Hwang Fu, Yu-Hsien; Chandrasekar, Sowmya; et el." }, { "id": "https://authors.library.caltech.edu/records/a6rt4-1wb44", "eprint_id": 97591, "eprint_status": "archive", "datestamp": "2023-08-22 02:35:31", "lastmod": "2023-10-18 16:07:08", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Wang-Shuai", "name": { "family": "Wang", "given": "Shuai" }, "orcid": "0000-0002-8920-969X" }, { "id": "Jomaa-A", "name": { "family": "Jomaa", "given": "Ahmad" }, "orcid": "0000-0002-4786-934X" }, { "id": "Jaskolowski-M", "name": { "family": "Jaskolowski", "given": "Mateusz" }, "orcid": "0000-0001-6618-7048" }, { "id": "Yang-Chien-I", "name": { "family": "Yang", "given": "Chien-I" }, "orcid": "0000-0001-8606-5013" }, { "id": "Ban-Nenad", "name": { "family": "Ban", "given": "Nenad" }, "orcid": "0000-0002-9527-210X" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "The molecular mechanism of cotranslational membrane protein recognition and targeting by SecA", "ispublished": "pub", "full_text_status": "public", "keywords": "Biochemistry; Cell biology; Structural biology", "note": "\u00a9 2019 Springer Nature Limited. \n\nReceived 04 May 2019; Accepted 12 August 2019; Published 30 September 2019. \n\nData Availability: Cryo-EM maps are deposited in the electron microscopy databank (EMDB) with accession codes EMD-10073 (RNCRodZ\u2013SecA) and EMD-10074 (SecA, local refinement), and model coordinates are deposited in the worldwide PDB with accession code PDB 6S0K. Other data are available from corresponding authors upon reasonable request. \n\nWe thank A. McDowall and H. Wang for assistance with negative stain electron microscopy data collection, J. Rothman for sharing plasmid ApoE422k, D. Boehringer and A. Scaiola for the support with EM data collection and processing, M. Leibundgut and M. Saurer for the support with model building and members of the Shan and Ban groups for discussions and comments on the manuscript. We also thank T. Miller, M. Zimmer and F. Huber for helpful discussions. Cryo-EM data was collected at the Scientific Center for Optical and Electron Microscopy at the ETH Zurich (ScopeM). We gratefully acknowledge the support of NVIDIA Corporation for the Titan Xp GPU used in this research through a GPU Grant program awarded to A.J.; M.J. was supported by the internal research grant of the ETH to N.B. (ETH-40 16-2). This work was supported by National Institutes of Health grant GM107368A and the Gordon and Betty Moore Foundation through grant GBMF2939 to S.S. and by the Swiss National Science Foundation (SNSF) (grant number 310030B_163478), National Center of Excellence in Research (NCCR) RNA & Disease Program of the SNSF (grant number 51NF40_141735) to N.B. \n\nAuthor Contributions: S.W. and S.S. conceived the project. S.W. performed most of the biochemical experiments and analyzed data. A.J. acquired cryo-electron microscopy data and performed reconstructions and model building. M.J. performed initial sample preparation for cryo-EM analysis. C.Y. measured the association rate constant of SecA binding to RNCRodZ. N.B. and S.S. supervised the structural and biochemical experiments, respectively. All authors interpreted the data and contributed to the final versions of the manuscript. \n\nThe authors declare no competing interests.\n\nAccepted Version - nihms-1537248.pdf
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Supplemental Material - 41594_2019_297_MOESM1_ESM.pdf
Supplemental Material - 41594_2019_297_MOESM2_ESM.pdf
", "abstract": "Cotranslational protein targeting is a conserved process for membrane protein biogenesis. In Escherichia coli, the essential ATPase SecA was found to cotranslationally target a subset of nascent membrane proteins to the SecYEG translocase at the plasma membrane. The molecular mechanism of this pathway remains unclear. Here we use biochemical and cryoelectron microscopy analyses to show that the amino-terminal amphipathic helix of SecA and the ribosomal protein uL23 form a composite binding site for the transmembrane domain (TMD) on the nascent protein. This binding mode further enables recognition of charged residues flanking the nascent TMD and thus explains the specificity of SecA recognition. Finally, we show that membrane-embedded SecYEG promotes handover of the translating ribosome from SecA to the translocase via a concerted mechanism. Our work provides a molecular description of the SecA-mediated cotranslational targeting pathway and demonstrates an unprecedented role of the ribosome in shielding nascent TMDs.", "date": "2019-10", "date_type": "published", "publication": "Nature Structural & Molecular Biology", "volume": "26", "number": "10", "publisher": "Nature Publishing Group", "pagerange": "919-929", "id_number": "CaltechAUTHORS:20190801-131335661", "issn": "1545-9985", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190801-131335661", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NVIDIA Corporation" }, { "agency": "ETH Zurich", "grant_number": "ETH-40 16-2" }, { "agency": "NIH", "grant_number": "GM107368A" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" }, { "agency": "Swiss National Science Foundation (SNSF)", "grant_number": "310030B_163478" }, { "agency": "Swiss National Science Foundation (SNSF)", "grant_number": "51NF40_141735" } ] }, "doi": "10.1038/s41594-019-0297-8", "pmcid": "PMC6858539", "primary_object": { "basename": "41594_2019_297_Fig12_ESM.jpg", "url": "https://authors.library.caltech.edu/records/a6rt4-1wb44/files/41594_2019_297_Fig12_ESM.jpg" }, "related_objects": [ { "basename": "41594_2019_297_Fig14_ESM.jpg", "url": "https://authors.library.caltech.edu/records/a6rt4-1wb44/files/41594_2019_297_Fig14_ESM.jpg" }, { "basename": "41594_2019_297_Fig15_ESM.jpg", "url": "https://authors.library.caltech.edu/records/a6rt4-1wb44/files/41594_2019_297_Fig15_ESM.jpg" }, { "basename": "41594_2019_297_Fig8_ESM.jpg", "url": "https://authors.library.caltech.edu/records/a6rt4-1wb44/files/41594_2019_297_Fig8_ESM.jpg" }, { "basename": "41594_2019_297_MOESM2_ESM.pdf", "url": "https://authors.library.caltech.edu/records/a6rt4-1wb44/files/41594_2019_297_MOESM2_ESM.pdf" }, { "basename": "41594_2019_297_Fig10_ESM.jpg", "url": "https://authors.library.caltech.edu/records/a6rt4-1wb44/files/41594_2019_297_Fig10_ESM.jpg" }, { "basename": "41594_2019_297_Fig11_ESM.jpg", "url": "https://authors.library.caltech.edu/records/a6rt4-1wb44/files/41594_2019_297_Fig11_ESM.jpg" }, { "basename": "41594_2019_297_Fig13_ESM.jpg", "url": "https://authors.library.caltech.edu/records/a6rt4-1wb44/files/41594_2019_297_Fig13_ESM.jpg" }, { "basename": "41594_2019_297_Fig9_ESM.jpg", "url": "https://authors.library.caltech.edu/records/a6rt4-1wb44/files/41594_2019_297_Fig9_ESM.jpg" }, { "basename": "41594_2019_297_MOESM1_ESM.pdf", "url": "https://authors.library.caltech.edu/records/a6rt4-1wb44/files/41594_2019_297_MOESM1_ESM.pdf" }, { "basename": "nihms-1537248.pdf", "url": "https://authors.library.caltech.edu/records/a6rt4-1wb44/files/nihms-1537248.pdf" } ], "resource_type": "article", "pub_year": "2019", "author_list": "Wang, Shuai; Jomaa, Ahmad; et el." }, { "id": "https://authors.library.caltech.edu/records/g7913-0z550", "eprint_id": 91991, "eprint_status": "archive", "datestamp": "2023-08-22 00:40:12", "lastmod": "2023-10-19 23:52:58", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Chio-Un-Seng", "name": { "family": "Chio", "given": "Un Seng" }, "orcid": "0000-0002-5295-2690" }, { "id": "Chung-SangYoon", "name": { "family": "Chung", "given": "SangYoon" }, "orcid": "0000-0002-0592-4099" }, { "id": "Weiss-Shimon", "name": { "family": "Weiss", "given": "Shimon" }, "orcid": "0000-0002-0720-5426" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "A Chaperone Lid Ensures Efficient and Privileged Client Transfer during Tail-Anchored Protein Targeting", "ispublished": "pub", "full_text_status": "public", "keywords": "chaperone; protein targeting; tail-anchored protein; ATPase; membrane protein", "note": "\u00a9 2018 The Authors. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). \n\nReceived 23 June 2018, Revised 20 September 2018, Accepted 7 December 2018, Available online 2 January 2019. \n\nWe thank J. Chartron for advice on generating yeast strains using CRISPR-Cas9; R.S. Hegde for CaM expression vectors; H. Cho, M. Rao, F. Liang, A. Siegel, and S. Wang for reagents; S. Hematian and the Beckman Institute Laser Resource Center for training and use of circular dichroism spectrometer; M. Rao for initial observations; and members of the Shan laboratory for critical discussions and comments on the manuscript. This work was supported by Dean Willard Chair funds to S.W., NIH grant GM107368, Gordon and Betty Moore Foundation grant GBMF2939, and a fellowship from the Weston Havens Foundation to S.S. \n\nAuthor Contributions: U.S.C. and S.S. designed the research. U.S.C. performed biochemical experiments. U.S.C. analyzed biochemical data with input from S.S. U.S.C. and S.C. performed single-molecule fluorescence experiments. U.S.C. and S.C. analyzed the data with input from S.W. and S.S. U.S.C and S.S. wrote the manuscript. All authors approved the final manuscript. \n\nThe authors declare no competing interests.\n\nPublished - 1-s2.0-S221112471831965X-main.pdf
Accepted Version - nihms-1023701.pdf
Supplemental Material - 1-s2.0-S221112471831965X-mmc1.pdf
", "abstract": "Molecular chaperones play key roles in maintaining cellular proteostasis. In addition to preventing client aggregation, chaperones often relay substrates within a network while preventing off-pathway chaperones from accessing the substrate. Here we show that a conserved lid motif lining the substrate-binding groove of the Get3 ATPase enables these important functions during the targeted delivery of tail-anchored membrane proteins (TAs) to the endoplasmic reticulum. The lid prevents promiscuous TA handoff to off-pathway chaperones, and more importantly, it cooperates with the Get4/5 scaffolding complex to enable rapid and privileged TA transfer from the upstream co-chaperone Sgt2 to Get3. These findings provide a molecular mechanism by which chaperones maintain the pathway specificity of client proteins in the crowded cytosolic environment.", "date": "2019-01-02", "date_type": "published", "publication": "Cell Reports", "volume": "26", "number": "1", "publisher": "Cell Press", "pagerange": "37-44", "id_number": "CaltechAUTHORS:20190102-135256811", "issn": "2211-1247", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190102-135256811", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Dean Willard Chair" }, { "agency": "NIH", "grant_number": "GM107368" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" }, { "agency": "Weston Havens Foundation" } ] }, "doi": "10.1016/j.celrep.2018.12.035", "pmcid": "PMC6689467", "primary_object": { "basename": "1-s2.0-S221112471831965X-main.pdf", "url": "https://authors.library.caltech.edu/records/g7913-0z550/files/1-s2.0-S221112471831965X-main.pdf" }, "related_objects": [ { "basename": "1-s2.0-S221112471831965X-mmc1.pdf", "url": "https://authors.library.caltech.edu/records/g7913-0z550/files/1-s2.0-S221112471831965X-mmc1.pdf" }, { "basename": "nihms-1023701.pdf", "url": "https://authors.library.caltech.edu/records/g7913-0z550/files/nihms-1023701.pdf" } ], "resource_type": "article", "pub_year": "2019", "author_list": "Chio, Un Seng; Chung, SangYoon; et el." }, { "id": "https://authors.library.caltech.edu/records/bxgje-prz14", "eprint_id": 90148, "eprint_status": "archive", "datestamp": "2023-08-22 00:23:49", "lastmod": "2023-10-18 23:14:27", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Cho-Hyunju", "name": { "family": "Cho", "given": "Hyunju" }, "orcid": "0000-0001-7393-657X" }, { "id": "Chio-Un-Seng", "name": { "family": "Chio", "given": "Un Seng" }, "orcid": "0000-0002-5295-2690" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "In vitro Assays for Targeting and Insertion of Tail\u2010Anchored Proteins Into the ER Membrane", "ispublished": "pub", "full_text_status": "public", "keywords": "amber suppression; chaperones; fluorescence; membrane proteins; protein targeting; tail\u2010anchored proteins", "note": "\u00a9 2018 by John Wiley & Sons, Inc. \n\nVersion of Record online: 25 September 2018. \n\nWe thank members of the Shan lab for comments on reagent preparation. This work was supported by NIH grant GM107368, Gordon and Betty Moore Foundation Grant GBMF2939, and a fellowship from the Weston Havens Foundation to S.-o.S.\n\nAccepted Version - nihms970064.pdf
", "abstract": "Membrane proteins mediate numerous essential cellular functions. Due to the aggregation propensity of hydrophobic transmembrane domains in aqueous environments, the targeting and insertion of membrane proteins pose major challenges to cells. In the Guided Entry of Tail\u2010anchored protein (GET) pathway, an essential class of newly synthesized tail\u2010anchored proteins (TAs) are chaperoned and guided by multiple targeting factors to the endoplasmic reticulum (ER) membrane. Deciphering the molecular mechanism of this cellular process has benefitted from successful in vitro reconstitution of individual molecular events in the GET pathway with purified components. Here we describe recently developed protocols for in vitro reconstitution of functional complexes of TA substrates with their targeting factors, for monitoring the transfer of TAs between targeting factors, and for the insertion of TA into the microsomal membrane. These procedures are generally applicable to the interrogation of other post\u2010translational membrane protein targeting pathways.", "date": "2018-12", "date_type": "published", "publication": "Current Protocols in Cell Biology", "volume": "81", "number": "1", "publisher": "Wiley", "pagerange": "Art. No. e63", "id_number": "CaltechAUTHORS:20181008-095135080", "issn": "1934-2616", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20181008-095135080", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM107368" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" }, { "agency": "Weston Havens Foundation" } ] }, "doi": "10.1002/cpcb.63", "pmcid": "PMC6263799", "primary_object": { "basename": "nihms970064.pdf", "url": "https://authors.library.caltech.edu/records/bxgje-prz14/files/nihms970064.pdf" }, "resource_type": "article", "pub_year": "2018", "author_list": "Cho, Hyunju; Chio, Un Seng; et el." }, { "id": "https://authors.library.caltech.edu/records/9cyf6-z8w66", "eprint_id": 87649, "eprint_status": "archive", "datestamp": "2023-08-19 10:11:59", "lastmod": "2023-10-18 21:20:23", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Cho-Hyunju", "name": { "family": "Cho", "given": "Hyunju" }, "orcid": "0000-0001-7393-657X" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Substrate relay in an Hsp70\u2010cochaperone cascade safeguards tail\u2010anchored membrane protein targeting", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2018 The Authors. \n\nReceived 16 February 2018; Revised 24 May 2018; Accepted 29 May 2018. \n\nWe thank E. A. Craig for the Ssa antibody and the SSA1 and ssa1ts strains, V. Denic for the SGT2FLAG\u2206get3 and sgt2TPRmtFLAG strains, U. S. Chio, M. Rao, A. Montequin, F.C. Liang, A. Siegel for reagents, and the Dougherty lab for use of HPLC. We thank Bil Clemons, Ray Deshaies, and members of the Shan, Clemons, and Deshaies laboratories for discussions and critical comments on the manuscript. This work was supported by NIH grant GM107368 and grant from the Weston Havens Foundation to S.S. \n\nAuthor contributions: HC and SS designed experiment; HC performed experiments and analyzed the data; HC and SS wrote the paper. \n\nThe authors declare that they have no conflict of interest.\n\nSupplemental Material - inline-supplementary-material-4.pdf
Supplemental Material - inline-supplementary-material-5.pdf
Supplemental Material - inline-supplementary-material-6.zip
", "abstract": "Membrane proteins are aggregation\u2010prone in aqueous environments, and their biogenesis poses acute challenges to cellular protein homeostasis. How the chaperone network effectively protects integral membrane proteins during their post\u2010translational targeting is not well understood. Here, biochemical reconstitutions showed that the yeast cytosolic Hsp70 is responsible for capturing newly synthesized tail\u2010anchored membrane proteins (TAs) in the soluble form. Moreover, direct interaction of Hsp70 with the cochaperone Sgt2 initiates a sequential series of TA relays to the dedicated TA targeting factor Get3. In contrast to direct loading of TAs to downstream chaperones, stepwise substrate loading via Hsp70 maintains the solubility and targeting competence of TAs, ensuring their efficient delivery to the endoplasmic reticulum (ER). Inactivation of cytosolic Hsp70 severely impairs TA translocation in vivo. Our results demonstrate a new role of cytosolic Hsp70 in directly assisting the targeting of an essential class of integral membrane proteins and provide a paradigm for how \"substrate funneling\" through a chaperone cascade preserves the conformational quality of nascent membrane proteins during their biogenesis.", "date": "2018-07-02", "date_type": "published", "publication": "EMBO Journal", "volume": "37", "number": "16", "publisher": "European Molecular Biology Organization", "pagerange": "Art. No. e99264", "id_number": "CaltechAUTHORS:20180709-130553195", "issn": "0261-4189", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20180709-130553195", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM107368" }, { "agency": "Weston Havens Foundation" } ] }, "doi": "10.15252/embj.201899264", "pmcid": "PMC6092619", "primary_object": { "basename": "inline-supplementary-material-4.pdf", "url": "https://authors.library.caltech.edu/records/9cyf6-z8w66/files/inline-supplementary-material-4.pdf" }, "related_objects": [ { "basename": "inline-supplementary-material-5.pdf", "url": "https://authors.library.caltech.edu/records/9cyf6-z8w66/files/inline-supplementary-material-5.pdf" }, { "basename": "inline-supplementary-material-6.zip", "url": "https://authors.library.caltech.edu/records/9cyf6-z8w66/files/inline-supplementary-material-6.zip" } ], "resource_type": "article", "pub_year": "2018", "author_list": "Cho, Hyunju and Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/5c4qm-vq361", "eprint_id": 86725, "eprint_status": "archive", "datestamp": "2023-08-21 23:32:10", "lastmod": "2023-10-18 19:56:11", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Lee-Jae-Ho", "name": { "family": "Lee", "given": "Jae Ho" }, "orcid": "0000-0002-8663-3209" }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Chung-SangYoon", "name": { "family": "Chung", "given": "SangYoon" }, "orcid": "0000-0002-0592-4099" }, { "id": "Fu-Yu-Hsien-Hwang", "name": { "family": "Fu", "given": "Yu-Hsien Hwang" } }, { "id": "Liu-Demi", "name": { "family": "Liu", "given": "Demi" } }, { "id": "Weiss-Shimon", "name": { "family": "Weiss", "given": "Shimon" }, "orcid": "0000-0002-0720-5426" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Sequential activation of human signal recognition particle by the ribosome and signal sequence drives efficient protein targeting", "ispublished": "pub", "full_text_status": "public", "keywords": "protein targeting; signal recognition particle; ribosome; fluorescence spectroscopy; single-molecule spectroscopy", "note": "\u00a9 2018 National Academy of Sciences. Published under the PNAS license. \n\nEdited by Joseph D. Puglisi, Stanford University School of Medicine, Stanford, CA, and approved May 2, 2018 (received for review February 6, 2018). Published ahead of print May 30, 2018. \n\nWe thank E. Menichelli, K. Nagai, E. Mandon, R. Gilmore, K. Strub, and C. Zwieb for the expression constructs and purification protocols on SRP proteins and SRP RNA; A. Sharma for advice on RRL reagents and protocols; H. Bernstein for sharing canine pancreatic microsomes; the laboratory of D. Rees for the use of MST; the laboratory of D. Dougherty for the use of HPLC; and K. Strub for advice on SRP assembly and purification procedures. This work was supported by National Institutes of Health Grant GM078024, Gordon and Betty Moore Foundation Grant GBMF2939 (to S.-o.S.), and Dean Willard Chair funds (to S.W.). \n\nAuthor contributions: J.H.L., S.W., and S.-o.S. designed research; J.H.L., S. Chandrasekar, S. Chung, Y.-H.H.F., D.L., and S.-o.S. performed research; J.H.L., S. Chandrasekar, S. Chung, Y.-H.H.F., D.L., and S.W. contributed new reagents/analytic tools; J.H.L., S. Chandrasekar, S. Chung, Y.-H.H.F., D.L., S.W., and S.-o.S. analyzed data; J.H.L. and S.-o.S. wrote the paper; S.W. led supervision on single molecule spectroscopy experiments; and S.-o.S. supervised the project. \n\nThe authors declare no conflict of interest. \n\nThis article is a PNAS Direct Submission. \n\nThis article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1802252115/-/DCSupplemental.\n\nPublished - E5487.full.pdf
Supplemental Material - pnas.1802252115.sapp.pdf
", "abstract": "Signal recognition particle (SRP) is a universally conserved targeting machine that mediates the targeted delivery of \u223c30% of the proteome. The molecular mechanism by which eukaryotic SRP achieves efficient and selective protein targeting remains elusive. Here, we describe quantitative analyses of completely reconstituted human SRP (hSRP) and SRP receptor (SR). Enzymatic and fluorescence analyses showed that the ribosome, together with a functional signal sequence on the nascent polypeptide, are required to activate SRP for rapid recruitment of the SR, thereby delivering translating ribosomes to the endoplasmic reticulum. Single-molecule fluorescence spectroscopy combined with cross-complementation analyses reveal a sequential mechanism of activation whereby the ribosome unlocks the hSRP from an autoinhibited state and primes SRP to sample a variety of conformations. The signal sequence further preorganizes the mammalian SRP into the optimal conformation for efficient recruitment of the SR. Finally, the use of a signal sequence to activate SRP for receptor recruitment is a universally conserved feature to enable efficient and selective protein targeting, and the eukaryote-specific components confer upon the mammalian SRP the ability to sense and respond to ribosomes.", "date": "2018-06-12", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "115", "number": "24", "publisher": "National Academy of Sciences", "pagerange": "E5487-E5496", "id_number": "CaltechAUTHORS:20180530-150151067", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20180530-150151067", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" }, { "agency": "Dean Willard Chair" } ] }, "doi": "10.1073/pnas.1802252115", "pmcid": "PMC6004459", "primary_object": { "basename": "pnas.1802252115.sapp.pdf", "url": "https://authors.library.caltech.edu/records/5c4qm-vq361/files/pnas.1802252115.sapp.pdf" }, "related_objects": [ { "basename": "E5487.full.pdf", "url": "https://authors.library.caltech.edu/records/5c4qm-vq361/files/E5487.full.pdf" } ], "resource_type": "article", "pub_year": "2018", "author_list": "Lee, Jae Ho; Chandrasekar, Sowmya; et el." }, { "id": "https://authors.library.caltech.edu/records/2f7mb-jzr72", "eprint_id": 86002, "eprint_status": "archive", "datestamp": "2023-08-19 09:44:27", "lastmod": "2023-10-18 19:10:25", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "McAvoy-Camille-Z", "name": { "family": "McAvoy", "given": "Camille" } }, { "id": "Siegel-Alex", "name": { "family": "Siegel", "given": "Alex" }, "orcid": "0000-0003-3601-5178" }, { "id": "Piszkiewicz-Samantha", "name": { "family": "Piszkiewicz", "given": "Samantha" } }, { "id": "Miaou-Emily", "name": { "family": "Miaou", "given": "Emily" } }, { "id": "Yu-Mansen", "name": { "family": "Yu", "given": "Mansen" } }, { "id": "Nguyen-Thang-X", "name": { "family": "Nguyen", "given": "Thang" } }, { "id": "Moradian-Annie", "name": { "family": "Moradian", "given": "Annie" }, "orcid": "0000-0002-0407-2031" }, { "id": "Sweredoski-Michael-J", "name": { "family": "Sweredoski", "given": "Michael J." }, "orcid": "0000-0003-0878-3831" }, { "id": "Hess-Sonja", "name": { "family": "Hess", "given": "Sonja" }, "orcid": "0000-0002-5904-9816" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Two Distinct Sites of client protein interaction with the chaperone cpSRP43", "ispublished": "pub", "full_text_status": "public", "keywords": "Signal Recognition Particle, protein targeting, chloroplast, chaperones, membrane proteins", "note": "\u00a9 2018 Published under license by The American Society for Biochemistry and Molecular Biology, Inc. \n\nReceived for publication, February 3, 2018, and in revised form, March 29, 2018 Published, Papers in Press, April 18, 2018. \n\nThis work was supported by Betty and Gordon Moore Foundation Grant 94-3397785 and National Institutes of Health Grant R01 GM114390 (to S.-o. S.) and by National Institutes of Health Training Grant 2 T32 GM 7616-36 (to C. Z. M.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. \n\nWe thank members of the Shan group for helpful comments on the manuscript. The Proteome Exploration Laboratory was supported by Betty and Gordon Moore Foundation Grant GBMF775 and the Beckman Institute at Caltech. \n\nAuthor contributions\u2014C. Z. M., A. S., S. P., E. M., T. N., A. M., M. J. S., and S. H. data curation; C. Z. M., A. S., S. P., E. M., M. Y., T. N., A. M., S. H., and S.-o. S. formal analysis; C. Z. M., A. S., S. P., E. M., M. Y., T. N., and S. H. investigation; C. M. writing-original\ndraft; A. S. and S.-o. S. validation; A. S., S. H., and S.-o. S. writing-review and editing; M. J. S. software; S. H. and S.-o. S. supervision; S.-o. S. conceptualization; S.-o. S. funding acquisition; S.-o. S. project administration.\n\nPublished - J._Biol._Chem.-2018-McAvoy-8861-73.pdf
Accepted Version - J._Biol._Chem.-2018-McAvoy-jbc.RA118.002215.pdf
Supplemental Material - 135723_1_supp_107935_p6dqkd.tif
", "abstract": "Integral membrane proteins are prone to aggregation and misfolding in aqueous environments and therefore require binding by molecular chaperones during their biogenesis. Chloroplast signal recognition particle 43 (cpSRP43) is an ATP-independent chaperone required for the biogenesis of the most abundant class of membrane proteins, the light-harvesting chlorophyll a/b-binding proteins (LHCPs). Previous work has shown that cpSRP43 specifically recognizes an L18 loop sequence conserved among LHCP paralogs. However, how cpSRP43 protects the transmembrane domains (TMDs) of LHCP from aggregation was unclear. In this work, alkylation-protection and site-specific cross-linking experiments found that cpSRP43 makes extensive contacts with all the TMDs in LHCP. Site-directed mutagenesis identified a class of cpSRP43 mutants that bind tightly to the L18 sequence but are defective in chaperoning full-length LHCP. These mutations mapped to hydrophobic surfaces on or near the bridging helix and the \u03b2-hairpins lining the ankyrin repeat motifs of cpSRP43, suggesting that these regions are potential sites for interaction with the client TMDs. Our results suggest a working model for client protein interactions in this membrane protein chaperone.", "date": "2018-06-08", "date_type": "published", "publication": "Journal of Biological Chemistry", "volume": "293", "number": "23", "publisher": "American Society for Biochemistry and Molecular Biology", "pagerange": "8861-8873", "id_number": "CaltechAUTHORS:20180423-092135973", "issn": "0021-9258", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20180423-092135973", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Gordon and Betty Moore Foundation", "grant_number": "94-3397785" }, { "agency": "NIH", "grant_number": "R01 GM114390" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "2 T32 GM 7616-36" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF775" }, { "agency": "Caltech Beckman Institute" } ] }, "doi": "10.1074/jbc.RA118.002215", "pmcid": "PMC5995501", "primary_object": { "basename": "135723_1_supp_107935_p6dqkd.tif", "url": "https://authors.library.caltech.edu/records/2f7mb-jzr72/files/135723_1_supp_107935_p6dqkd.tif" }, "related_objects": [ { "basename": "J._Biol._Chem.-2018-McAvoy-8861-73.pdf", "url": "https://authors.library.caltech.edu/records/2f7mb-jzr72/files/J._Biol._Chem.-2018-McAvoy-8861-73.pdf" }, { "basename": "J._Biol._Chem.-2018-McAvoy-jbc.RA118.002215.pdf", "url": "https://authors.library.caltech.edu/records/2f7mb-jzr72/files/J._Biol._Chem.-2018-McAvoy-jbc.RA118.002215.pdf" } ], "resource_type": "article", "pub_year": "2018", "author_list": "McAvoy, Camille; Siegel, Alex; et el." }, { "id": "https://authors.library.caltech.edu/records/aa816-66378", "eprint_id": 85432, "eprint_status": "archive", "datestamp": "2023-08-19 08:50:37", "lastmod": "2023-10-18 18:13:28", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Kobayashi-Kan", "name": { "family": "Kobayashi", "given": "Kan" } }, { "id": "Jomaa-Ahmad", "name": { "family": "Jomaa", "given": "Ahmad" }, "orcid": "0000-0002-4786-934X" }, { "id": "Lee-Jae-Ho", "name": { "family": "Lee", "given": "Jae Ho" }, "orcid": "0000-0002-8663-3209" }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Boehringer-Daniel", "name": { "family": "Boehringer", "given": "Daniel" }, "orcid": "0000-0002-6666-8447" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Ban-Nenad", "name": { "family": "Ban", "given": "Nenad" }, "orcid": "0000-0002-9527-210X" } ] }, "title": "Structure of a prehandover mammalian ribosomal SRP\u2022SRP receptor targeting complex", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2018 American Association for the Advancement of Science. \n\nReceived for publication December 18, 2017. Accepted for publication March 12, 2018. \n\nCryo-EM data was collected at the Scientific Center for Optical and Electron Microscopy at the ETH Zurich (ScopeM). We thank P. Tittmann and A. Scaiola for support with data collection and C. Aylett and M. Itten for support with data processing. We also thank P. Bieri for initial help with microsome preparation and structure model building. We are grateful for M. Leibundgut for the critical comments on our structure model and manuscript. \n\nFunding: K.K. was supported by Long-Term Fellowships from Toyobo Biotechnology Foundation and European Molecular Biology Organization (EMBO) (ALTF 660-2015). This study was supported by the Swiss National Science Foundation (SNSF) (grant number 310030B_163478), National Center of Excellence in Research (NCCR) RNA & Disease Program of the SNSF (grant number 51NF40_141735) (to N.B.), NIH grant GM107368, Gordon and Betty Moore Foundation grant GBMF2939, and a fellowship from the Weston Havens Foundation (to S.-o.S.). \n\nAuthor contributions: K.K., A.J., S.-o.S., and N.B. designed the experimental strategy. K.K. prepared samples and grids for the cryo-EM analysis. K.K., A.J., and D.B. collected cryo-EM data. K.K. and A.J. processed data, built the structure model, and refined it. J.H.L. and S.C. prepared samples for the functional assays and performed them. K.K. and A.J. wrote the initial draft of the manuscript, and all authors contributed to the final version. \n\nCompeting interests: None declared. \n\nData and materials availability: The atomic coordinates have been deposited in the Protein Data Bank (PDB ID: 6FRK). The cryo-EM density map has been deposited in Electron Microscopy Data Bank (EMD-4300).\n\nAccepted Version - nihms-997827.pdf
Supplemental Material - aar7924_Kobayashi_SM.pdf
", "abstract": "Signal recognition particle (SRP) targets proteins to the endoplasmic reticulum (ER). SRP recognizes the ribosome synthesizing a signal sequence and delivers it to the SRP receptor (SR) on the ER membrane followed by the transfer of the signal sequence to the translocon. Here, we present the cryo-EM structure of the mammalian translating ribosome in complex with SRP and SR in a conformation preceding signal sequence handover. The structure visualizes all eukaryotic-specific SRP and SR proteins and reveals their roles in stabilizing this conformation by forming a large protein assembly at the distal site of SRP RNA. We provide biochemical evidence that the GTP hydrolysis of SRP\u2022SR is delayed at this stage, possibly to provide a time window for signal sequence handover to the translocon.", "date": "2018-04-20", "date_type": "published", "publication": "Science", "volume": "360", "number": "6386", "publisher": "American Association for the Advancement of Science", "pagerange": "323-327", "id_number": "CaltechAUTHORS:20180326-131741830", "issn": "0036-8075", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20180326-131741830", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Toyobo Biotechnology Foundation" }, { "agency": "European Molecular Biology Organization (EMBO)", "grant_number": "ALTF 660-2015" }, { "agency": "Swiss National Science Foundation (SNSF)", "grant_number": "310030B_163478" }, { "agency": "Swiss National Science Foundation (SNSF)", "grant_number": "51NF40_141735" }, { "agency": "NIH", "grant_number": "GM107368" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" }, { "agency": "Weston Havens Foundation" } ] }, "doi": "10.1126/science.aar7924", "pmcid": "PMC6309883", "primary_object": { "basename": "nihms-997827.pdf", "url": "https://authors.library.caltech.edu/records/aa816-66378/files/nihms-997827.pdf" }, "related_objects": [ { "basename": "aar7924_Kobayashi_SM.pdf", "url": "https://authors.library.caltech.edu/records/aa816-66378/files/aar7924_Kobayashi_SM.pdf" } ], "resource_type": "article", "pub_year": "2018", "author_list": "Kobayashi, Kan; Jomaa, Ahmad; et el." }, { "id": "https://authors.library.caltech.edu/records/5xhfs-tpq38", "eprint_id": 85436, "eprint_status": "archive", "datestamp": "2023-08-21 23:09:41", "lastmod": "2023-10-18 18:13:41", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Wang-Peng", "name": { "family": "Wang", "given": "Peng" }, "orcid": "0000-0001-8420-5326" }, { "id": "Liang-Fu-Cheng", "name": { "family": "Liang", "given": "Fu-Cheng" } }, { "id": "Wittmann-Daniel", "name": { "family": "Wittmann", "given": "Daniel" } }, { "id": "Siegel-Alex", "name": { "family": "Siegel", "given": "Alex" }, "orcid": "0000-0003-3601-5178" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Grimm-Bernhard", "name": { "family": "Grimm", "given": "Bernhard" }, "orcid": "0000-0002-9730-1074" } ] }, "title": "Chloroplast SRP43 acts as a chaperone for glutamyl-tRNA reductase, the rate-limiting enzyme in tetrapyrrole biosynthesis", "ispublished": "pub", "full_text_status": "public", "keywords": "chloroplast signal recognition particle; chaperone; tetrapyrrole \nbiosynthesis; 5-aminolevulinic acid; chloroplast biogenesis", "note": "\u00a9 2018 National Academy of Sciences. Published under the PNAS license. \n\nEdited by Donald R. Ort, University of Illinois at Urbana\u2013Champaign, Urbana, IL, and approved March 7, 2018 (received for review November 13, 2017) \n\nWe thank Prof. Danja Sch\u00fcnemann (Ruhr University Bochum) for the help with Arabidopsis cpsrp mutants, GST-cpSRP43 expression vector, and antibodies against cpSRP components. We thank Dr. B. Hedtke and J. Apitz (B.G. laboratory) for help with research materials. This work was supported by a fellowship from the Alexander von Humboldt Foundation (to P.W.) and by grants from the Betty and Gordon Moore Foundation (Grant 94-3397785) and NIH (Grant R01 GM114390) (both to S.-o.S.) and a grant from the Deutsche Forschungsgemeinschaft (FOR2092, Grant GR 936/18-1 to B.G.). \n\nAuthor contributions: P.W., F.-C.L., D.W., A.S., S.-o.S., and B.G. designed research; P.W., F.-C.L., D.W., and A.S. performed research; P.W., F.-C.L., D.W., A.S., S.-o.S., and B.G. analyzed data; and P.W., S.-o.S., and B.G. wrote the paper. \n\nThe authors declare no conflict of interest. \n\nThis article is a PNAS Direct Submission. \n\nThis article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1719645115/-/DCSupplemental.\n\nPublished - E3588.full.pdf
Supplemental Material - pnas.1719645115.sapp.pdf
", "abstract": "Assembly of light-harvesting complexes requires synchronization of chlorophyll (Chl) biosynthesis with biogenesis of light-harvesting Chl a/b-binding proteins (LHCPs). The chloroplast signal recognition particle (cpSRP) pathway is responsible for transport of nucleus-encoded LHCPs in the stroma of the plastid and their integration into the thylakoid membranes. Correct folding and assembly of LHCPs require the incorporation of Chls, whose biosynthesis must therefore be precisely coordinated with membrane insertion of LHCPs. How the spatiotemporal coordination between the cpSRP machinery and Chl biosynthesis is achieved is poorly understood. In this work, we demonstrate a direct interaction between cpSRP43, the chaperone that mediates LHCP targeting and insertion, and glutamyl-tRNA reductase (GluTR), a rate-limiting enzyme in tetrapyrrole biosynthesis. Concurrent deficiency for cpSRP43 and the GluTR-binding protein (GBP) additively reduces GluTR levels, indicating that cpSRP43 and GBP act nonredundantly to stabilize GluTR. The substrate-binding domain of cpSRP43 binds to the N-terminal region of GluTR, which harbors aggregation-prone motifs, and the chaperone activity of cpSRP43 efficiently prevents aggregation of these regions. Our work thus reveals a function of cpSRP43 in Chl biosynthesis and suggests a striking mechanism for posttranslational coordination of LHCP insertion with Chl biosynthesis.", "date": "2018-04-10", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "115", "number": "15", "publisher": "National Academy of Sciences", "pagerange": "E3588-E3596", "id_number": "CaltechAUTHORS:20180326-161145347", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20180326-161145347", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Alexander von Humboldt Foundation" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "94-3397785" }, { "agency": "NIH", "grant_number": "R01 GM114390" }, { "agency": "Deutsche Forschungsgemeinschaft (DFG)", "grant_number": "GR 936/18-1" } ] }, "doi": "10.1073/pnas.1719645115", "pmcid": "PMC5899456", "primary_object": { "basename": "E3588.full.pdf", "url": "https://authors.library.caltech.edu/records/5xhfs-tpq38/files/E3588.full.pdf" }, "related_objects": [ { "basename": "pnas.1719645115.sapp.pdf", "url": "https://authors.library.caltech.edu/records/5xhfs-tpq38/files/pnas.1719645115.sapp.pdf" } ], "resource_type": "article", "pub_year": "2018", "author_list": "Wang, Peng; Liang, Fu-Cheng; et el." }, { "id": "https://authors.library.caltech.edu/records/09chm-xpa03", "eprint_id": 86340, "eprint_status": "archive", "datestamp": "2023-08-19 07:43:32", "lastmod": "2023-10-18 19:29:15", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Chio-Un-Seng", "name": { "family": "Chio", "given": "Un Seng" }, "orcid": "0000-0002-5295-2690" } ] }, "title": "A Protean Clamp Guides Membrane Targeting of Tail-Anchored Proteins", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2018 Elsevier Inc. \n\nAvailable online 6 February 2018.", "abstract": "To maintain cellular organization, many chaperones and targeting factors escort nascent proteins to membrane destinations. It was generally thought that substrate proteins preferably bind conformationally closed chaperones. However, how the nature and dynamics of the targeting complex help guide substrate proteins to the target membrane is not understood for most pathways. We addressed this question for the conserved ATPase Get3, which targets the essential class of tail-anchored proteins (TAs) to the endoplasmic reticulum (ER).", "date": "2018-02-02", "date_type": "published", "publication": "Biophysical Journal", "volume": "114", "number": "3", "publisher": "Biophysical Society", "pagerange": "553a", "id_number": "CaltechAUTHORS:20180510-133648234", "issn": "0006-3495", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20180510-133648234", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1016/j.bpj.2017.11.3022", "resource_type": "article", "pub_year": "2018", "author_list": "Shan, Shu-ou and Chio, Un Seng" }, { "id": "https://authors.library.caltech.edu/records/ds6sa-rnv50", "eprint_id": 86336, "eprint_status": "archive", "datestamp": "2023-08-19 07:43:05", "lastmod": "2023-10-18 19:28:58", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Hsieh-Hao-Hsuan", "name": { "family": "Hsieh", "given": "Hao Hsuan" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Co-translational Targeting by Signal Recognition Particle Activates Only after Cytosolic Exposure of Signal Sequence", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2018 Elsevier Inc. \n\nAvailable online 6 February 2018.", "abstract": "Signal recognition particle (SRP) and its receptor (SR) co-translationally target membrane and secretory proteins to membrane specifically and efficiently through multiple layers of regulation. Differential SRP binding to ribosome, SRP-SR association rate, and GTP hydrolysis ensure specificity for substrate over non-substrate. However, there is a debate on whether SRP interacts with its substrate when a target sequence is still buried inside ribosome exit tunnel during the early stage of translation. Previous studies, which only focus on SRP binding to ribosome, produce contradictory results. In this study, we examine the full picture of SRP's multiple layers of regulation at each translation intermediate carrying increasing length of nascent chain. FRET-based SRP-ribosome binding affinity assay and fluorescence detected SRP-SR association rate measurement give the formation of key intermediates in SRP targeting cycle as a function of nascent chain length. Both parameters differ for more than two orders of magnitude between N-terminal and full exposure of targeting sequence from the end of ribosome exit tunnel. Such discrimination is comparable to the difference between substrate and non-substrate. Using e. coli as a model system, our results show that SRP targets its substrate exclusively after the emergence of a transmembrane domain or signal sequence from ribosome exit tunnel.", "date": "2018-02-02", "date_type": "published", "publication": "Biophysical Journal", "volume": "114", "number": "3", "publisher": "Biophysical Society", "pagerange": "69a", "id_number": "CaltechAUTHORS:20180510-112234591", "issn": "0006-3495", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20180510-112234591", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1016/j.bpj.2017.11.425", "resource_type": "article", "pub_year": "2018", "author_list": "Hsieh, Hao Hsuan and Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/javx8-m0a93", "eprint_id": 86573, "eprint_status": "archive", "datestamp": "2023-08-19 07:43:47", "lastmod": "2023-10-18 19:43:51", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Fu-Yu-Hsien-Hwang", "name": { "family": "Fu", "given": "Yu-Hsien Hwang" } }, { "id": "Huang-William-Y-C", "name": { "family": "Huang", "given": "William Y. C." } }, { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Groves-Jay-T", "name": { "family": "Groves", "given": "Jay T." } }, { "id": "Miller-T-F-III", "name": { "family": "Miller", "given": "Thomas" }, "orcid": "0000-0002-1882-5380" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Two-Step Membrane Binding by the Bacterial SRP Receptor Enables Efficient and Accurate Co-Translational Protein Targeting", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2018 Biophysical Society. \n\nAvailable online 6 February 2018.", "abstract": "The signal recognition particle (SRP) delivers \u223c30% of the proteome to the eukaryotic endoplasmic reticulum, or the bacterial plasma membrane. The precise mechanism by which the bacterial SRP receptor, FtsY, interacts with and is regulated at the target membrane remain unclear. Here, quantitative analysis of FtsY-lipid interactions at single-molecule resolution revealed a two-step mechanism in which FtsY initially contacts membrane via a Dynamic mode, followed by an SRP-induced conformational transition to a Stable mode that activates FtsY for downstream steps. Importantly, mutational analyses revealed extensive auto-inhibitory mechanisms that prevent free FtsY from engaging membrane in the Stable mode; an engineered FtsY pre-organized into the Stable mode led to indiscriminate targeting in vitro and disrupted FtsY function in vivo. Our results show that the two-step lipid-binding mechanism uncouples the membrane association of FtsY from its conformational activation, thus optimizing the balance between the efficiency and fidelity of co-translational protein targeting.", "date": "2018-02-02", "date_type": "published", "publication": "Biophysical Journal", "volume": "114", "number": "3", "publisher": "Biophysical Society", "pagerange": "209A", "id_number": "CaltechAUTHORS:20180523-143936913", "issn": "0006-3495", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20180523-143936913", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1016/j.bpj.2017.11.1170", "resource_type": "article", "pub_year": "2018", "author_list": "Fu, Yu-Hsien Hwang; Huang, William Y. C.; et el." }, { "id": "https://authors.library.caltech.edu/records/231w2-n9z65", "eprint_id": 84287, "eprint_status": "archive", "datestamp": "2023-08-19 06:22:32", "lastmod": "2023-10-18 16:04:43", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "McAvoy-C-Z", "name": { "family": "McAvoy", "given": "Camille" } }, { "id": "Liang-Fu-Cheng", "name": { "family": "Liang", "given": "Fu-Cheng" } }, { "id": "Miaou-Emily", "name": { "family": "Miaou", "given": "Emily" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Dynamics of Membrane Protein-Chaperone Interaction", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2017 The Protein Society.\n\nFirst published: 27 December 2017.", "abstract": "[No abstract]", "date": "2017-12", "date_type": "published", "publication": "Protein Science", "volume": "26", "number": "S1", "publisher": "Wiley", "pagerange": "29-30", "id_number": "CaltechAUTHORS:20180112-075000705", "issn": "0961-8368", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20180112-075000705", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1002/pro.3349", "resource_type": "article", "pub_year": "2017", "author_list": "McAvoy, Camille; Liang, Fu-Cheng; et el." }, { "id": "https://authors.library.caltech.edu/records/9tz8f-ny151", "eprint_id": 81798, "eprint_status": "archive", "datestamp": "2023-08-19 05:57:32", "lastmod": "2023-10-17 21:49:49", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Wang-Shuai", "name": { "family": "Wang", "given": "Shuai" }, "orcid": "0000-0002-8920-969X" }, { "id": "Yang-Chien-I", "name": { "family": "Yang", "given": "Chien-I" }, "orcid": "0000-0001-8606-5013" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "SecA mediates cotranslational targeting and translocation of an inner membrane protein", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2017 Wang et al. This article is distributed under the terms of an Attribution\u2013Noncommercial\u2013Share Alike\u2013No Mirror Sites license for the first six months after the publication date (see http ://www .rupress .org /terms/). After six months it is available under a Creative Commons License (Attribution\u2013Noncommercial\u2013Share Alike 4.0 International license, as described at https ://creativecommons .org /licenses /by -nc-sa /4.0/). \n\nSubmitted: 5 April 2017; Revision received 17 July 2017; Accepted: 2 August 2017; Published September 19, 2017. \n\nWe thank Stephen H. White and Eric Lindner for discussions and members of the Shan group for comments on the manuscript. \n\nThis work was supported by National Institutes of Health grant GM107368A and the Gordon and Betty Moore Foundation through grant GBMF2939 to S.-o. Shan. \n\nThe authors declare no competing financial interests. \n\nAuthor contributions: S. Wang designed and performed most of the experiments. C.-I Yang assisted in measuring the binding affinity of RNCs to SecA and SRP. S. Wang and S.-o. Shan wrote the paper. S.-o. Shan conceived the project. All authors reviewed the manuscript.\n\nPublished - 3639.full.pdf
Supplemental Material - JCB_201704036_TableS1.pdf
Supplemental Material - JCB_201704036_sm.pdf
", "abstract": "Protein targeting to the bacterial plasma membrane was generally thought to occur via two major pathways: cotranslational targeting by signal recognition particle (SRP) and posttranslational targeting by SecA and SecB. Recently, SecA was found to also bind ribosomes near the nascent polypeptide exit tunnel, but the function of this SecA\u2013ribosome contact remains unclear. In this study, we show that SecA cotranslationally recognizes the nascent chain of an inner membrane protein, RodZ, with high affinity and specificity. In vitro reconstitution and in vivo targeting assays show that SecA is necessary and sufficient to direct the targeting and translocation of RodZ to the bacterial plasma membrane in an obligatorily cotranslational mechanism. Sequence elements upstream and downstream of the RodZ transmembrane domain dictate nascent polypeptide selection by SecA instead of the SRP machinery. These findings identify a new route for the targeting of inner membrane proteins in bacteria and highlight the diversity of targeting pathways that enables an organism to accommodate diverse nascent proteins.", "date": "2017-11-06", "date_type": "published", "publication": "Journal of Cell Biology", "volume": "216", "number": "11", "publisher": "Rockefeller University Press", "pagerange": "3639-3653", "id_number": "CaltechAUTHORS:20170925-085525865", "issn": "0021-9525", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20170925-085525865", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM107368A" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" } ] }, "doi": "10.1083/jcb.201704036", "pmcid": "PMC5674894", "primary_object": { "basename": "3639.full.pdf", "url": "https://authors.library.caltech.edu/records/9tz8f-ny151/files/3639.full.pdf" }, "related_objects": [ { "basename": "JCB_201704036_TableS1.pdf", "url": "https://authors.library.caltech.edu/records/9tz8f-ny151/files/JCB_201704036_TableS1.pdf" }, { "basename": "JCB_201704036_sm.pdf", "url": "https://authors.library.caltech.edu/records/9tz8f-ny151/files/JCB_201704036_sm.pdf" } ], "resource_type": "article", "pub_year": "2017", "author_list": "Wang, Shuai; Yang, Chien-I; et el." }, { "id": "https://authors.library.caltech.edu/records/4ds6e-83b64", "eprint_id": 81834, "eprint_status": "archive", "datestamp": "2023-08-21 21:57:56", "lastmod": "2023-10-17 21:52:06", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Chio-Un-Seng", "name": { "family": "Chio", "given": "Un Seng" }, "orcid": "0000-0002-5295-2690" }, { "id": "Chung-SangYoon", "name": { "family": "Chung", "given": "SangYoon" }, "orcid": "0000-0002-0592-4099" }, { "id": "Weiss-Shimon", "name": { "family": "Weiss", "given": "Shimon" }, "orcid": "0000-0002-0720-5426" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "A protean clamp guides membrane targeting of tail-anchored proteins", "ispublished": "pub", "full_text_status": "public", "keywords": "protein targeting; chaperones; protein dynamics; single-molecule spectroscopy; ATPases", "note": "\u00a9 2017 National Academy of Sciences. \n\nEdited by Taekjip Ha, Johns Hopkins University, Baltimore, MD, and approved September 5, 2017 (received for review May 25, 2017). Published online before print September 26, 2017. \n\nWe thank Xavier Michalet and Antonino Ingargiola for help with analysis of \u03bcs-ALEX data; H. Cho, M. Rao, and S. Wang for reagents; the D. A. Dougherty laboratory for use of HPLC; and W. M. Clemons, A. Hoelz, R. Deshaies, M. Zimmer, T. F. Miller, M. Rome, and members of the S.-o.S. and S.W. laboratories for critical discussions and comments on the manuscript. This work was supported by NIH Grant GM107368, Gordon and Betty Moore Foundation Grant GBMF2939, and a fellowship from the Weston Havens Foundation (to S.-o.S.). \n\nAuthor contributions: U.S.C. and S.-o.S. designed research; U.S.C. and S.C. performed research; S.W. contributed new reagents/analytic tools; U.S.C. and S.C. analyzed data; and U.S.C. and S.-o.S. wrote the paper. \n\nThe authors declare no conflict of interest. \n\nThis article is a PNAS Direct Submission. \n\nThis article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1708731114/-/DCSupplemental.\n\nPublished - PNAS-2017-Chio-E8585-94.pdf
Supplemental Material - pnas.201708731SI.pdf
", "abstract": "Proper localization of proteins to target membranes is a fundamental cellular process. How the nature and dynamics of the targeting complex help guide substrate proteins to the target membrane is not understood for most pathways. Here, we address this question for the conserved ATPase guided entry of tail-anchored protein 3 (Get3), which targets the essential class of tail-anchored proteins (TAs) to the endoplasmic reticulum (ER). Single-molecule fluorescence spectroscopy showed that, contrary to previous models of a static closed Get3\u2022TA complex, Get3 samples open conformations on the submillisecond timescale upon TA binding, generating a fluctuating \"protean clamp\" that stably traps the substrate. Point mutations at the ATPase site bias Get3 toward closed conformations, uncouple TA binding from induced Get3\u2022Get4/5 disassembly, and inhibit the ER targeting of the Get3\u2022TA complex. These results demonstrate an essential role of substrate-induced Get3 dynamics in driving TA targeting to the membrane, and reveal a tightly coupled channel of communication between the TA-binding site, ATPase site, and effector interaction surfaces of Get3. Our results provide a precedent for large-scale dynamics in a substrate-bound chaperone, which provides an effective mechanism to retain substrate proteins with high affinity while also generating functional switches to drive vectorial cellular processes.", "date": "2017-10-10", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "114", "number": "41", "publisher": "National Academy of Sciences", "pagerange": "E8585-E8594", "id_number": "CaltechAUTHORS:20170926-110202110", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20170926-110202110", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM107368" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" }, { "agency": "Weston Havens Foundation" } ] }, "doi": "10.1073/pnas.1708731114", "pmcid": "PMC5642712", "primary_object": { "basename": "PNAS-2017-Chio-E8585-94.pdf", "url": "https://authors.library.caltech.edu/records/4ds6e-83b64/files/PNAS-2017-Chio-E8585-94.pdf" }, "related_objects": [ { "basename": "pnas.201708731SI.pdf", "url": "https://authors.library.caltech.edu/records/4ds6e-83b64/files/pnas.201708731SI.pdf" } ], "resource_type": "article", "pub_year": "2017", "author_list": "Chio, Un Seng; Chung, SangYoon; et el." }, { "id": "https://authors.library.caltech.edu/records/w6vr2-95t93", "eprint_id": 82316, "eprint_status": "archive", "datestamp": "2023-08-19 05:21:17", "lastmod": "2023-10-17 22:11:29", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Chio-Un-Seng", "name": { "family": "Chio", "given": "Un Seng" }, "orcid": "0000-0002-5295-2690" }, { "id": "Cho-Hyunjun", "name": { "family": "Cho", "given": "Hyunjun" }, "orcid": "0000-0002-8963-5525" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Mechanisms of Tail-Anchored Membrane Protein Targeting and Insertion", "ispublished": "pub", "full_text_status": "public", "keywords": "tail-anchored protein, protein targeting, ATPase, membrane protein biogenesis, chaperones, protein quality control", "note": "\u00a9 2017 Annual Reviews. \n\nThis work was supported by NIH grant GM107368 to S.-o.S. \n\nThe authors are not aware of any affiliations, memberships, funding, or financial holdings that might be perceived as affecting the objectivity of this review.\n\nAccepted Version - nihms-1001158.pdf
", "abstract": "Proper localization of membrane proteins is essential for the function of biological membranes and for the establishment of organelle identity within a cell. Molecular machineries that mediate membrane protein biogenesis need to not only achieve a high degree of efficiency and accuracy, but also prevent off-pathway aggregation events that can be detrimental to cells. The posttranslational targeting of tail-anchored proteins (TAs) provides tractable model systems to probe these fundamental issues. Recent advances in understanding TA-targeting pathways reveal sophisticated molecular machineries that drive and regulate these processes. These findings also suggest how an interconnected network of targeting factors, cochaperones, and quality control machineries together ensures robust membrane protein biogenesis.", "date": "2017-10", "date_type": "published", "publication": "Annual Review of Cell and Developmental Biology", "volume": "33", "publisher": "Annual Reviews", "pagerange": "417-438", "id_number": "CaltechAUTHORS:20171012-133314474", "issn": "1081-0706", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20171012-133314474", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM107368" } ] }, "doi": "10.1146/annurev-cellbio-100616-060839", "pmcid": "PMC6343671", "primary_object": { "basename": "nihms-1001158.pdf", "url": "https://authors.library.caltech.edu/records/w6vr2-95t93/files/nihms-1001158.pdf" }, "resource_type": "article", "pub_year": "2017", "author_list": "Chio, Un Seng; Cho, Hyunjun; et el." }, { "id": "https://authors.library.caltech.edu/records/j7835-h0d89", "eprint_id": 79584, "eprint_status": "archive", "datestamp": "2023-08-19 04:15:00", "lastmod": "2023-10-26 16:57:49", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Fu-Yu-Hsien-Hwang", "name": { "family": "Fu", "given": "Yu-Hsien Hwang" } }, { "id": "Huang-William-Y-C", "name": { "family": "Huang", "given": "William Y. C." } }, { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Groves-Jay-T", "name": { "family": "Groves", "given": "Jay T." } }, { "id": "Miller-T-F-III", "name": { "family": "Miller", "given": "Thomas" }, "orcid": "0000-0002-1882-5380" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Two-step membrane binding by the bacterial SRP receptor enable efficient and accurate Co-translational protein targeting", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2017 Hwang Fu et al. This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited. \n\nReceived: 12 February 2017; Accepted: 28 June 2017; Published: 28 July 2017. \n\nWe thank Connie Wang and members of the Shan lab for critical discussions and comments on the manuscript; Wen-Chen Lin, and Meredith Triplet from the Groves lab for the helps on SLB techniques; Oliver Loson from the Chan lab alumni for help on checking the SLB quality; Heun Jin Lee and Tal Einav from the Phillips Lab for helping us set up real-time delivery instruments and for modeling discussions, respectively. This work was supported by NIH grant GM078024 and the Gordon and Betty Moore Foundation through Grant GBMF2939 to SS. \n\nThe funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.\n\nAuthor Contributions: Y-HHF, Conceptualization, Data curation, Software, Formal analysis, Investigation, Visualization, Methodology, Writing\u2014original draft, Writing\u2014review and editing; WYCH, Validation, Methodology, Writing\u2014review and editing; KS, Validation, Investigation, Methodology, Writing\u2014review and\nediting; JTG, Resources, Supervision, Validation, Methodology, Writing\u2014review and editing; TM, Formal analysis, Methodology, Writing\u2014review and editing; S-oS, Conceptualization, Formal analysis, Supervision, Funding acquisition, Visualization, Writing\u2014review and editing. \n\nThe authors declare that no competing interests exist.\n\nPublished - elife-25885-v1.pdf
", "abstract": "The signal recognition particle (SRP) delivers ~30% of the proteome to the eukaryotic endoplasmic reticulum, or the bacterial plasma membrane. The precise mechanism by which the bacterial SRP receptor, FtsY, interacts with and is regulated at the target membrane remain unclear. Here, quantitative analysis of FtsY-lipid interactions at single-molecule resolution revealed a two-step mechanism in which FtsY initially contacts membrane via a Dynamic mode, followed by an SRP-induced conformational transition to a Stable mode that activates FtsY for downstream steps. Importantly, mutational analyses revealed extensive auto-inhibitory mechanisms that prevent free FtsY from engaging membrane in the Stable mode; an engineered FtsY pre-organized into the Stable mode led to indiscriminate targeting in vitro and disrupted FtsY function in vivo. Our results show that the two-step lipid-binding mechanism uncouples the membrane association of FtsY from its conformational activation, thus optimizing the balance between the efficiency and fidelity of co-translational protein targeting.", "date": "2017-07-28", "date_type": "published", "publication": "eLife", "volume": "6", "publisher": "eLife Sciences Publications", "pagerange": "Art. No. e25885", "id_number": "CaltechAUTHORS:20170731-091633412", "issn": "2050-084X", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20170731-091633412", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" } ] }, "doi": "10.7554/eLife.25885", "pmcid": "PMC5533587", "primary_object": { "basename": "elife-25885-v1.pdf", "url": "https://authors.library.caltech.edu/records/j7835-h0d89/files/elife-25885-v1.pdf" }, "resource_type": "article", "pub_year": "2017", "author_list": "Fu, Yu-Hsien Hwang; Huang, William Y. C.; et el." }, { "id": "https://authors.library.caltech.edu/records/zbhb7-ybg73", "eprint_id": 77627, "eprint_status": "archive", "datestamp": "2023-08-19 03:11:30", "lastmod": "2023-10-25 23:18:43", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Jomaa-Ahmad", "name": { "family": "Jomaa", "given": "Ahmad" }, "orcid": "0000-0002-4786-934X" }, { "id": "Fu-Yu-Hsien-Hwang", "name": { "family": "Fu", "given": "Yu-Hsien Hwang" } }, { "id": "Boehringer-Daniel", "name": { "family": "Boehringer", "given": "Daniel" }, "orcid": "0000-0002-6666-8447" }, { "id": "Leibundgut-Marc", "name": { "family": "Leibundgut", "given": "Marc" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Ban-Nenad", "name": { "family": "Ban", "given": "Nenad" }, "orcid": "0000-0002-9527-210X" } ] }, "title": "Structure of the quaternary complex between SRP, SR, and translocon bound to the translating ribosome", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2017 The Author(s). This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. \n\nReceived: 25 January 2017; Accepted: 29 March 2017; Published online: 19 May 2017. \n\nWe would like to thank David Ramrath, Philipp Bieri and members of the Ban lab for discussions. Cryo-EM data was collected at the Scientific Center for Optical and Electron Microscopy at the ETH Zurich (ScopeM). We thank Shuai Wang for preparation of translation extracts and SecYEG used in the GTPase assays. We are thankful to Peter Tittmann and members of the ScopeM facility for technical support. We acknowledge the use of computing infrastructure provided by the Central Information Technology Services of the ETH Zurich. This work was supported by the Swiss National Science Foundation (SNSF), the National Center of Excellence in Research (NCCR) Structural Biology and RNA & Disease programs of the SNSF, and European Research Council grant 250071 under the European Community's Seventh Framework Programme (to N.B.) and by National Institutes of Health (NIH) Grant GM078024 (to S.-o.S). \n\nAuthor Contributions: N.B. and A.J. conceived the study. N.B., S.-o.S. and A.J. designed experiments. A.J., D.B. and M.L. performed cryo-EM data collection, image processing and modelling. Y.-H.H.F. performed GTPase stimulation assays. A.J., N.B. and S.-o.S. wrote the manuscript. All authors discussed the results, commented on the data and contributed to the final version of the manuscript. \n\nThe authors declare no competing financial interests.\n\nPublished - ncomms15470.pdf
Supplemental Material - ncomms15470-s1.pdf
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Supplemental Material - ncomms15470-s3.mov
Supplemental Material - ncomms15470-s4.mov
Supplemental Material - ncomms15470-s5.mov
Supplemental Material - ncomms15470-s6.mov
", "abstract": "During co-translational protein targeting, the signal recognition particle (SRP) binds to the translating ribosome displaying the signal sequence to deliver it to the SRP receptor (SR) on the membrane, where the signal peptide is transferred to the translocon. Using electron cryo-microscopy, we have determined the structure of a quaternary complex of the translating Escherichia coli ribosome, the SRP\u2013SR in the 'activated' state and the translocon. Our structure, supported by biochemical experiments, reveals that the SRP RNA adopts a kinked and untwisted conformation to allow repositioning of the 'activated' SRP\u2013SR complex on the ribosome. In addition, we observe the translocon positioned through interactions with the SR in the vicinity of the ribosome exit tunnel where the signal sequence is extending beyond its hydrophobic binding groove of the SRP M domain towards the translocon. Our study provides new insights into the mechanism of signal sequence transfer from the SRP to the translocon.", "date": "2017-05-19", "date_type": "published", "publication": "Nature Communications", "volume": "8", "publisher": "Nature Publishing Group", "pagerange": "Art. No. 15470", "id_number": "CaltechAUTHORS:20170522-110612032", "issn": "2041-1723", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20170522-110612032", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Swiss National Science Foundation (SNSF)" }, { "agency": "European Research Council (ERC)", "grant_number": "250071" }, { "agency": "NIH", "grant_number": "GM078024" } ] }, "doi": "10.1038/ncomms15470", "pmcid": "PMC5454536", "primary_object": { "basename": "ncomms15470-s1.pdf", "url": "https://authors.library.caltech.edu/records/zbhb7-ybg73/files/ncomms15470-s1.pdf" }, "related_objects": [ { "basename": "ncomms15470-s2.mov", "url": "https://authors.library.caltech.edu/records/zbhb7-ybg73/files/ncomms15470-s2.mov" }, { "basename": "ncomms15470-s3.mov", "url": "https://authors.library.caltech.edu/records/zbhb7-ybg73/files/ncomms15470-s3.mov" }, { "basename": "ncomms15470-s4.mov", "url": "https://authors.library.caltech.edu/records/zbhb7-ybg73/files/ncomms15470-s4.mov" }, { "basename": "ncomms15470-s5.mov", "url": "https://authors.library.caltech.edu/records/zbhb7-ybg73/files/ncomms15470-s5.mov" }, { "basename": "ncomms15470-s6.mov", "url": "https://authors.library.caltech.edu/records/zbhb7-ybg73/files/ncomms15470-s6.mov" }, { "basename": "ncomms15470.pdf", "url": "https://authors.library.caltech.edu/records/zbhb7-ybg73/files/ncomms15470.pdf" } ], "resource_type": "article", "pub_year": "2017", "author_list": "Jomaa, Ahmad; Fu, Yu-Hsien Hwang; et el." }, { "id": "https://authors.library.caltech.edu/records/456yp-w8w75", "eprint_id": 72562, "eprint_status": "archive", "datestamp": "2023-08-22 19:30:36", "lastmod": "2023-10-23 22:43:38", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Anionic Phospholipids and the Albino3 Translocase Activate Signal Recognition Particle-Receptor Interaction during Light-harvesting Chlorophyll a/b-binding Protein Targeting", "ispublished": "pub", "full_text_status": "public", "keywords": "chloroplast; GTPase; phospholipid; protein targeting; signal recognition particle (SRP); GTPases", "note": "\u00a9 2017 The American Society for Biochemistry and Molecular Biology, Inc. Under an Attribution 4.0 International (CC BY 4.0). \n\nReceived August 9, 2016; Revision received October 14, 2016; First Published on November 28, 2016. \n\nThis work was supported by National Institutes of Health Grant 1R01GM114390 and fellowships from the Gordon and Betty Moore Foundation and American Federation for Aging Research (to S. S.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. \n\nWe thank members of the Shan laboratory for comments on the manuscript. \n\nAuthor Contributions: S. S. conceived and coordinated the study and wrote the paper. S. C. designed, performed, and analyzed the experiments and wrote the paper. All authors reviewed the results and approved the final version of the manuscript.\n\nPublished - J._Biol._Chem.-2017-Chandrasekar-397-406.pdf
", "abstract": "The universally conserved signal recognition particle (SRP) co-translationally delivers newly synthesized membrane and secretory proteins to the target cellular membrane. The only exception is found in the chloroplast of green plants, where the chloroplast SRP (cpSRP) post-translationally targets light-harvesting chlorophyll a/b-binding proteins (LHCP) to the thylakoid membrane. The mechanism and regulation of this post-translational mode of targeting by cpSRP remain unclear. Using biochemical and biophysical methods, here we show that anionic phospholipids activate the cpSRP receptor cpFtsY to promote rapid and stable cpSRP54\u00b7cpFtsY complex assembly. Furthermore, the stromal domain of the Alb3 translocase binds with high affinity to and regulates GTP hydrolysis in the cpSRP54\u00b7cpFtsY complex, suggesting that cpFtsY is primarily responsible for initial recruitment of the targeting complex to Alb3. These results suggest a new model for the sequential recruitment, remodeling, and unloading of the targeting complex at membrane translocase sites in the post-translational cpSRP pathway.", "date": "2017-01-06", "date_type": "published", "publication": "Journal of Biological Chemistry", "volume": "292", "number": "1", "publisher": "American Society for Biochemistry and Molecular Biology", "pagerange": "397-406", "id_number": "CaltechAUTHORS:20161205-134337068", "issn": "0021-9258", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20161205-134337068", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "1R01GM114390" }, { "agency": "Gordon and Betty Moore Foundation" }, { "agency": "American Federation for Aging Research" } ] }, "doi": "10.1074/jbc.M116.752956", "pmcid": "PMC5217697", "primary_object": { "basename": "J._Biol._Chem.-2017-Chandrasekar-397-406.pdf", "url": "https://authors.library.caltech.edu/records/456yp-w8w75/files/J._Biol._Chem.-2017-Chandrasekar-397-406.pdf" }, "resource_type": "article", "pub_year": "2017", "author_list": "Chandrasekar, Sowmya and Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/1hmcv-k2d90", "eprint_id": 72552, "eprint_status": "archive", "datestamp": "2023-08-22 19:30:28", "lastmod": "2023-10-23 22:43:07", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Sweredoski-Michael-J", "name": { "family": "Sweredoski", "given": "Michael J." }, "orcid": "0000-0003-0878-3831" }, { "id": "Sohn-Chang-Ho", "name": { "family": "Sohn", "given": "Chang Ho" } }, { "id": "Hess-Sonja", "name": { "family": "Hess", "given": "Sonja" }, "orcid": "0000-0002-5904-9816" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Co-evolution of two GTPases enables efficient protein targeting in an RNA-less chloroplast Signal Recognition Particle pathway", "ispublished": "pub", "full_text_status": "public", "keywords": "chloroplast; fluorescence; GTPase; protein cross-linking; protein evolution; protein targeting; signal recognition particle (SRP); GTPases", "note": "\u00a9 2017 The American Society for Biochemistry and Molecular Biology, Inc. \n\nReceived August 9, 2016; Revision received November 17, 2016; First Published on November 28, 2016. \n\nThis work was supported by National Institutes of Health Grant 1R01GM114390 and fellowships from the Gordon and Betty Moore Foundation and American Federation for Aging Research (to S. S.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. \n\nWe thank Jesse Beauchamp for helpful discussions on the cross-linking and mass spectrometry experiments; members of the Proteome Exploration Laboratory for help with mass spectrometry experiments and analyses; the Guttman laboratory for help with phylogenic analysis, and members of the Shan laboratory for comments on the manuscript. The Proteome Exploration Laboratory was supported by the Beckman Institute and the Gordon and Betty Moore Foundation through Grant GBMF775. \n\nAuthor Contributions: S. S. conceived and coordinated the study and wrote the paper. S. C. designed, performed, and analyzed the experiments shown in Figs. 1\u20135, carried out the analyses in Fig. 6 and Table 1, and wrote the paper. S. C., C. H. S., S. H., and M. J. S. designed, performed, and analyzed the experiments shown in Fig. 2 and Table 1. All authors reviewed the results and approved the final version of the manuscript.\n\nPublished - J._Biol._Chem.-2017-Chandrasekar-386-96.pdf
", "abstract": "The signal recognition particle (SRP) is an essential ribonucleoprotein particle that mediates the co-translational targeting of newly synthesized proteins to cellular membranes. The SRP RNA is a universally conserved component of SRP that mediates key interactions between two GTPases in SRP and its receptor, thus enabling rapid delivery of cargo to the target membrane. Notably, this essential RNA is bypassed in the chloroplast (cp) SRP of green plants. Previously, we showed that the cpSRP and cpSRP receptor GTPases (cpSRP54 and cpFtsY, respectively) interact efficiently by themselves without the SRP RNA. Here, we explore the molecular mechanism by which this is accomplished. Fluorescence analyses showed that, in the absence of SRP RNA, the M-domain of cpSRP54 both accelerates and stabilizes complex assembly between cpSRP54 and cpFtsY. Cross-linking coupled with mass spectrometry and mutational analyses identified a new interaction between complementarily charged residues on the cpFtsY G-domain and the vicinity of the cpSRP54 M-domain. These residues are specifically conserved in plastids, and their evolution coincides with the loss of SRP RNA in green plants. These results provide an example of how proteins replace the functions of RNA during evolution.", "date": "2017-01-06", "date_type": "published", "publication": "Journal of Biological Chemistry", "volume": "292", "number": "1", "publisher": "American Society for Biochemistry and Molecular Biology", "pagerange": "386-396", "id_number": "CaltechAUTHORS:20161205-103747522", "issn": "0021-9258", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20161205-103747522", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "1R01GM114390" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF775" }, { "agency": "American Federation for Aging Research" }, { "agency": "Caltech Beckman Institute" } ] }, "doi": "10.1074/jbc.M116.752931", "pmcid": "PMC5217696", "primary_object": { "basename": "J._Biol._Chem.-2017-Chandrasekar-386-96.pdf", "url": "https://authors.library.caltech.edu/records/1hmcv-k2d90/files/J._Biol._Chem.-2017-Chandrasekar-386-96.pdf" }, "resource_type": "article", "pub_year": "2017", "author_list": "Chandrasekar, Sowmya; Sweredoski, Michael J.; et el." }, { "id": "https://authors.library.caltech.edu/records/mqs9g-yn993", "eprint_id": 72720, "eprint_status": "archive", "datestamp": "2023-08-19 00:28:48", "lastmod": "2023-10-23 22:52:55", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Rao-Meera", "name": { "family": "Rao", "given": "Meera" } }, { "id": "Okreglak-V", "name": { "family": "Okreglak", "given": "Voytek" } }, { "id": "Chio-Un-Seng", "name": { "family": "Chio", "given": "Un Seng" }, "orcid": "0000-0002-5295-2690" }, { "id": "Cho-Hyunjun", "name": { "family": "Cho", "given": "Hyunjun" } }, { "id": "Walter-Peter", "name": { "family": "Walter", "given": "Peter" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Multiple selection filters ensure accurate tail-anchored membrane protein targeting", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2016, Rao et al. This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited. \n\nReceived September 06, 2016; Accepted December 06, 2016; Published December 07, 2016. \n\nWe thank Bil Clemons, Michael Rome, Kuang Shen, Xin Zhang, and members of the Shan and Clemons labs for critical discussions and comments on the manuscript, and the Dougherty lab for use of their HPLC. This work was supported by NIH grant GM107368 \nand Gordon and Betty Moore Foundation Grant GBMF2939 to S.S., NIH grants R01GM32384 & U01GM098254 to P.W., NIH training grant 5T32GM007616-33 to M.R, and the Leukemia and Lymphoma Society fellowship to V.O. P.W. is an Investigator of the Howard Hughes Medical Institute. \n\nThe funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication. \n\nThe authors declare that no competing interests exist.\n\nPublished - e21301-download.pdf
", "abstract": "Accurate protein localization is crucial to generate and maintain organization in all cells. Achieving accuracy is challenging, as the molecular signals that dictate a protein's cellular destination are often promiscuous. A salient example is the targeting of an essential class of tail-anchored (TA) proteins, whose sole defining feature is a transmembrane domain near their C-terminus. Here we show that the Guided Entry of Tail-anchored protein (GET) pathway selects TA proteins destined to the endoplasmic reticulum (ER) utilizing distinct molecular steps, including differential binding by the co-chaperone Sgt2 and kinetic proofreading after ATP hydrolysis by the targeting factor Get3. Further, the different steps select for distinct physicochemical features of the TA substrate. The use of multiple selection filters may be general to protein biogenesis pathways that must distinguish correct and incorrect substrates based on minor differences.", "date": "2016-12-07", "date_type": "published", "publication": "eLife", "volume": "5", "publisher": "eLife Sciences Publications", "pagerange": "Art. No. e21301", "id_number": "CaltechAUTHORS:20161212-104646015", "issn": "2050-084X", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20161212-104646015", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM107368" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" }, { "agency": "NIH", "grant_number": "R01GM32384" }, { "agency": "NIH", "grant_number": "U01GM098254" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "5T32GM007616-33" }, { "agency": "Leukemia and Lymphoma Society" }, { "agency": "Howard Hughes Medical Institute (HHMI)" } ] }, "doi": "10.7554/eLife.21301", "pmcid": "PMC5214336", "primary_object": { "basename": "e21301-download.pdf", "url": "https://authors.library.caltech.edu/records/mqs9g-yn993/files/e21301-download.pdf" }, "resource_type": "article", "pub_year": "2016", "author_list": "Rao, Meera; Okreglak, Voytek; et el." }, { "id": "https://authors.library.caltech.edu/records/4rdv8-cvx45", "eprint_id": 70419, "eprint_status": "archive", "datestamp": "2023-08-22 19:10:56", "lastmod": "2023-10-20 22:56:46", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "ATPase and GTPase Tangos Drive Intracellular Protein Transport", "ispublished": "pub", "full_text_status": "public", "keywords": "ATPases; Get3; GTPases; molecular recognition and regulation; protein targeting; signal recognition particle", "note": "\u00a9 2016 Elsevier. \n\nAvailable online 19 September 2016. \n\nThe author thanks members of her laboratory for helpful comments on the manuscript. This work was supported by NIH grants GM078024 and GM107368 and by the Gordon and Betty Moore Foundation through Grant GBMF2939 to S.S.\n\nAccepted Version - nihms817981.pdf
", "abstract": "The GTPase superfamily of proteins provides molecular switches to regulate numerous cellular processes. The 'GTPase switch' paradigm, in which external regulatory factors control the switch of a GTPase between 'on' and 'off' states, has been used to interpret the regulatory mechanism of many GTPases. However, recent work unveiled a class of nucleotide hydrolases that do not adhere to this classical paradigm. Instead, they use nucleotide-dependent dimerization cycles to regulate key cellular processes. In this review article, recent studies of dimeric GTPases and ATPases involved in intracellular protein targeting are summarized. It is suggested that these proteins can use the conformational plasticity at their dimer interface to generate multiple points of regulation, thereby providing the driving force and spatiotemporal coordination of complex cellular pathways.", "date": "2016-12", "date_type": "published", "publication": "Trends in Biochemical Sciences", "volume": "41", "number": "12", "publisher": "Cell Press", "pagerange": "1050-1060", "id_number": "CaltechAUTHORS:20160919-100630353", "issn": "0968-0004", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20160919-100630353", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "NIH", "grant_number": "GM107368" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" } ] }, "doi": "10.1016/j.tibs.2016.08.012", "pmcid": "PMC5627767", "primary_object": { "basename": "nihms817981.pdf", "url": "https://authors.library.caltech.edu/records/4rdv8-cvx45/files/nihms817981.pdf" }, "resource_type": "article", "pub_year": "2016", "author_list": "Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/d44kg-76h80", "eprint_id": 72088, "eprint_status": "archive", "datestamp": "2023-08-20 14:13:54", "lastmod": "2023-10-23 17:54:24", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Chen-Yang", "name": { "family": "Chen", "given": "Yang" }, "orcid": "0000-0002-8964-0084" }, { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" }, "orcid": "0000-0002-0873-758X" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Kou-S-C", "name": { "family": "Kou", "given": "S. C." } } ] }, "title": "Analyzing Single-Molecule Protein Transportation Experiments via Hierarchical Hidden Markov Models", "ispublished": "pub", "full_text_status": "public", "keywords": "Conformational change, FRET, Hierarchical model, MCMC (Markov chain Monte Carlo), Model checking, Protein targeting", "note": "\u00a9 2016 American Statistical Association. \n\nReceived 01 Aug 2014, Accepted author version posted online: 02 Feb 2016, Published online: 18 Oct 2016. \n\nS. Shan's research is supported in part by NIH grant GM078024 and the Gordon and Betty Moore Foundation through Grant GBMF2939. S. C. Kou's research is supported in part by grants from NSF and ARO.\n\nAccepted Version - nihms814888.pdf
", "abstract": "To maintain proper cellular functions, over 50% of proteins encoded in the genome need to be transported to cellular membranes. The molecular mechanism behind such a process, often referred to as protein targeting, is not well understood. Single-molecule experiments are designed to unveil the detailed mechanisms and reveal the functions of different molecular machineries involved in the process. The experimental data consist of hundreds of stochastic time traces from the fluorescence recordings of the experimental system. We introduce a Bayesian hierarchical model on top of hidden Markov models (HMMs) to analyze these data and use the statistical results to answer the biological questions. In addition to resolving the biological puzzles and delineating the regulating roles of different molecular complexes, our statistical results enable us to propose a more detailed mechanism for the late stages of the protein targeting process.", "date": "2016-10-18", "date_type": "published", "publication": "Journal of the American Statistical Association", "volume": "111", "number": "515", "publisher": "Taylor & Francis", "pagerange": "951-966", "id_number": "CaltechAUTHORS:20161117-073554872", "issn": "0162-1459", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20161117-073554872", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" }, { "agency": "NSF" }, { "agency": "Army Research Office (ARO)" } ] }, "doi": "10.1080/01621459.2016.1140050", "pmcid": "PMC5606165", "primary_object": { "basename": "nihms814888.pdf", "url": "https://authors.library.caltech.edu/records/d44kg-76h80/files/nihms814888.pdf" }, "resource_type": "article", "pub_year": "2016", "author_list": "Chen, Yang; Shen, Kuang; et el." }, { "id": "https://authors.library.caltech.edu/records/9z85m-rwb97", "eprint_id": 67340, "eprint_status": "archive", "datestamp": "2023-08-22 17:41:44", "lastmod": "2023-10-18 21:06:28", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Liu-Wenpeng", "name": { "family": "Liu", "given": "Wenpeng" } }, { "id": "Zhou-Mian", "name": { "family": "Zhou", "given": "Mian" } }, { "id": "Li-Zhengke", "name": { "family": "Li", "given": "Zhengke" } }, { "id": "Li-Hongzhi", "name": { "family": "Li", "given": "Hongzhi" } }, { "id": "Polaczek-Piotr", "name": { "family": "Polaczek", "given": "Piotr" } }, { "id": "Dai-Huifang", "name": { "family": "Dai", "given": "Huifang" } }, { "id": "Wu-Qiong", "name": { "family": "Wu", "given": "Qiong" } }, { "id": "Liu-Changwei", "name": { "family": "Liu", "given": "Changwei" } }, { "id": "Karanja-Kenneth-K", "name": { "family": "Karanja", "given": "Kenneth K." } }, { "id": "Popuri-Vencat", "name": { "family": "Popuri", "given": "Vencat" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Schlacher-Katharina", "name": { "family": "Schlacher", "given": "Katharina" }, "orcid": "0000-0001-7226-6391" }, { "id": "Zheng-Li", "name": { "family": "Zheng", "given": "Li" }, "orcid": "0000-0002-3744-185X" }, { "id": "Campbell-J-L", "name": { "family": "Campbell", "given": "Judith L." } }, { "id": "Shen-Binghui", "name": { "family": "Shen", "given": "Binghui" }, "orcid": "0000-0002-4408-407X" } ] }, "title": "A Selective Small Molecule DNA2 Inhibitor for Sensitization of Human Cancer Cells to Chemotherapy", "ispublished": "pub", "full_text_status": "public", "keywords": "DNA2 inhibitor; Nuclease; Helicase; DNA binding; DNA end resection; DNA replicatoin fork protection; Chemotherapy; Sensitizer; Camptothecin; PARP inhibitor; Cancer", "note": "\u00a9 2016 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). \n\nReceived 22 February 2016; Received in revised form 29 February 2016; Accepted 29 February 2016; Available online 10 March 2016. \n\nFunding Sources: The work was supported by an NIH grant RO1 CA085344, the Margaret E. Early Medical Research Trust grant ADF-1481M to B.H.S., Margaret Early Medical Research Trust grant and ARO W81XWH-09-1-0041 to J.L.C., a Caltech-City of Hope Biomedical Research Initiative to B.H.S. and J.L.C., and K22 grant 5K22CA175262\u201304 to K.S. Wenpeng Liu is supported by the China Scholarship Council (CSC) during his visit to California Institute of Technology and City of Hope, and K.S. is a CPRIT scholar in cancer biology. \n\nWe thank the City of Hope Microscopy and Bioinformatics Core Facilities, National Cancer Institute (NCI) designated cancer center support grant P30 CA033572, and Nancy Linford, Ph.D. for her critical reading and editing of the manuscript and S. Howard and J. Stark for their technical support. \n\nAuthor Contributions: L.Z., J.L.C. and B.S. designed the study and supervised the entire project. L.Z., J.L.C., B.S. and K.S. wrote the manuscript. H.L. established the 3-D model and virtually screened the chemical inhibitors. W.L., P.P., J.L.C., Q.W., C.L, and K.K.J. purified the proteins and performed all of the biochemical experiments in the current study. K.S. and V.P. designed and executed the fork protection experiments. Z.L. performed the DNA fiber experiments. Correspondence concerning fork protection experiments may be addressed to K.S. and V.P. M.Z., Z.L., H.D., K.K.J., L.C., performed the cellular functional (SSA, HR, P-RPA resection, replication fork restart) and toxicological assays. S.S. supervised the enzyme kinetics experiments. \n\nWe declare no financial or other relationships that may lead to a conflict of interest in this study.\n\nPublished - 1-s2.0-S2352396416300822-main.pdf
Supplemental Material - mmc1.pdf
", "abstract": "Cancer cells frequently up-regulate DNA replication and repair proteins such as the multifunctional DNA2 nuclease/helicase, counteracting DNA damage due to replication stress and promoting survival. Therefore, we hypothesized that blocking both DNA replication and repair by inhibiting the bifunctional DNA2 could be a potent strategy to sensitize cancer cells to stresses from radiation or chemotherapeutic agents. We show that homozygous deletion of DNA2 sensitizes cells to ionizing radiation and camptothecin (CPT). Using a virtual high throughput screen, we identify 4-hydroxy-8-nitroquinoline-3-carboxylic acid (C5) as an effective and selective inhibitor of DNA2. Mutagenesis and biochemical analysis define the C5 binding pocket at a DNA-binding motif that is shared by the nuclease and helicase activities, consistent with structural studies that suggest that DNA binding to the helicase domain is necessary for nuclease activity. C5 targets the known functions of DNA2 in vivo: C5 inhibits resection at stalled forks as well as reducing recombination. C5 is an even more potent inhibitor of restart of stalled DNA replication forks and over-resection of nascent DNA in cells defective in replication fork protection, including BRCA2 and BOD1L. C5 sensitizes cells to CPT and synergizes with PARP inhibitors.", "date": "2016-04", "date_type": "published", "publication": "EBioMedicine", "volume": "6", "publisher": "Elsevier", "pagerange": "73-86", "id_number": "CaltechAUTHORS:20160525-094720168", "issn": "2352-3964", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20160525-094720168", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "RO1 CA085344" }, { "agency": "Margaret E. Early Medical Research Trust", "grant_number": "ADF-1481M" }, { "agency": "Army Research Office (ARO)", "grant_number": "W81XWH-09-1-0041" }, { "agency": "Caltech-City of Hope Biomedical Initiative" }, { "agency": "NIH", "grant_number": "5K22CA175262\u201304" }, { "agency": "China Scholarship Council" } ] }, "doi": "10.1016/j.ebiom.2016.02.043", "pmcid": "PMC4856754", "primary_object": { "basename": "1-s2.0-S2352396416300822-main.pdf", "url": "https://authors.library.caltech.edu/records/9z85m-rwb97/files/1-s2.0-S2352396416300822-main.pdf" }, "related_objects": [ { "basename": "mmc1.pdf", "url": "https://authors.library.caltech.edu/records/9z85m-rwb97/files/mmc1.pdf" } ], "resource_type": "article", "pub_year": "2016", "author_list": "Liu, Wenpeng; Zhou, Mian; et el." }, { "id": "https://authors.library.caltech.edu/records/d89nh-76s96", "eprint_id": 65172, "eprint_status": "archive", "datestamp": "2023-08-22 17:38:26", "lastmod": "2023-10-17 23:15:04", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Liang-Fu-Cheng", "name": { "family": "Liang", "given": "Fu-Cheng" } }, { "id": "Kroon-Gerard-J", "name": { "family": "Kroon", "given": "Gerard" }, "orcid": "0000-0001-8038-7476" }, { "id": "McAvoy-Camille-Z", "name": { "family": "McAvoy", "given": "Camille Z." } }, { "id": "Chi-Chris", "name": { "family": "Chi", "given": "Chris" } }, { "id": "Wright-Peter-E", "name": { "family": "Wright", "given": "Peter E." }, "orcid": "0000-0002-1368-0223" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Conformational dynamics of a membrane protein chaperone enables spatially regulated substrate capture and release", "ispublished": "pub", "full_text_status": "public", "keywords": "membrane protein biogenesis; molecular chaperone; signal recognition particle; protein dynamics; NMR spectroscopy", "note": "\u00a9 2016 National Academy of Sciences. \n\nEdited by Gerhard Wagner, Harvard Medical School, Boston, MA, and approved February 4, 2016 (received for review December 16, 2015).\nPublished online before print March 7, 2016. \n\nWe thank R. Dalbey and P. Jaru-Ampornpan for plasmid encoding Alb3CT, V. Q. Lam and M. Yamout for initial optimization of the isotope labeling condition, P. Aoto for advice on NMR assignments, S. Chandrasekar and S. Lieblich (S. Mayo's laboratory) for help with analytical ultracentrifugation, and members of the S.-o.S. laboratory for comments on the manuscript. This work was supported by fellowships from the Gordon and Betty Moore Foundation and American Federation for Aging Research. (to S.-o.S.) and the Skaggs Institute of Chemical Biology (to P.E.W.). \n\nAuthor contributions: F.-C.L., G.K., C.Z.M., P.E.W., and S.-o.S. designed research; F.-C.L., G.K., and C.Z.M. performed research; F.-C.L., G.K., C.Z.M., C.C., P.E.W., and S.-o.S. analyzed data; and F.-C.L., P.E.W., and S.-o.S. wrote the paper. \n\nThe authors declare no conflict of interest. \n\nThis article is a PNAS Direct Submission. \n\nThis article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1524777113/-/DCSupplemental.\n\nPublished - PNAS-2016-Liang-E1615-24.pdf
Supplemental Material - pnas.201524777SI.pdf
", "abstract": "Membrane protein biogenesis poses enormous challenges to cellular protein homeostasis and requires effective molecular chaperones. Compared with chaperones that promote soluble protein folding, membrane protein chaperones require tight spatiotemporal coordination of their substrate binding and release cycles. Here we define the chaperone cycle for cpSRP43, which protects the largest family of membrane proteins, the light harvesting chlorophyll a/b-binding proteins (LHCPs), during their delivery. Biochemical and NMR analyses demonstrate that cpSRP43 samples three distinct conformations. The stromal factor cpSRP54 drives cpSRP43 to the active state, allowing it to tightly bind substrate in the aqueous compartment. Bidentate interactions with the Alb3 translocase drive cpSRP43 to a partially inactive state, triggering selective release of LHCP's transmembrane domains in a productive unloading complex at the membrane. Our work demonstrates how the intrinsic conformational dynamics of a chaperone enables spatially coordinated substrate capture and release, which may be general to other ATP-independent chaperone systems.", "date": "2016-03-22", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "113", "number": "12", "publisher": "National Academy of Sciences", "pagerange": "E1615-E1624", "id_number": "CaltechAUTHORS:20160308-073550753", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20160308-073550753", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Gordon and Betty Moore Foundation" }, { "agency": "American Federation for Aging Research" }, { "agency": "Skaggs Institute of Chemical Biology" } ] }, "doi": "10.1073/pnas.1524777113", "pmcid": "PMC4812700", "primary_object": { "basename": "PNAS-2016-Liang-E1615-24.pdf", "url": "https://authors.library.caltech.edu/records/d89nh-76s96/files/PNAS-2016-Liang-E1615-24.pdf" }, "related_objects": [ { "basename": "pnas.201524777SI.pdf", "url": "https://authors.library.caltech.edu/records/d89nh-76s96/files/pnas.201524777SI.pdf" } ], "resource_type": "article", "pub_year": "2016", "author_list": "Liang, Fu-Cheng; Kroon, Gerard; et el." }, { "id": "https://authors.library.caltech.edu/records/mysna-gfz86", "eprint_id": 67674, "eprint_status": "archive", "datestamp": "2023-08-20 10:22:24", "lastmod": "2023-10-18 21:22:54", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Wang-Connie-Y", "name": { "family": "Wang", "given": "Connie" }, "orcid": "0000-0003-2971-3971" }, { "id": "Wang-Shuai", "name": { "family": "Wang", "given": "Shuai" }, "orcid": "0000-0002-8920-969X" }, { "id": "Niesen-M-J-M", "name": { "family": "Niesen", "given": "Michiel" }, "orcid": "0000-0002-9255-6203" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Miller-T-F-III", "name": { "family": "Miller", "given": "Thomas F., III" }, "orcid": "0000-0002-1882-5380" } ] }, "title": "Inversion of Signal Sequence Topology during Membrane Integration", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2016 Biophysical Society. Published by Elsevier Inc.", "abstract": "In many cases, the topology of membrane proteins is established during co-translational membrane integration. This process involves the Sec translocon, a heterotrimeric protein-conducting channel that allows for both the translocation of secreted domains across the membrane through the central pore and the integration of membrane domains directly into the lipid bilayer through a lateral opening. For many proteins, the N-terminus is retained on the cytosolic side of the membrane and forms an inverted (type II) topology that threads the C-terminus through the channel. This inverted topology is hypothesized to involve a head-first intermediate which then undergoes a step-wise inversion process to its final topology. \n\nWe use a newly developed coarse-grained model to simulate the integration of the signal sequence during the elongation of the nascent chain on the minute-long timescales that are relevant to the biological process. This coarse-grained simulation method enables direct comparisons to experimentally measured energetics of ribosome-nascent chain to translocon interactions. We observe a series of pulling and pushing forces on the ribosome-nascent chain as translation proceeds and identify a head-first intermediate whose inversion is driven by the entropic confinement of nascent chain residues in the ribosome-translocon junction.", "date": "2016-02-16", "date_type": "published", "publication": "Biophysical Journal", "volume": "110", "number": "3", "publisher": "Biophysical Society", "pagerange": "226A-227A", "id_number": "CaltechAUTHORS:20160606-080929408", "issn": "0006-3495", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20160606-080929408", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1016/j.bpj.2015.11.1252", "resource_type": "article", "pub_year": "2016", "author_list": "Wang, Connie; Wang, Shuai; et el." }, { "id": "https://authors.library.caltech.edu/records/sg29c-c5129", "eprint_id": 61251, "eprint_status": "archive", "datestamp": "2023-08-20 09:28:17", "lastmod": "2023-10-25 14:39:33", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Gristick-Harry-B", "name": { "family": "Gristick", "given": "Harry B." }, "orcid": "0000-0002-1957-2821" }, { "id": "Rome-Michael-E", "name": { "family": "Rome", "given": "Michael E." } }, { "id": "Chartron-Justin-W", "name": { "family": "Chartron", "given": "Justin W." }, "orcid": "0000-0003-1734-3424" }, { "id": "Rao-Meera", "name": { "family": "Rao", "given": "Meera" }, "orcid": "0000-0001-8650-6253" }, { "id": "Hess-Sonja", "name": { "family": "Hess", "given": "Sonja" }, "orcid": "0000-0002-5904-9816" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Clemons-W-M-Jr", "name": { "family": "Clemons", "given": "William M., Jr." }, "orcid": "0000-0002-0021-889X" } ] }, "title": "Mechanism of assembly of a substrate-transfer complex during tail-anchored protein targeting", "ispublished": "pub", "full_text_status": "public", "keywords": "Tail-anchor targeting, GET pathway, protein transport, X-ray crystallography", "note": "\u00a9 2015 The American Society for Biochemistry and Molecular Biology, Inc. \n\nReceived July 7, 2015; Revision received October 2, 2015; First Published on October 7, 2015. \n\nThis work was supported by National Science Foundation Graduate Research Fellowship DGE-1144469 (to M. E. R.) and National Institutes of Health Training Grant 5T32GM007616-33 (to H. B. G. and M. R.) and Research Grants R01GM097572 (to W. M. C.) and R01GM107368 (to S. S.). The Proteome Exploration Laboratory was supported by Gordon and Betty Moore Foundation Grant GBMF775 and the Beckman Institute (to S. H.). \n\n\nThe atomic coordinates and structure factors (codes 5BW8 and 5BWK) have been deposited in the Protein Data Bank (http://wwpdb.org/). \n\nThe authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. \n\nWe thank Graeme Card, Ana Gonzalez, and Michael Soltis for help with data collection at Stanford Synchrotron Radiation Lightsource Beamline 12-2. We are grateful to Gordon and Betty Moore for support of the Molecular Observatory at Caltech. Operations at Stanford Synchrotron Radiation Lightsource are supported by the United States Department of Energy and National Institutes of Health. \n\nAuthor Contributions: H. B. G., M. E. R., S. S., and W. M. C. designed the study and wrote the paper. H. B. G. determined the 2.8-\u00c5 crystal structure and characterized the intermediate complex. M. E. R. performed experiments to define stoichiometry and kinetic analysis. J. W. C. determined the structure of the 6.0-\u00c5 complex. M. R. helped conceptually with the study. S. H. assisted in the mass spectrometry analysis. All authors reviewed the results and approved the final version of the manuscript.\n\nPublished - J._Biol._Chem.-2015-Gristick-30006-17.pdf
", "abstract": "Tail-anchored (TA) proteins, defined as having a single transmembrane helix at their C terminus, are post-translationally targeted to the endoplasmic reticulum membrane by the guided entry of TA proteins (GET) pathway. In yeast, the handover of TA substrates is mediated by the heterotetrameric Get4/Get5 complex (Get4/5), which tethers the co-chaperone Sgt2 to the targeting factor, the Get3 ATPase. Binding of Get4/5 to Get3 is critical for efficient TA targeting; however, questions remain about the formation of the Get3\u00b7Get4/5 complex. Here we report crystal structures of a Get3\u00b7Get4/5 complex from Saccharomyces cerevisiae at 2.8 and 6.0 \u00c5 that reveal a novel interface between Get3 and Get4 dominated by electrostatic interactions. Kinetic and mutational analyses strongly suggest that these structures represent an on-pathway intermediate that rapidly assembles and then rearranges to the final Get3\u00b7Get4/5 complex. Furthermore, we provide evidence that the Get3\u00b7Get4/5 complex is dominated by a single Get4/5 heterotetramer bound to one monomer of a Get3 dimer, uncovering an intriguing asymmetry in the Get4/5 heterotetramer upon Get3 binding. Ultrafast diffusion-limited electrostatically driven Get3\u00b7Get4/5 association enables Get4/5 to rapidly sample and capture Get3 at different stages of the GET pathway.", "date": "2015-12-11", "date_type": "published", "publication": "Journal of Biological Chemistry", "volume": "290", "number": "50", "publisher": "American Society for Biochemistry and Molecular Biology", "pagerange": "30006-30017", "id_number": "CaltechAUTHORS:20151019-101737280", "issn": "0021-9258", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20151019-101737280", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NSF Graduate Research Fellowship", "grant_number": "DGE-1144469" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "5T32GM007616-33" }, { "agency": "NIH", "grant_number": "R01GM097572" }, { "agency": "NIH", "grant_number": "R01GM107368" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF775" }, { "agency": "Caltech Beckman Institute" }, { "agency": "Department of Energy (DOE)" } ] }, "doi": "10.1074/jbc.M115.677328", "pmcid": "PMC4705998", "primary_object": { "basename": "J._Biol._Chem.-2015-Gristick-30006-17.pdf", "url": "https://authors.library.caltech.edu/records/sg29c-c5129/files/J._Biol._Chem.-2015-Gristick-30006-17.pdf" }, "resource_type": "article", "pub_year": "2015", "author_list": "Gristick, Harry B.; Rome, Michael E.; et el." }, { "id": "https://authors.library.caltech.edu/records/1579d-b6a61", "eprint_id": 58125, "eprint_status": "archive", "datestamp": "2023-08-22 15:45:33", "lastmod": "2023-10-23 19:04:50", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Ariosa-Aileen-R", "name": { "family": "Ariosa", "given": "Aileen" } }, { "id": "Lee-Jae-Ho", "name": { "family": "Lee", "given": "Jae Ho" }, "orcid": "0000-0002-8663-3209" }, { "id": "Wang-Shuai", "name": { "family": "Wang", "given": "Shuai" }, "orcid": "0000-0002-8920-969X" }, { "id": "Saraogi-Ishu", "name": { "family": "Saraogi", "given": "Ishu" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Regulation by a chaperone improves substrate selectivity during cotranslational protein targeting", "ispublished": "pub", "full_text_status": "public", "keywords": "signal recognition particle; trigger factor; ribosome; protein biogenesis; GTPases", "note": "\u00a9 2015 National Academy of Sciences.\n\nEdited by Jonathan S. Weissman, University of California, San Francisco, Howard Hughes Medical Institute, and California Institute for Quantitative\nBiosciences, San Francisco, CA, and approved May 5, 2015 (received for review November 25, 2014).\nPublished online before print June 8, 2015.\n\nWe thank Xin Zhang and Kuang Shen for insightful\ndiscussions, and Harris Bernstein and members of the S.-o.S. group for\ncomments on the manuscript. This work was supported by National Institutes\nof Health (NIH) Grant GM078024 to S.-o.S. and the NIH NIGMS Ruth L.\nKirschstein National Research Service Award (F31GM095294) to A.A. This\nproject was funded in part by the Henry Dreyfus Teacher-Scholar Award, the\nPackard and Lucile Fellowship in science and engineering, and the Gordon\nand Betty Moore Foundation through Grant GBMF2939 (to S.-o.S.).\n\nAuthor contributions: A.A., J.H.L., and S.-o.S. designed research; A.A., J.H.L., S.W., I.S., and\nS.-o.S. performed research; A.A., J.H.L., S.W., I.S., and S.-o.S. analyzed data; and A.A. and\nS.-o.S. wrote the paper.\n\nThe authors declare no conflict of interest.\n\nThis article is a PNAS Direct Submission.\n\nThis article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1422594112/-/DCSupplemental.\n\nPublished - PNAS-2015-Ariosa-E3169-78.pdf
Supplemental Material - pnas.201422594SI.pdf
", "abstract": "The ribosome exit site is a crowded environment where numerous factors contact nascent polypeptides to influence their folding, localization, and quality control. Timely and accurate selection of nascent polypeptides into the correct pathway is essential for proper protein biogenesis. To understand how this is accomplished, we probe the mechanism by which nascent polypeptides are accurately sorted between the major cotranslational chaperone trigger factor (TF) and the essential cotranslational targeting machinery, signal recognition particle (SRP). We show that TF regulates SRP function at three distinct stages, including binding of the translating ribosome, membrane targeting via recruitment of the SRP receptor, and rejection of ribosome-bound nascent polypeptides beyond a critical length. Together, these mechanisms enhance the specificity of substrate selection into both pathways. Our results reveal a multilayered mechanism of molecular interplay at the ribosome exit site, and provide a conceptual framework to understand how proteins are selected among distinct biogenesis machineries in this crowded environment.", "date": "2015-06-23", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "112", "number": "25", "publisher": "National Academy of Sciences", "pagerange": "E3169-E3178", "id_number": "CaltechAUTHORS:20150609-122913362", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20150609-122913362", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "F31GM095294" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" } ] }, "doi": "10.1073/pnas.1422594112", "pmcid": "PMC4485088", "primary_object": { "basename": "PNAS-2015-Ariosa-E3169-78.pdf", "url": "https://authors.library.caltech.edu/records/1579d-b6a61/files/PNAS-2015-Ariosa-E3169-78.pdf" }, "related_objects": [ { "basename": "pnas.201422594SI.pdf", "url": "https://authors.library.caltech.edu/records/1579d-b6a61/files/pnas.201422594SI.pdf" } ], "resource_type": "article", "pub_year": "2015", "author_list": "Ariosa, Aileen; Lee, Jae Ho; et el." }, { "id": "https://authors.library.caltech.edu/records/x1837-13n39", "eprint_id": 55881, "eprint_status": "archive", "datestamp": "2023-08-22 15:14:20", "lastmod": "2023-10-20 23:21:46", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "von-Loeffelholz-Ottilie", "name": { "family": "von Loeffelholz", "given": "Ottilie" } }, { "id": "Jiang-Qiyang", "name": { "family": "Jiang", "given": "Qiyang" } }, { "id": "Ariosa-Aileen-R", "name": { "family": "Ariosa", "given": "Aileen R." } }, { "id": "Karuppasamy-Manikandan", "name": { "family": "Karuppasamy", "given": "Manikandan" } }, { "id": "Huard-Karine", "name": { "family": "Huard", "given": "Karine" } }, { "id": "Berger-Imre", "name": { "family": "Berger", "given": "Imre" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Schaffitzel-Christiane", "name": { "family": "Schaffitzel", "given": "Christiane" } } ] }, "title": "Ribosome\u2013SRP\u2013FtsY cotranslational targeting complex in the closed state", "ispublished": "pub", "full_text_status": "public", "keywords": "protein targeting; signal recognition particle; signal sequence; ribosome; single-particle electron cryomicroscopy", "note": "\u00a9 2015 National Academy of Sciences. \n\nEdited by Joachim Frank, Howard Hughes Medical Institute, Columbia University, New York, NY, and approved February 20, 2015 (received for review December 30, 2014). Published online before print March 16, 2015, doi: 10.1073/pnas.1424453112 .\n\nWe thank Sejeong Lee and Wolfgang Wintermeyer (Max Planck Institute for Biophysical Chemistry, G\u00f6ttingen, Germany) for discussions and information about Lep50; Wim Hagen for data collection; the protein expression facility at EMBL Heidelberg and the Partnership for Structural Biology in Grenoble for support; and all members of C.S.'s group for discussion and advice with image processing. The Polara microscope is part of the Institut de Biologie Structurale Structural Biology and Dynamics GIS-IBISA\u2013labeled platform. Support was provided by ERC Starting Grant\nProject 281331 (to C.S.). \n\nAuthor contributions: O.v.L., S.-o.S., and C.S. designed research; O.v.L., Q.J., A.A., M.K., and K.H. performed research; O.v.L., Q.J., A.A., M.K., and C.S. analyzed data; and O.v.L., Q.J., I.B., S.-o.S., and C.S. wrote the paper.\n\nThe authors declare no conflict of interest.\n\nThis article is a PNAS Direct Submission.\n\nData deposition: The EM maps and atomic models have been deposited in the Protein Data Bank, www.pdb.org (PDB ID code 5AKA), and EMDataBank, www.emdatabank.org (accession no. EMD-2917).\n\nThis article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1424453112/-/DCSupplemental.\n\nPublished - 3943.full.pdf
Supplemental Material - pnas.1424453112.sapp.pdf
", "abstract": "The signal recognition particle (SRP)-dependent pathway is essential for correct targeting of proteins to the membrane and subsequent insertion in the membrane or secretion. In Escherichia coli, the SRP and its receptor FtsY bind to ribosome\u2013nascent chain complexes with signal sequences and undergo a series of distinct conformational changes, which ensures accurate timing and fidelity of protein targeting. Initial recruitment of the SRP receptor FtsY to the SRP\u2013RNC complex results in GTP-independent binding of the SRP\u2013FtsY GTPases at the SRP RNA tetraloop. In the presence of GTP, a closed state is adopted by the SRP\u2013FtsY complex. The cryo-EM structure of the closed state reveals an ordered SRP RNA and SRP M domain with a signal sequence-bound. Van der Waals interactions between the finger loop and ribosomal protein L24 lead to a constricted signal sequence-binding pocket possibly preventing premature release of the signal sequence. Conserved M-domain residues contact ribosomal RNA helices 24 and 59. The SRP\u2013FtsY GTPases are detached from the RNA tetraloop and flexible, thus liberating the ribosomal exit site for binding of the translocation machinery.", "date": "2015-03-31", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "112", "number": "13", "publisher": "National Academy of Sciences", "pagerange": "3943-3948", "id_number": "CaltechAUTHORS:20150318-091732909", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20150318-091732909", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "European Research Council (ERC)", "grant_number": "281331" } ] }, "doi": "10.1073/pnas.1424453112", "pmcid": "PMC4386334", "primary_object": { "basename": "3943.full.pdf", "url": "https://authors.library.caltech.edu/records/x1837-13n39/files/3943.full.pdf" }, "related_objects": [ { "basename": "pnas.1424453112.sapp.pdf", "url": "https://authors.library.caltech.edu/records/x1837-13n39/files/pnas.1424453112.sapp.pdf" } ], "resource_type": "article", "pub_year": "2015", "author_list": "von Loeffelholz, Ottilie; Jiang, Qiyang; et el." }, { "id": "https://authors.library.caltech.edu/records/vg4yw-j0s29", "eprint_id": 61924, "eprint_status": "archive", "datestamp": "2023-08-20 04:39:53", "lastmod": "2023-10-25 16:04:12", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Fu-Yu-Hsien-Hwang", "name": { "family": "Fu", "given": "Yu-Hsien Hwang" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Distinct Membrane Association Modes Facilitate Co-Translational Protein Targeting", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2015 Biophysical Society.", "abstract": "Deciphering the nature of protein interactions at the membrane surface is crucial to understanding the molecular mechanism, timing, and regulation of many essential cellular processes. The signal recognition particle (SRP) and its receptor (SR), which target the translating ribosomes to translocation machineries at the target membrane, provide an ideal model system for investigating the mechanism of membrane recruitment of protein machineries.\n\nHere we show that FtsY, the E. coli SR, interacts with the membrane through two distinct modes. Using single-molecule techniques that directly observe membrane association of FtsY in a supported lipid bilayer environment, we were able to detect and differentiate between the previously identified stable association mode as well as a transient interaction mode that is also crucial for targeting. Transition between these two modes is regulated by the SRP. Functional assays show that switching between transient and stabilized modes upon SRP binding is critical for successful targeting. We propose that, in contrast to currently accepted models in which FtsY is anchored on the membrane and awaits SRP encounter, this receptor rapidly cycles between cytosol and membrane and actively searches for cargo-loaded SRP. This active search could provide a mechanism to enhance efficient and accurate membrane localization of the targeting complex.", "date": "2015-01-27", "date_type": "published", "publication": "Biophysical Journal", "volume": "108", "number": "2", "publisher": "Biophysical Society", "pagerange": "253A", "id_number": "CaltechAUTHORS:20151106-080405216", "issn": "0006-3495", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20151106-080405216", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1016/j.bpj.2014.11.1401", "resource_type": "article", "pub_year": "2015", "author_list": "Fu, Yu-Hsien Hwang and Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/ssrma-1ng38", "eprint_id": 62615, "eprint_status": "archive", "datestamp": "2023-08-20 04:27:11", "lastmod": "2023-10-25 17:16:49", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Liang-Fu-Cheng", "name": { "family": "Liang", "given": "Fu-Cheng" } }, { "id": "McAvoy-C", "name": { "family": "McAvoy", "given": "Camille" } }, { "id": "Piszkiewicz-S", "name": { "family": "Piszkiewicz", "given": "Samantha" } }, { "id": "Kroon-G-J", "name": { "family": "Kroon", "given": "Gerard J." } }, { "id": "Yamout-M", "name": { "family": "Yamout", "given": "Maria" } }, { "id": "Wright-P", "name": { "family": "Wright", "given": "Peter" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Inter-Domain Dynamics of a Novel Chaperone Enables Effective Capture of Membrane Protein Substrates", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2015 Biophysical Society. Published by Elsevier Inc.", "abstract": "Protein homeostasis is essential for all cells and requires proper control of the folding, localization and interactions of proteins. The biogenesis of membrane proteins poses a particular challenge to the protein homeostasis network, wherein molecular chaperones play an important role by preventing or reversing membrane protein aggregation, maintaining them in translocation competent states before they reach the membrane destination", "date": "2015-01", "date_type": "published", "publication": "Biophysical Journal", "volume": "108", "number": "2", "publisher": "Biophysical Society", "pagerange": "53a", "id_number": "CaltechAUTHORS:20151204-105119596", "issn": "0006-3495", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20151204-105119596", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1016/j.bpj.2014.11.323", "resource_type": "article", "pub_year": "2015", "author_list": "Liang, Fu-Cheng; McAvoy, Camille; et el." }, { "id": "https://authors.library.caltech.edu/records/aeabp-gfj80", "eprint_id": 51289, "eprint_status": "archive", "datestamp": "2023-08-22 14:13:39", "lastmod": "2023-10-18 16:15:54", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Rome-M-E", "name": { "family": "Rome", "given": "Michael E." } }, { "id": "Chio-Un-Seng", "name": { "family": "Chio", "given": "Un Seng" }, "orcid": "0000-0002-5295-2690" }, { "id": "Rao-Meera", "name": { "family": "Rao", "given": "Meera" } }, { "id": "Gristick-H-B", "name": { "family": "Gristick", "given": "Harry" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Differential gradients of interaction affinities drive efficient targeting and recycling in the GET pathway", "ispublished": "pub", "full_text_status": "public", "keywords": "protein targeting; tail-anchored protein; ATPase; protein interaction cascades; fluorescence", "note": "\u00a9 2014 National Academy of Sciences.\n\nEdited by Douglas C. Rees, Howard Hughes Medical Institute, Caltech, Pasadena, CA, and approved October 2, 2014 (received for review June 20, 2014).\n\nPublished online before print November 3, 2014.\n\nWe thank Bil Clemons for Get1/2-CD expression\nconstructs, purification protocols, critical discussions, and sharing unpublished\nstructural data; Bob Keenan and Manu Hegde for Get1/2-FL expression\nvectors; Vlad Denic for mini-Get1/2 constructs; Peter Walter for\nKar2 antibody; David Akopian for help with PLs; Dennis Woo for help\nwith yeast insertion assays; and members of the laboratory of S.-o.S.\nfor critical comments on the manuscript. This work was supported by\nNational Science Foundation Graduate Research Fellowship DGE-1144469\n(to M.E.R.), National Institutes of Health Training Grant 5T32GM007616-\n33 (to M.R.), and the David and Lucile Packard Foundation Career Award\n(to S.-o.S.).\n\nAuthor contributions: M.E.R. and S.-o.S. designed research; M.E.R., U.S.C., and M.R. performed\nresearch; M.E.R. and H.G. contributed new reagents/analytic tools; M.E.R., U.S.C.,\nM.R., and S.-o.S. analyzed data; and M.E.R., U.S.C., and S.-o.S. wrote the paper.\n\nThe authors declare no conflict of interest.\n\nThis Direct Submission article had a prearranged editor.\n\nThis article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.\n1073/pnas.1411284111/-/DCSupplemental.\n\nPublished - E4929.full.pdf
Supplemental Material - pnas.201411284SI.pdf
", "abstract": "Efficient and accurate localization of membrane proteins requires a complex cascade of interactions between protein machineries. This requirement is exemplified in the guided entry of tail-anchored (TA) protein (GET) pathway, where the central targeting factor Get3 must sequentially interact with three distinct binding partners to ensure the delivery of TA proteins to the endoplasmic reticulum (ER) membrane. To understand the molecular principles that provide the vectorial driving force of these interactions, we developed quantitative fluorescence assays to monitor Get3\u2013effector interactions at each stage of targeting. We show that nucleotide and substrate generate differential gradients of interaction energies that drive the ordered interaction of Get3 with successive effectors. These data also provide more molecular details on how the targeting complex is captured and disassembled by the ER receptor and reveal a previously unidentified role for Get4/5 in recycling Get3 from the ER membrane at the end of the targeting reaction. These results provide general insights into how complex protein interaction cascades are coupled to energy inputs in biological systems.", "date": "2014-11-06", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "111", "number": "46", "publisher": "National Academy of Sciences", "pagerange": "E4929-E4935", "id_number": "CaltechAUTHORS:20141105-102115424", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20141105-102115424", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NSF Graduate Research Fellowship", "grant_number": "DGE-1144469" }, { "agency": "NIH", "grant_number": "5T32GM007616-33" }, { "agency": "David and Lucile Packard Foundation" } ] }, "doi": "10.1073/pnas.1411284111", "pmcid": "PMC4246279", "primary_object": { "basename": "E4929.full.pdf", "url": "https://authors.library.caltech.edu/records/aeabp-gfj80/files/E4929.full.pdf" }, "related_objects": [ { "basename": "pnas.201411284SI.pdf", "url": "https://authors.library.caltech.edu/records/aeabp-gfj80/files/pnas.201411284SI.pdf" } ], "resource_type": "article", "pub_year": "2014", "author_list": "Rome, Michael E.; Chio, Un Seng; et el." }, { "id": "https://authors.library.caltech.edu/records/hh3bw-wsh14", "eprint_id": 43221, "eprint_status": "archive", "datestamp": "2023-08-22 13:16:40", "lastmod": "2023-10-25 23:22:31", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Saraogi-Ishu", "name": { "family": "Saraogi", "given": "Ishu" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Co-translational protein targeting to the bacterial membrane", "ispublished": "pub", "full_text_status": "public", "keywords": "Protein targeting; SecYEG; GTPases; Molecular recognition and regulation; Ribosome", "note": "\u00a9 2013 Elsevier B.V.\n\nReceived 10 August 2013; Received in revised form 9 October 2013;\nAccepted 16 October 2013; Available online 24 October 2013.\n\nWe thank Sandra Schmid, Jennifer Doudna, Peter Walter, Ramanujan Hegde, Douglas Rees, Raymond Deshaies and Bil Clemons for support and insightful discussions over the years, and David Akopian for critical reading of the manuscript. S.S. was supported by NIH grant GM078024\nand by career awards from the Henry and Camille Dreyfus foundation, the Arnold and Mabel Beckman foundation, and the David and Lucile Packard foundation. I.S. was supported by a grant from the Betty and Gordon Moore Foundation.\n\nAccepted Version - nihms534497.pdf
", "abstract": "Co-translational protein targeting by the Signal Recognition Particle (SRP) is an essential cellular pathway that couples the synthesis of nascent proteins to their proper cellular localization. The bacterial SRP, which contains the minimal ribonucleoprotein core of this universally conserved targeting machine, has served as a paradigm for understanding the molecular basis of protein localization in all cells. In this review, we highlight recent biochemical and structural insights into the molecular mechanisms by which fundamental challenges faced by protein targeting machineries are met in the SRP pathway. Collectively, these studies elucidate how an essential SRP RNA and two regulatory GTPases in the SRP and SRP receptor (SR) enable this targeting machinery to recognize, sense and respond to its biological effectors, i.e. the cargo protein, the target membrane and the translocation machinery, thus driving efficient and faithful co-translational protein targeting. This article is part of a Special Issue entitled: Protein trafficking & Secretion.", "date": "2014-08", "date_type": "published", "publication": "Biochimica et Biophysica Acta - Molecular Cell Research", "volume": "1843", "number": "8", "publisher": "Elsevier", "pagerange": "1433-1441", "id_number": "CaltechAUTHORS:20140106-100007866", "issn": "0167-4889", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140106-100007866", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Gordon and Betty Moore Foundation" } ] }, "doi": "10.1016/j.bbamcr.2013.10.013", "pmcid": "PMC3999308", "primary_object": { "basename": "nihms534497.pdf", "url": "https://authors.library.caltech.edu/records/hh3bw-wsh14/files/nihms534497.pdf" }, "resource_type": "article", "pub_year": "2014", "author_list": "Saraogi, Ishu and Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/k40hn-71q52", "eprint_id": 46260, "eprint_status": "archive", "datestamp": "2023-08-20 01:22:39", "lastmod": "2023-10-26 19:37:50", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Saraogi-Ishu", "name": { "family": "Saraogi", "given": "Ishu" } }, { "id": "Akopian-D", "name": { "family": "Akopian", "given": "David" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Regulation of cargo recognition, commitment, and unloading drives cotranslational protein targeting", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2014 Saraogi et al. This article is distributed under the terms of an Attribution\u2013Noncommercial\u2013Share Alike\u2013No Mirror Sites license for the first six months after the publication date (see\nhttp://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution\u2013Noncommercial\u2013Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/). \n\nSubmitted: 7 November 2013; Accepted: 1 May 2014; Published June 9, 2014. \n\nWe thank Tom Miller and Connie Wang for discussions and members of the Shan group for comments on the manuscript. \n\nThis work was supported by National Institutes of Health grant\nGM078024 to S.-o. Shan. S.-o. Shan was supported by the Henry Dreyfus teacher-scholar award and the Packard and Lucile Fellowship in science and engineering. This project is funded in part by the Gordon and Betty Moore Foundation through grant GBMF2939 to S.-o. Shan. \n\nThe authors declare no competing financial interests.\n\nPublished - J_Cell_Biol-2014-Saraogi-693-706.pdf
Supplemental Material - JCB_201311028_sm.pdf
", "abstract": "Efficient and accurate protein localization is essential\nto cells and requires protein-targeting machineries\nto both effectively capture the cargo in the\ncytosol and productively unload the cargo at the membrane.\nTo understand how these challenges are met, we\nfollowed the interaction of translating ribosomes during\ntheir targeting by the signal recognition particle (SRP)\nusing a site-specific fluorescent probe in the nascent\nprotein. We show that initial recruitment of SRP receptor\n(SR) selectively enhances the affinity of SRP for correct\ncargos, thus committing SRP-dependent substrates to the\npathway. Real-time measurement of cargo transfer from\nthe targeting to translocation machinery revealed multiple\nfactors that drive this event, including GTPase rearrangement\nin the SRP\u2013SR complex, stepwise displacement\nof SRP from the ribosome and signal sequence by SecYEG,\nand elongation of the nascent polypeptide. Our\nresults elucidate how active and sequential regulation of\nthe SRP\u2013cargo interaction drives efficient and faithful\nprotein targeting.", "date": "2014-06-09", "date_type": "published", "publication": "Journal of Cell Biology", "volume": "205", "number": "5", "publisher": "Rockefeller University Press", "pagerange": "693-706", "id_number": "CaltechAUTHORS:20140613-090320700", "issn": "0021-9525", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140613-090320700", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Gordon and Betty Moore Foundation", "grant_number": "GBMF2939" } ] }, "doi": "10.1083/jcb.201311028", "pmcid": "PMC4050729", "primary_object": { "basename": "JCB_201311028_sm.pdf", "url": "https://authors.library.caltech.edu/records/k40hn-71q52/files/JCB_201311028_sm.pdf" }, "related_objects": [ { "basename": "J_Cell_Biol-2014-Saraogi-693-706.pdf", "url": "https://authors.library.caltech.edu/records/k40hn-71q52/files/J_Cell_Biol-2014-Saraogi-693-706.pdf" } ], "resource_type": "article", "pub_year": "2014", "author_list": "Saraogi, Ishu; Akopian, David; et el." }, { "id": "https://authors.library.caltech.edu/records/fn0dz-hdj69", "eprint_id": 47517, "eprint_status": "archive", "datestamp": "2023-08-22 12:55:22", "lastmod": "2023-10-26 20:40:10", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Guo-Huan", "name": { "family": "Guo", "given": "Huan" } }, { "id": "Xiong-Yi", "name": { "family": "Xiong", "given": "Yi" } }, { "id": "Witkowski-P", "name": { "family": "Witkowski", "given": "Piotr" } }, { "id": "Cui-Jingqing", "name": { "family": "Cui", "given": "Jingqing" } }, { "id": "Wang-Ling-jia", "name": { "family": "Wang", "given": "Ling-jia" } }, { "id": "Sun-Jinhong", "name": { "family": "Sun", "given": "Jinhong" } }, { "id": "Lara-Lemus-R", "name": { "family": "Lara-Lemus", "given": "Roberto" } }, { "id": "Haataja-L", "name": { "family": "Haataja", "given": "Leena" } }, { "id": "Hutchison-K", "name": { "family": "Hutchison", "given": "Kathryn" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Arvan-P", "name": { "family": "Arvan", "given": "Peter" } }, { "id": "Liu-Ming", "name": { "family": "Liu", "given": "Ming" } } ] }, "title": "Inefficient Translocation of Preproinsulin Contributes to Pancreatic \u03b2 Cell Failure and Late-onset Diabetes", "ispublished": "pub", "full_text_status": "public", "keywords": "\u03b2 Cell; Diabetes; Insulin Synthesis; Mutant; Protein Translocation; Cytosolic Protein Accumulation; Preproinsulin; Proinsulin", "note": "\u00a9 2014 The American Society for Biochemistry and Molecular Biology, Inc.\n\nReceived for publication, March 2, 2014, and in revised form, March 26, 2014 Published, JBC Papers in Press, April 25, 2014.\n\nWe thank Bill and Dee Brehm for helping to establish the Brehm Center for Diabetes Research at the University of Michigan and Drs. Donald F. Steiner, Michael A. Weiss, Graeme Bell, and Yanzhuang Wang for encouragement and discussions. We also thank Dr. Martin Jendrisak and the entire team of the Gift of Hope Organ and Tissue Donor Network in Chicago for the human pancreas tissues used in this study. \n\nThis work was supported, in whole or in part, by National Institutes of Health Grants RO1-DK088856 (to M. L.), RO1-DK-48280 (to P. A.), RO1 GM078024 (to S. S.), and P60-DK-20572 (to the Molecular Biology and DNA Sequencing Core of the Diabetes Research and Training Center). This work was also supported by National Natural Science Foundation of China Research Grant 81070629 (to M. L.), by a research grant from the March of Dimes Foundation (to M. L.), and by the Protein Folding Disease Initiatives of the University of Michigan. Human pancreatic islet processing was supported by Illinois Department of Public Health Grant \"Pancreatic Islets Transplantation.\"\n\nPublished - J._Biol._Chem.-2014-Guo-16290-302.pdf
", "abstract": "Among the defects in the early events of insulin biosynthesis, proinsulin misfolding and endoplasmic reticulum (ER) stress have drawn increasing attention as causes of \u03b2 cell failure. However, no studies have yet addressed potential defects at the cytosolic entry point of preproinsulin into the secretory pathway. Here, we provide the first evidence that inefficient translocation of preproinsulin (caused by loss of a positive charge in the n region of its signal sequence) contributes to \u03b2 cell failure and diabetes. Specifically, we find that, after targeting to the ER membrane, preproinsulin signal peptide (SP) mutants associated with autosomal dominant late-onset diabetes fail to be fully translocated across the ER membrane. The newly synthesized, untranslocated preproinsulin remains strongly associated with the ER membrane, exposing its proinsulin moiety to the cytosol. Rather than accumulating in the ER and inducing ER stress, untranslocated preproinsulin accumulates in a juxtanuclear compartment distinct from the Golgi complex, induces the expression of heat shock protein 70 (HSP70), and promotes \u03b2 cell death. Restoring an N-terminal positive charge to the mutant preproinsulin SP significantly improves the translocation defect. These findings not only reveal a novel molecular pathogenesis of \u03b2 cell failure and diabetes but also provide the first evidence of the physiological and pathological significance of the SP n region positive charge of secretory proteins.", "date": "2014-06-06", "date_type": "published", "publication": "Journal of Biological Chemistry", "volume": "289", "number": "23", "publisher": "American Society for Biochemistry and Molecular Biology", "pagerange": "16290-16302", "id_number": "CaltechAUTHORS:20140728-091816472", "issn": "0021-9258", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140728-091816472", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "P30-DK020595" }, { "agency": "NIH", "grant_number": "RO1-DK088856" }, { "agency": "NIH", "grant_number": "RO1-DK-48280" }, { "agency": "NIH", "grant_number": "RO1-GM078024" }, { "agency": "NIH", "grant_number": "P60-DK-20572" }, { "agency": "National Nature Science Foundation of China", "grant_number": "81070629" }, { "agency": "March of Dimes Foundation" }, { "agency": "University of Michigan" }, { "agency": "Illinois Department of Public Health" } ] }, "doi": "10.1074/jbc.M114.562355", "pmcid": "PMC4047398", "primary_object": { "basename": "J._Biol._Chem.-2014-Guo-16290-302.pdf", "url": "https://authors.library.caltech.edu/records/fn0dz-hdj69/files/J._Biol._Chem.-2014-Guo-16290-302.pdf" }, "resource_type": "article", "pub_year": "2014", "author_list": "Guo, Huan; Xiong, Yi; et el." }, { "id": "https://authors.library.caltech.edu/records/j52np-qf218", "eprint_id": 45135, "eprint_status": "archive", "datestamp": "2023-08-20 00:31:54", "lastmod": "2023-10-26 17:35:14", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Gristick-H-B", "name": { "family": "Gristick", "given": "Harry B." } }, { "id": "Rao-M", "name": { "family": "Rao", "given": "Meera" } }, { "id": "Chartron-J-W", "name": { "family": "Chartron", "given": "Justin W." } }, { "id": "Rome-M-E", "name": { "family": "Rome", "given": "Michael E." } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Clemons-W-M-Jr", "name": { "family": "Clemons", "given": "William M., Jr." }, "orcid": "0000-0002-0021-889X" } ] }, "title": "Crystal structure of ATP-bound Get3\u2013Get4\u2013Get5 complex reveals regulation of Get3 by Get4", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2014 Macmillan Publishers Limited.\n\nReceived: 28 August 2013; Accepted: 24 March 2014;\nPublished online: 13 April 2014.\n\nWe are grateful to V. Denic (Harvard University) for providing the \u0394get3 \u0394get5 yeast strain. We thank G. Card, A. Gonzalez and M. Soltis for help with data collection at the Stanford Synchrotron Radiation Lightsource (SSRL) beamline 12-2. We are grateful to G. Moore and B. Moore for support of the Molecular Observatory at the California Institute of Technology. Operations at the SSRL are supported by the US Department of Energy and US National Institutes of Health (NIH). This work was supported by career awards from the David and Lucile Packard Foundation and the Henry Dreyfus Foundation (S.S.), US National Science Foundation Graduate Research Fellowship DGE-1144469 (M.E.R.), NIH training grant 5T32GM007616-33 (H.B.G. and M.R.) and NIH research grant R01GM097572 (W.M.C. Jr.).\n\nAuthor Contributions: H.B.G. performed all experiments except those noted. M.R. performed Get4 inhibition studies, and M.E.R. performed translocation experiments, both supervised by S.S. J.W.C. initiated the project and aided in refinement.\nS.S. contributed to experimental design and interpretation. W.M.C. Jr. conceived and supervised the project. H.B.G. and W.M.C. Jr. wrote the manuscript. All authors discussed the results and implications and commented on the manuscript.\n\nAccepted Version - nihms579145.pdf
Supplemental Material - nsmb.2813-S1.pdf
Supplemental Material - nsmb.2813-SF1.jpg
Supplemental Material - nsmb.2813-SF2.jpg
Supplemental Material - nsmb.2813-SF3.jpg
Supplemental Material - nsmb.2813-SF4.jpg
Supplemental Material - nsmb.2813-SF5.jpg
", "abstract": "Correct localization of membrane proteins is essential to all cells. Chaperone cascades coordinate the capture and handover of substrate proteins from the ribosomes to the target membranes, yet the mechanistic and structural details of these processes remain unclear. Here we investigate the conserved GET pathway, in which the Get4\u2013Get5 complex mediates the handover of tail-anchor (TA) substrates from the cochaperone Sgt2 to the Get3 ATPase, the central targeting factor. We present a crystal structure of a yeast Get3\u2013Get4\u2013Get5 complex in an ATP-bound state and show how Get4 primes Get3 by promoting the optimal configuration for substrate capture. Structure-guided biochemical analyses demonstrate that Get4-mediated regulation of ATP hydrolysis by Get3 is essential to efficient TA-protein targeting. Analogous regulation of other chaperones or targeting factors could provide a general mechanism for ensuring effective substrate capture during protein biogenesis.", "date": "2014-05", "date_type": "published", "publication": "Nature Structural & Molecular Biology", "volume": "21", "number": "5", "publisher": "Nature Publishing Group", "pagerange": "437-442", "id_number": "CaltechAUTHORS:20140423-093035077", "issn": "1545-9985", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140423-093035077", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "NSF Graduate Research Fellowship", "grant_number": "DGE-1144469" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "5T32GM007616-33" }, { "agency": "NIH", "grant_number": "R01GM097572" } ] }, "doi": "10.1038/nsmb.2813", "pmcid": "PMC4386898", "primary_object": { "basename": "nsmb.2813-SF1.jpg", "url": "https://authors.library.caltech.edu/records/j52np-qf218/files/nsmb.2813-SF1.jpg" }, "related_objects": [ { "basename": "nsmb.2813-SF2.jpg", "url": "https://authors.library.caltech.edu/records/j52np-qf218/files/nsmb.2813-SF2.jpg" }, { "basename": "nsmb.2813-SF3.jpg", "url": "https://authors.library.caltech.edu/records/j52np-qf218/files/nsmb.2813-SF3.jpg" }, { "basename": "nsmb.2813-SF4.jpg", "url": "https://authors.library.caltech.edu/records/j52np-qf218/files/nsmb.2813-SF4.jpg" }, { "basename": "nsmb.2813-SF5.jpg", "url": "https://authors.library.caltech.edu/records/j52np-qf218/files/nsmb.2813-SF5.jpg" }, { "basename": "nihms579145.pdf", "url": "https://authors.library.caltech.edu/records/j52np-qf218/files/nihms579145.pdf" }, { "basename": "nsmb.2813-S1.pdf", "url": "https://authors.library.caltech.edu/records/j52np-qf218/files/nsmb.2813-S1.pdf" } ], "resource_type": "article", "pub_year": "2014", "author_list": "Gristick, Harry B.; Rao, Meera; et el." }, { "id": "https://authors.library.caltech.edu/records/80rpe-vw885", "eprint_id": 46097, "eprint_status": "archive", "datestamp": "2023-08-20 00:46:15", "lastmod": "2023-10-26 18:41:22", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Zhang-Xin", "name": { "family": "Zhang", "given": "Xin" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Fidelity of Cotranslational Protein Targeting by the Signal Recognition Particle", "ispublished": "pub", "full_text_status": "public", "keywords": "protein biogenesis, signal sequence, GTPases, RNA, ribosome, protein\ntranslocation", "note": "\u00a9 2014 Annual Reviews. \n\nThe authors are not aware of any affiliations, memberships, funding, or financial holdings that might be perceived as affecting the objectivity of this review.\n\nAccepted Version - nihms689575.pdf
", "abstract": "Accurate folding, assembly, localization, and maturation of newly synthesized proteins are essential to all cells and require high fidelity in the protein biogenesis machineries that mediate these processes. Here, we review our current understanding of how high fidelity is achieved in one of these processes, the cotranslational targeting of nascent membrane and secretory proteins by the signal recognition particle (SRP). Recent biochemical, biophysical, and structural studies have elucidated how the correct substrates drive a series of elaborate conformational rearrangements in the SRP and SRP receptor GTPases; these rearrangements provide effective fidelity checkpoints to reject incorrect substrates and enhance the fidelity of this essential cellular pathway. The mechanisms used by SRP to ensure fidelity share important conceptual analogies with those used by cellular machineries involved in DNA replication, transcription, and translation, and these mechanisms likely represent general principles for other complex cellular pathways.", "date": "2014-05", "date_type": "published", "publication": "Annual Review of Biophysics", "volume": "43", "publisher": "Annual Reviews", "pagerange": "381-408", "id_number": "CaltechAUTHORS:20140605-104144841", "issn": "1936-122X", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140605-104144841", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1146/annurev-biophys-051013-022653", "pmcid": "PMC4444370", "primary_object": { "basename": "nihms689575.pdf", "url": "https://authors.library.caltech.edu/records/80rpe-vw885/files/nihms689575.pdf" }, "resource_type": "article", "pub_year": "2014", "author_list": "Zhang, Xin and Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/8xjd9-v1h23", "eprint_id": 54428, "eprint_status": "archive", "datestamp": "2023-08-20 00:16:29", "lastmod": "2023-10-20 16:26:51", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Losn-O", "name": { "family": "Losn", "given": "Oliver" } }, { "id": "Rome-M-E", "name": { "family": "Rome", "given": "Michael" } }, { "id": "Kaiser-J-T", "name": { "family": "Kaiser", "given": "Jens" }, "orcid": "0000-0002-5948-5212" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Chan-D-C", "name": { "family": "Chan", "given": "David" }, "orcid": "0000-0002-0191-2154" } ] }, "title": "Regulation of mitochondrial fission by MiD51", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2014 Federation of American Societies for Experimental Biology.", "abstract": "Mitochondrial fission requires the recruitment of dynamin-related protein 1 (Drp1) to mitochondria and activation of its GTP-dependent scission function. In mammals, the recruitment of Drp1 is primarily regulated by the outer membrane receptors Mff, MiD49 and MiD51, while Fis1 plays a minor role. We recently demonstrated that these receptors can function independently of one another to recruit Drp1 and mediate mitochondrial division. Intriguingly, exogenous expression of MiD49 and MiD51 causes constitutive recruitment of Drp1 but inhibits its scission activity. With mitochondrial stress, however, these receptors can rapidly activate pre-recruited Drp1 and cause rapid mitochondrial fragmentation independently of Fis1 and Mff. This behavior suggests an ability to sense mitochondrial function, but the molecular mechanism is unknown. To better understand the role of the MiDs in regulating mitochondrial fission, we produced recombinant protein pertaining to the cytosolic domain of each MiD, and performed crystallographic screens. We obtained crystals for MiD51 that diffracted to high resolution, and we were able to determine its structure. MiD51 possesses a nucleotidyl transferase fold and uses a variant set of residues to tightly bind a ligand. We have performed a structure-function analysis of MiD51 by mutating structurally implicated residues and studying their importance in vivo and in vitro to activate Drp1. Our analysis reveals how MiD51 activates mitochondrial fission during mitochondrial stress.", "date": "2014-04", "date_type": "published", "publication": "FASEB Journal", "volume": "28", "number": "1", "publisher": "Federation of American Societies for Experimental Biology", "pagerange": "757.1", "id_number": "CaltechAUTHORS:20150205-131139873", "issn": "0892-6638", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20150205-131139873", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "resource_type": "article", "pub_year": "2014", "author_list": "Losn, Oliver; Rome, Michael; et el." }, { "id": "https://authors.library.caltech.edu/records/bej14-qz924", "eprint_id": 54359, "eprint_status": "archive", "datestamp": "2023-08-20 00:15:49", "lastmod": "2023-10-20 16:21:03", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Gristick-H-B", "name": { "family": "Gristick", "given": "Harry" }, "orcid": "0000-0002-1957-2821" }, { "id": "Rao-Meera", "name": { "family": "Rao", "given": "Meera" } }, { "id": "Chartron-J-W", "name": { "family": "Chartron", "given": "Justin" } }, { "id": "Rome-M", "name": { "family": "Rome", "given": "Mike" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Clemons-W-M-Jr", "name": { "family": "Clemons", "given": "William M., Jr." }, "orcid": "0000-0002-0021-889X" } ] }, "title": "The structure of a tail-anchor membrane protein-binding complex reveals the regulation of Get3 by Get4", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2015 by the Federation of American Societies for Experimental Biology. April 2014. \n\nGrant Funding Source: NIH", "abstract": "Tail-anchored (TA) membrane proteins are targeted to the endoplasmic reticulum via the post-translational GET pathway. A critical step in this essential pathway is the transfer of TA substrates from the co-chaperone Sgt2 to the Get3 ATPase, mediated by the hetero-tetrameric complex Get4/Get5 (Get4/5). However, a major outstanding question is how Get4 binds to Get3 and regulates its activity. Here we report a 5.4 \u00c5 crystal structure of an ATP-bound Get3-Get4/5 complex from Saccharomyces cerevisiae (Sc), the largest pathway components solved to date. Our structure reveals the role of nucleotide in complex formation and the presence of two functionally distinct binding interfaces. Mutational analysis confirms that one interface is required for the specificity of binding, while the other interface is involved in the regulation of Get3 ATPase activity. Additional functional assays demonstrate that Get4/5-mediated regulation of ATP hydrolysis by Get3 is essential for efficient TA protein targeting. Our work also supports a model for Get3/TA complex formation in which two opposing Get3 dimers are bridged by a single Get4/5 hetero-tetramer. Finally, we present a 2.8 \u00c5 crystal structure of an apo ScGet3-Get4/5 complex that provides evidence for a rapid initial binding state mediated by electrostatics. This work illustrates how Get4/5 regulates Get3, priming it for TA loading, a critical step in this important pathway.", "date": "2014-04", "date_type": "published", "publication": "FASEB Journal", "volume": "28", "number": "1", "publisher": "Federation of American Societies for Experimental Biology", "pagerange": "Art. No. 950.4", "id_number": "CaltechAUTHORS:20150204-084954787", "issn": "0892-6638", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20150204-084954787", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH" } ] }, "resource_type": "article", "pub_year": "2014", "author_list": "Gristick, Harry; Rao, Meera; et el." }, { "id": "https://authors.library.caltech.edu/records/vfbc2-1p098", "eprint_id": 43840, "eprint_status": "archive", "datestamp": "2023-08-19 23:52:11", "lastmod": "2023-10-25 23:56:01", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Los\u00f3n-O-C", "name": { "family": "Los\u00f3n", "given": "Oliver C." } }, { "id": "Liu-Raymond", "name": { "family": "Liu", "given": "Raymond" } }, { "id": "Rome-M-E", "name": { "family": "Rome", "given": "Michael E." } }, { "id": "Meng-Shuxia", "name": { "family": "Meng", "given": "Shuxia" } }, { "id": "Kaiser-J-T", "name": { "family": "Kaiser", "given": "Jens T." }, "orcid": "0000-0002-5948-5212" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Chan-D-C", "name": { "family": "Chan", "given": "David C." }, "orcid": "0000-0002-0191-2154" } ] }, "title": "The Mitochondrial Fission Receptor MiD51 Requires ADP as a Cofactor", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2014 Elsevier Ltd.\n\nReceived: December 31, 2013; Revised: December 31, 2013\nAccepted: January 2, 2014; Published: February 6, 2014.\n\nSupplemental Information includes five figures and can be found with this article online at http://dx.doi.org/10.1016/j.str.2014.01.001.\n\nWe are grateful to Meera Rao for assistance with the GTPase assay and Alasdair McDowell for guidance with EM. We thank the Beckman Foundation at Caltech for support of the EM resource and the Gordon and Betty Moore Foundation and Augoron Institute for support of the Grant Jensen lab microscopy center. We acknowledge the Gordon and Betty Moore Foundation, the Beckman Institute, and the Sanofi-Aventis Bioengineering Research Program at Caltech for their generous support of the Molecular Observatory at Caltech. Operations at SSRL are supported by the US Department of Energy and the National Institutes of Health (NIH). This work was supported by a grant from the NIH (GM062967). O.C.L. was supported by an R. L. Kirschstein National Research Service Award (5F31GM089327) and an American Physiological Society William Townsend Porter predoctoral fellowship. \n\nPDB Accession Numbers: 4OAF (native structure), 4OAG (bound to ADP), 4OAH (H201A mutant structure), 4OAI (dimer mutant structure)\n\nAccepted Version - nihms576282.pdf
Supplemental Material - mmc1.pdf
", "abstract": "Mitochondrial fission requires recruitment of dynamin-\nrelated protein 1 (Drp1) to the mitochondrial surface\nand activation of its GTP-dependent scission\nfunction. The Drp1 receptors MiD49 and MiD51 recruit\nDrp1 to facilitate mitochondrial fission, but their\nmechanism of action is poorly understood. Using\nX-ray crystallography, we demonstrate that MiD51\ncontains a nucleotidyl transferase domain that binds\nADP with high affinity. MiD51 recruits Drp1 via a surface\nloop that functions independently of ADP binding.\nHowever, in the absence of nucleotide binding,\nthe recruited Drp1 cannot be activated for fission.\nPurified MiD51 strongly inhibits Drp1 assembly\nand GTP hydrolysis in the absence of ADP. Addition\nof ADP relieves this inhibition and promotes Drp1\nassembly into spirals with enhanced GTP hydrolysis.\nOur results reveal ADP as an essential cofactor for\nMiD51 during mitochondrial fission.", "date": "2014-03-04", "date_type": "published", "publication": "Structure", "volume": "22", "number": "3", "publisher": "Cell Press", "pagerange": "367-377", "id_number": "CaltechAUTHORS:20140214-120458864", "issn": "0969-2126", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140214-120458864", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Gordon and Betty Moore Foundation" }, { "agency": "Caltech Beckman Institute" }, { "agency": "Caltech Sanofi-Aventis Bioengineering Research Program" }, { "agency": "NIH", "grant_number": "GM062967" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "5F31GM089327" }, { "agency": "American Physiological Society William Townsend Porter Predoctoral Fellowship" } ] }, "doi": "10.1016/j.str.2014.01.001", "pmcid": "PMC4066849", "primary_object": { "basename": "mmc1.pdf", "url": "https://authors.library.caltech.edu/records/vfbc2-1p098/files/mmc1.pdf" }, "related_objects": [ { "basename": "nihms576282.pdf", "url": "https://authors.library.caltech.edu/records/vfbc2-1p098/files/nihms576282.pdf" } ], "resource_type": "article", "pub_year": "2014", "author_list": "Los\u00f3n, Oliver C.; Liu, Raymond; et el." }, { "id": "https://authors.library.caltech.edu/records/s4g06-dym72", "eprint_id": 43515, "eprint_status": "archive", "datestamp": "2023-08-22 11:09:44", "lastmod": "2023-10-25 23:37:11", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Wang-Yaqiang", "name": { "family": "Wang", "given": "Yaqiang" } }, { "id": "Fu-Yu-Hsien-Hwang", "name": { "family": "Fu", "given": "Yu-Hsien Hwang" } }, { "id": "Zhang-Qi", "name": { "family": "Zhang", "given": "Qi" }, "orcid": "0000-0002-5203-8778" }, { "id": "Feigon-Juli", "name": { "family": "Feigon", "given": "Juli" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Molecular Mechanism of GTPase Activation at the Signal Recognition Particle (SRP) RNA Distal End", "ispublished": "pub", "full_text_status": "public", "keywords": "G Proteins; Nuclear Magnetic Resonance; Protein Targeting; Protein-Nucleic Acid Interaction; RNA; Single Molecule Biophysics", "note": "\u00a9 2013 by The American Society for Biochemistry and Molecular Biology, Inc.\n\nReceived August 24, 2013. Revision received October 9, 2013. First Published on October 22, 2013.\n\nThis work was supported, in whole or in part, by National Institutes of Health Grants GM078024 (to S.-o.S.) and GM048123 (to J. F.) and a National Institutes of Health instrument supplement to Grant GM45162 (to D. C. Rees). This work was also supported by Caltech Matching Fund 350270 for the single molecule instruments, the David and Lucile Packard Fellowship in Science and Engineering (to S.-o.S.), and a Henry Dreyfus teacher-scholar award (to S.-o.S.).\n\nPublished - J._Biol._Chem.-2013-Shen-36385-97.pdf
Supplemental Material - jbc.M113.513614-1.docx
", "abstract": "The signal recognition particle (SRP) RNA is a universally conserved and essential component of the SRP that mediates the co-translational targeting of proteins to the correct cellular membrane. During the targeting reaction, two functional ends in the SRP RNA mediate distinct functions. Whereas the RNA tetraloop facilitates initial assembly of two GTPases between the SRP and SRP receptor, this GTPase complex subsequently relocalizes \u223c100 \u00c5 to the 5\u2032,3\u2032-distal end of the RNA, a conformation crucial for GTPase activation and cargo handover. Here we combined biochemical, single molecule, and NMR studies to investigate the molecular mechanism of this large scale conformational change. We show that two independent sites contribute to the interaction of the GTPase complex with the SRP RNA distal end. Loop E plays a crucial role in the precise positioning of the GTPase complex on these two sites by inducing a defined bend in the RNA helix and thus generating a preorganized recognition surface. GTPase docking can be uncoupled from its subsequent activation, which is mediated by conserved bases in the next internal loop. These results, combined with recent structural work, elucidate how the SRP RNA induces GTPase relocalization and activation at the end of the protein targeting reaction.", "date": "2013-12-20", "date_type": "published", "publication": "Journal of Biological Chemistry", "volume": "288", "number": "51", "publisher": "American Society for Biochemistry and Molecular Biology", "pagerange": "36385-36397", "id_number": "CaltechAUTHORS:20140124-141625084", "issn": "0021-9258", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140124-141625084", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "NIH", "grant_number": "GM048123" }, { "agency": "NIH", "grant_number": "GM45162" }, { "agency": "Caltech Matching Fund", "grant_number": "350270" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" } ] }, "doi": "10.1074/jbc.M113.513614", "pmcid": "PMC3868752", "primary_object": { "basename": "J._Biol._Chem.-2013-Shen-36385-97.pdf", "url": "https://authors.library.caltech.edu/records/s4g06-dym72/files/J._Biol._Chem.-2013-Shen-36385-97.pdf" }, "related_objects": [ { "basename": "jbc.M113.513614-1.docx", "url": "https://authors.library.caltech.edu/records/s4g06-dym72/files/jbc.M113.513614-1.docx" } ], "resource_type": "article", "pub_year": "2013", "author_list": "Shen, Kuang; Wang, Yaqiang; et el." }, { "id": "https://authors.library.caltech.edu/records/86zsy-tsg90", "eprint_id": 43131, "eprint_status": "archive", "datestamp": "2023-08-19 22:34:57", "lastmod": "2023-10-25 23:18:18", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Voigts-Hoffmann-F", "name": { "family": "Voigts-Hoffmann", "given": "Felix" } }, { "id": "Schmitz-N", "name": { "family": "Schmitz", "given": "Nikolaus" } }, { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Ataide-S-F", "name": { "family": "Ataide", "given": "Sandro F." } }, { "id": "Ban-Nenad", "name": { "family": "Ban", "given": "Nenad" }, "orcid": "0000-0002-9527-210X" } ] }, "title": "The Structural Basis of FtsY Recruitment and GTPase Activation by SRP RNA", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2013 Elsevier Inc. Received: March 5, 2013. Revised: August 27, 2013. Accepted: October 4, 2013. Published: November 7, 2013. We acknowledge support by the Swiss National Science Foundation (SNSF), the National Center of Excellence in Research (NCCR) Structural Biology program of the SNSF, and European Research Council grant 250071 under the European Community's Seventh Framework Programme. N.S. was supported by the Boehringer-Ingelheim Fonds. S.S. was supported by NIH grant R01GM078024 and the Packard and Lucile Award in Science and Engineering. We thank Daniel Boehringer and Marc Leibundgut for critical discussions of the manuscript and Stephan Imseng for contributions to the initial stages of the project.\n\nAccepted Version - nihms540393.pdf
Supplemental Material - mmc1.pdf
", "abstract": "The universally conserved signal recognition particle (SRP) system mediates the targeting of membrane proteins to the translocon in a multistep process controlled by GTP hydrolysis. Here we present the 2.6 \u00c5 crystal structure of the GTPase domains of the E. coli SRP protein (Ffh) and its receptor (FtsY) in complex with the tetraloop and the distal region of SRP-RNA, trapped in the activated state in presence of GDP:AlF_4. The structure reveals the atomic details of FtsY recruitment and, together with biochemical experiments, pinpoints G83 as the key RNA residue that stimulates GTP hydrolysis. Insertion of G83 into the FtsY active site orients a single glutamate residue provided by Ffh (E277), triggering GTP hydrolysis and complex disassembly at the end of the targeting cycle. The complete conservation of the key residues of the SRP-RNA and the SRP protein implies that the suggested chemical mechanism of GTPase activation is applicable across all kingdoms.", "date": "2013-12-12", "date_type": "published", "publication": "Molecular Cell", "volume": "52", "number": "5", "publisher": "Elsevier", "pagerange": "643-654", "id_number": "CaltechAUTHORS:20131223-080921779", "issn": "1097-2765", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20131223-080921779", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Swiss National Science Foundation (SNSF)" }, { "agency": "European Research Council (ERC)", "grant_number": "250071" }, { "agency": "Boehringer-Ingelheim Fonds" }, { "agency": "NIH", "grant_number": "R01 GM078024" }, { "agency": "David and Lucile Packard Foundation" } ] }, "doi": "10.1016/j.molcel.2013.10.005", "pmcid": "PMC3910249", "primary_object": { "basename": "mmc1.pdf", "url": "https://authors.library.caltech.edu/records/86zsy-tsg90/files/mmc1.pdf" }, "related_objects": [ { "basename": "nihms540393.pdf", "url": "https://authors.library.caltech.edu/records/86zsy-tsg90/files/nihms540393.pdf" } ], "resource_type": "article", "pub_year": "2013", "author_list": "Voigts-Hoffmann, Felix; Schmitz, Nikolaus; et el." }, { "id": "https://authors.library.caltech.edu/records/cwf8e-5yw82", "eprint_id": 40921, "eprint_status": "archive", "datestamp": "2023-08-22 09:31:46", "lastmod": "2023-10-24 23:18:17", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Akopian-D", "name": { "family": "Akopian", "given": "David" } }, { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Zhang-Xin", "name": { "family": "Zhang", "given": "Xin" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Signal Recognition Particle: An Essential Protein-Targeting Machine", "ispublished": "pub", "full_text_status": "public", "keywords": "GTPases; protein biogenesis; ribosome; signal recognition particle RNA; signal sequence", "note": "\u00a9 2013 Annual Reviews. \n\nFirst published online as a Review in Advance on February 13, 2013.\n\nWe are indebted to Sandra Schmid, Jennifer Doudna, Peter Walter, Douglas Rees, Raymond\nDeshaies, and William M. Clemons Jr. for support and insightful discussions over the years.\nS.S. was supported by NIH grant GM078024 and career awards from the Henry and Camille\nDreyfus Foundation, the Arnold and Mabel Beckman Foundation, and the David and Lucile\nPackard Foundation. D.A. was supported by NIH/NRSA training grant 5T32GM07616. X.Z.\nwas supported by a fellowship from the Ulric B. and Evelyn L. Bray Endowment Fund. X.Z.'s\ncurrent address is Department of Molecular and Experimental Medicine, The Scripps Research\nInstitute, La Jolla, California 92037.\n\nAccepted Version - nihms505529.pdf
", "abstract": "The signal recognition particle (SRP) and its receptor compose a universally\nconserved and essential cellular machinery that couples the synthesis\nof nascent proteins to their proper membrane localization. The\npast decade has witnessed an explosion in in-depth mechanistic investigations\nof this targeting machine at increasingly higher resolutions. In\nthis review, we summarize recent work that elucidates how the SRP and\nSRP receptor interact with the cargo protein and the target membrane,\nrespectively, and how these interactions are coupled to a novel GTPase\ncycle in the SRP\u00b7SRP receptor complex to provide the driving force\nand enhance the fidelity of this fundamental cellular pathway. We also\ndiscuss emerging frontiers in which important questions remain to be\naddressed.", "date": "2013-06", "date_type": "published", "publication": "Annual Review of Biochemistry", "volume": "82", "publisher": "Annual Reviews", "pagerange": "693-721", "id_number": "CaltechAUTHORS:20130826-105419836", "issn": "0066-4154", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130826-105419836", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "5T32GM07616" }, { "agency": "Ulric B. and Evelyn L. Bray Endowment Fund" } ] }, "doi": "10.1146/annurev-biochem-072711-164732", "pmcid": "PMC3805129", "primary_object": { "basename": "nihms505529.pdf", "url": "https://authors.library.caltech.edu/records/cwf8e-5yw82/files/nihms505529.pdf" }, "resource_type": "article", "pub_year": "2013", "author_list": "Akopian, David; Shen, Kuang; et el." }, { "id": "https://authors.library.caltech.edu/records/ne124-mtx73", "eprint_id": 39016, "eprint_status": "archive", "datestamp": "2023-08-19 19:55:22", "lastmod": "2023-10-24 14:58:09", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Nguyen-Thang-X", "name": { "family": "Nguyen", "given": "Thang X." } }, { "id": "Jaru-Ampornpan-P", "name": { "family": "Jaru-Ampornpan", "given": "Peera" } }, { "id": "Lam-Vinh-Q", "name": { "family": "Lam", "given": "Vinh Q." } }, { "id": "Cao-Peigen", "name": { "family": "Cao", "given": "Peigen" } }, { "id": "Piszkiewicz-S", "name": { "family": "Piszkiewicz", "given": "Samantha" } }, { "id": "Hess-S", "name": { "family": "Hess", "given": "Sonja" }, "orcid": "0000-0002-5904-9816" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Mechanism of an ATP-independent Protein Disaggregase - I. Structure of a Membrane Protein Aggregate Reveals a Mechanism of Recognition by its Chaperone", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2013 by The American Society for Biochemistry and Molecular Biology, Inc.\nReceived for publication, February 18, 2013, and in revised form, March 13, 2013 Published, JBC Papers in Press, March 22, 2013.\nSupported by a grant from the Betty and Gordon Moore Foundation.\nSupported by the David and Lucile Packard Fellowship in science and engineering,\nthe Henry Dreyfus Teacher-Scholar Award, and the Breakthroughs\nin Gerontology award from the American Federation for Aging\nResearch.\nWe thank A. N. Murray and Dr. J. W. Kelly for\nrecrystallized ThT and fibrilized A\u03b2_(1\u201340), A. McDowall for EM analyses,\nmembers of the Proteome Exploration Laboratory for mass spectrometry\nassistance, and members of the Shan group for helpful comments\non the manuscript. The Proteome Exploration Laboratory is\nsupported by the Betty and Gordon Moore Foundation and the Beckman\nInstitute. AFM studies were supported by United States Department\nof Energy Grant DE-FG03-01ER46175 (to J. Heath).\n\nPublished - J._Biol._Chem.-2013-Nguyen-13420-30.pdf
Supplemental Material - jbc.M113.462812-1.docx
", "abstract": "Protein aggregation is detrimental to the maintenance of proper protein homeostasis in all cells. To overcome this problem, cells have evolved a network of molecular chaperones to prevent protein aggregation and even reverse existing protein aggregates. The most extensively studied disaggregase systems are ATP-driven macromolecular machines. Recently, we reported an alternative disaggregase system in which the 38-kDa subunit of chloroplast signal recognition particle (cpSRP43) efficiently reverses the aggregation of its substrates, the light-harvesting chlorophyll a/b-binding (LHC) proteins, in the absence of external energy input. To understand the molecular mechanism of this novel activity, here we used biophysical and biochemical methods to characterize the structure and nature of LHC protein aggregates. We show that LHC proteins form micellar, disc-shaped aggregates that are kinetically stable and detergent-resistant. Despite the nonamyloidal nature, the LHC aggregates have a defined global organization, displaying the chaperone recognition motif on its solvent-accessible surface. These findings suggest an attractive mechanism for recognition of the LHC aggregate by cpSRP43 and provide important constraints to define the capability of this chaperone.", "date": "2013-05-10", "date_type": "published", "publication": "Journal of Biological Chemistry", "volume": "288", "number": "19", "publisher": "American Society for Biochemistry and Molecular Biology", "pagerange": "13420-13430", "id_number": "CaltechAUTHORS:20130621-085950473", "issn": "0021-9258", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130621-085950473", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Gordon and Betty Moore Foundation" }, { "agency": "Caltech Beckman Institute" }, { "agency": "Department of Energy (DOE)", "grant_number": "DE-FG03-01ER46175" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "American Federation for Aging Research" } ] }, "doi": "10.1074/jbc.M113.462812", "pmcid": "PMC3650380", "primary_object": { "basename": "J._Biol._Chem.-2013-Nguyen-13420-30.pdf", "url": "https://authors.library.caltech.edu/records/ne124-mtx73/files/J._Biol._Chem.-2013-Nguyen-13420-30.pdf" }, "related_objects": [ { "basename": "jbc.M113.462812-1.docx", "url": "https://authors.library.caltech.edu/records/ne124-mtx73/files/jbc.M113.462812-1.docx" } ], "resource_type": "article", "pub_year": "2013", "author_list": "Nguyen, Thang X.; Jaru-Ampornpan, Peera; et el." }, { "id": "https://authors.library.caltech.edu/records/fg5n4-8zh83", "eprint_id": 39030, "eprint_status": "archive", "datestamp": "2023-08-22 09:20:48", "lastmod": "2023-10-24 14:59:13", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Jaru-Ampornpan-P", "name": { "family": "Jaru-Ampornpan", "given": "Peera" } }, { "id": "Liang-Fu-Cheng", "name": { "family": "Liang", "given": "Fu-Cheng" } }, { "id": "Nisthal-A", "name": { "family": "Nisthal", "given": "Alex" } }, { "id": "Nguyen-Thang-X", "name": { "family": "Nguyen", "given": "Thang X." } }, { "id": "Wang-Pengcheng", "name": { "family": "Wang", "given": "Pengcheng" } }, { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Mayo-S-L", "name": { "family": "Mayo", "given": "Steven L." }, "orcid": "0000-0002-9785-5018" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Mechanism of an ATP-independent Protein Disaggregase. II. Distinct Molecular Interactions Drive Multiple Steps During Aggregate Disassembly", "ispublished": "pub", "full_text_status": "public", "keywords": "Enzyme Mechanisms; Kinetics; Molecular Chaperone; Mutagenesis Mechanisms; Protein Aggregation; ATP-independent Disaggregase; Membrane Proteins; Protein Biogenesis; Signal Recognition Particle.", "note": "\u00a9 2013 by The American Society for Biochemistry and Molecular Biology, Inc. \n\nReceived February 18, 2013; Revision received March 12, 2013. \n\nSupported by the Department of Defense, National Security Science and Engineering Faculty Fellowship. \n\nSupported by the David and Lucile Packard Fellowship in science and engineering, the Henry Dreyfus Teacher-Scholar Award, and the Breakthroughs in Gerontology award from the American Federation for Aging Research.\n\nWe thank Drs. W. M. Clemons, J. Chartron, and C. Suloway for the plasmids of SERP1, Sec61b_s, and cytochrome b5 and the Shan laboratory for helpful comments on the manuscript.\n\nPublished - J._Biol._Chem.-2013-Jaru-Ampornpan-13431-45.pdf
", "abstract": "The ability of molecular chaperones to overcome the misfolding and aggregation of proteins is essential for the maintenance of proper protein homeostasis in all cells. Thus far, the best studied disaggregase systems are the Clp/Hsp100 family of \"ATPases associated with various cellular activities\" (AAA^+) ATPases, which use mechanical forces powered by ATP hydrolysis to remodel protein aggregates. An alternative system to disassemble large protein aggregates is provided by the 38-kDa subunit of the chloroplast signal recognition particle (cpSRP43), which uses binding energy with its substrate proteins to drive disaggregation. The mechanism of this novel chaperone remains unclear. Here, molecular genetics and structure-activity analyses show that the action of cpSRP43 can be dissected into two steps with distinct molecular requirements: (i) initial recognition, during which cpSRP43 binds specifically to a recognition motif displayed on the surface of the aggregate; and (ii) aggregate remodeling, during which highly adaptable binding interactions of cpSRP43 with hydrophobic transmembrane domains of the substrate protein compete with the packing interactions within the aggregate. This establishes a useful framework to understand the molecular mechanism by which binding interactions from a molecular chaperone can be used to overcome protein aggregates in the absence of external energy input from ATP.", "date": "2013-05-10", "date_type": "published", "publication": "Journal of Biological Chemistry", "volume": "288", "number": "19", "publisher": "American Society for Biochemistry and Molecular Biology", "pagerange": "13431-13445", "id_number": "CaltechAUTHORS:20130621-133910862", "issn": "0021-9258", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130621-133910862", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "National Security Science and Engineering Faculty Fellowship" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "American Federation for Aging Research" } ] }, "doi": "10.1074/jbc.M113.462861", "pmcid": "PMC3650381", "primary_object": { "basename": "J._Biol._Chem.-2013-Jaru-Ampornpan-13431-45.pdf", "url": "https://authors.library.caltech.edu/records/fg5n4-8zh83/files/J._Biol._Chem.-2013-Jaru-Ampornpan-13431-45.pdf" }, "resource_type": "article", "pub_year": "2013", "author_list": "Jaru-Ampornpan, Peera; Liang, Fu-Cheng; et el." }, { "id": "https://authors.library.caltech.edu/records/e1ew6-rg373", "eprint_id": 39135, "eprint_status": "archive", "datestamp": "2023-08-22 09:18:52", "lastmod": "2023-10-24 16:27:39", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Rome-M-E", "name": { "family": "Rome", "given": "Michael E." } }, { "id": "Rao-M", "name": { "family": "Rao", "given": "Meera" } }, { "id": "Clemons-W-M-Jr", "name": { "family": "Clemons", "given": "William M." }, "orcid": "0000-0002-0021-889X" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Precise timing of ATPase activation drives targeting of tail-anchored proteins", "ispublished": "pub", "full_text_status": "public", "keywords": "allostery; GTPases; mechanistic enzymology; translocation", "note": "\u00a9 2013 National Academy of Sciences.\n\nEdited by Douglas C. Rees, Howard Hughes Medical Institute, Caltech, Pasadena, CA, and approved March 26, 2013 (received for review December 19, 2012).\nPublished online before print April 22, 2013.\n\n\nWe thank J. Chartron, H. Gristick, and C. J. M.\nSuloway for expression constructs, purification protocols, and critical\ndiscussions; M. Sachs and C. Wu for help with yeast translation extracts;\nR. Schekman for help with yeast microsomes; and D. C. Rees and members\nof the S.-o.S. and W.M.C. groups for helpful comments. This work was\nsupported by career awards from the David and Lucile Packard Foundation\nand the Henry Dreyfus Foundation (to S.-o.S.), National Science Foundation\nGraduate Research Fellowship DGE-1144469 (to M.E.R.), National Institutes\nof Health (NIH) Training Grant 5T32GM007616-33 (to M.R.), and NIH Grant\nR01 GM097572 (to W.M.C.).\n\n\nAuthor contributions: M.E.R., M.R., and S.-o.S. designed research; M.E.R. and M.R. performed\nresearch; W.M.C. contributed new reagents/analytic tools; M.E.R., M.R., and S.-o.S.\nanalyzed data; and M.E.R., M.R., and S.-o.S. wrote the paper.\n\nPublished - PNAS-2013-Rome-7666-71.pdf
Supplemental Material - pnas.201222054SI.pdf
", "abstract": "The localization of tail-anchored (TA) proteins, whose transmembrane domain resides at the extreme C terminus, presents major challenges to cellular protein targeting machineries. In eukaryotic cells, the highly conserved ATPase, guided entry of tail-anchored protein 3 (Get3), coordinates the delivery of TA proteins to the endoplasmic reticulum. How Get3 uses its ATPase cycle to drive this fundamental process remains unclear. Here, we establish a quantitative framework for the Get3 ATPase cycle and show that ATP specifically induces multiple conformational changes in Get3 that culminate in its ATPase activation through tetramerization. Further, upstream and downstream components actively regulate the Get3 ATPase cycle to ensure the precise timing of ATP hydrolysis in the pathway: the Get4/5 TA loading complex locks Get3 in the ATP-bound state and primes it for TA protein capture, whereas the TA substrate induces tetramerization of Get3 and activates its ATPase reaction 100-fold. Our results establish a precise model for how Get3 harnesses the energy from ATP to drive the membrane localization of TA proteins and illustrate how dimerization-activated nucleotide hydrolases regulate diverse cellular processes.", "date": "2013-05-07", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "110", "number": "19", "publisher": "National Academy of Sciences", "pagerange": "7666-7671", "id_number": "CaltechAUTHORS:20130628-090524393", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130628-090524393", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "NSF Graduate Research Fellowship", "grant_number": "DGE-1144469" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "5T32GM007616-33" }, { "agency": "NIH", "grant_number": "R01 GM097572" } ] }, "doi": "10.1073/pnas.1222054110", "pmcid": "PMC3651441", "primary_object": { "basename": "PNAS-2013-Rome-7666-71.pdf", "url": "https://authors.library.caltech.edu/records/e1ew6-rg373/files/PNAS-2013-Rome-7666-71.pdf" }, "related_objects": [ { "basename": "pnas.201222054SI.pdf", "url": "https://authors.library.caltech.edu/records/e1ew6-rg373/files/pnas.201222054SI.pdf" } ], "resource_type": "article", "pub_year": "2013", "author_list": "Rome, Michael E.; Rao, Meera; et el." }, { "id": "https://authors.library.caltech.edu/records/gvbcy-72522", "eprint_id": 39075, "eprint_status": "archive", "datestamp": "2023-08-19 19:42:51", "lastmod": "2023-10-24 16:22:10", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "von-Loeffelholz-O", "name": { "family": "von Loeffelholz", "given": "Ottilie" } }, { "id": "Knoops-K", "name": { "family": "Knoops", "given": "K\u00e8vin" } }, { "id": "Ariosa-A-R", "name": { "family": "Ariosa", "given": "Aileen" } }, { "id": "Zhang-Xin", "name": { "family": "Zhang", "given": "Xin" } }, { "id": "Karuppasamy-M", "name": { "family": "Karuppasamy", "given": "Manikandan" } }, { "id": "Huard-K", "name": { "family": "Huard", "given": "Karine" } }, { "id": "Schoehn-G", "name": { "family": "Schoehn", "given": "Guy" } }, { "id": "Berger-I", "name": { "family": "Berger", "given": "Imre" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Schaffitzel-C", "name": { "family": "Schaffitzel", "given": "Christiane" } } ] }, "title": "Structural basis of signal sequence surveillance and selection by the SRP\u2013FtsY complex", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2013 Nature Publishing Group, a division of Macmillan Publishers Limited. \n\nReceived 8 November 2012; accepted 21 February 2013; published online 7 April 2013.\n\nWe thank T. Shaikh for advice with Spider refinement, P. Penczek for assistance with SPARX, M. Bacia for excellent technical assistance and members of the protein expression facility at European Molecular Biology Laboratory Heidelberg as well as the Partnership for Structural Biology in Grenoble for support. The Polara microscope is part of the Structural Biology and Dynamics Groupement d'int\u00e9r\u00eat scientifique\u2013Infrastrutures en Biologie Sante et Agronomie platform of the Institut de Biologie Structurale. C.S. acknowledges support by the Agence Nationale de la Recherche (ANR-09-JCJC-0044), the region Rh\u00f4ne-Alpes\n(CIBLE_1976) and the European Research Council Starting grant (project 281331). K.K. was supported by a postdoctoral European Molecular Biology Organization fellowship. We thank I. Saraogi in the Shan laboratory for sharing unpublished results and for critical reading of the manuscript. S.S. is supported by US National Institutes of Health grant R01 GM078024, and the Fellowship for science and engineering from the David and Lucile Packard foundation. A.A. was supported by the US National Institute of General Medical Sciences Ruth L. Kirschstein National Research Service Award (F31GM095294) and the National Institutes of Health National Research Service Award Training grant 5T32GM07616. X.Z. is supported by the Howard Hughes Medical Institute Fellowship of the Helen Hay Whitney Foundation.\n\nAuthor Contributions:\n\nC.S., I.B., X.Z. and S.S. designed experiments; C.S., K.H., O.v.L., A.A. and X.Z. prepared samples; A.A. and X.Z. carried out biochemical experiments; K.K., G.S. and M.K. performed the electron microscopy; O.v.L., M.K. and C.S. performed image analysis and model building; C.S., O.v.L., A.A., X.Z. and S.S. prepared\nthe manuscript.\n\nAccepted Version - nihms505534.pdf
Supplemental Material - nsmb.2546-S1.pdf
", "abstract": "Signal-recognition particle (SRP)-dependent targeting of translating ribosomes to membranes is a multistep quality-control process. Ribosomes that are translating weakly hydrophobic signal sequences can be rejected from the targeting reaction even after they are bound to the SRP. Here we show that the early complex, formed by Escherichia coli SRP and its receptor FtsY with ribosomes translating the incorrect cargo EspP, is unstable and rearranges inefficiently into subsequent conformational states, such that FtsY dissociation is favored over successful targeting. The N-terminal extension of EspP is responsible for these defects in the early targeting complex. The cryo-electron microscopy structure of this 'false' early complex with EspP revealed an ordered M domain of SRP protein Ffh making two ribosomal contacts, and the NG domains of Ffh and FtsY forming a distorted, flexible heterodimer. Our results provide a structural basis for SRP-mediated signal-sequence selection during recruitment of the SRP receptor.", "date": "2013-05", "date_type": "published", "publication": "Nature Structural & Molecular Biology", "volume": "20", "number": "5", "publisher": "Nature Publishing Group", "pagerange": "604-610", "id_number": "CaltechAUTHORS:20130625-102457328", "issn": "1545-9985", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130625-102457328", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Agence Nationale de la Recherche (ANR)", "grant_number": "ANR-09-JCJC-0044" }, { "agency": "Region Rh\u00f4ne-Alpes", "grant_number": "CIBLE_1976" }, { "agency": "European Research Council (ERC)", "grant_number": "281331" }, { "agency": "European Molecular Biology Organization (EMBO)" }, { "agency": "NIH", "grant_number": "R01 GM078024" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "NIH Postdoctoral Fellowship", "grant_number": "F31GM095294" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "5T32GM07616" }, { "agency": "Howard Hughes Medical Institute (HHMI)" }, { "agency": "Helen Hay Whitney Foundation" } ] }, "doi": "10.1038/nsmb.2546", "pmcid": "PMC3874396", "primary_object": { "basename": "nsmb.2546-S1.pdf", "url": "https://authors.library.caltech.edu/records/gvbcy-72522/files/nsmb.2546-S1.pdf" }, "related_objects": [ { "basename": "nihms505534.pdf", "url": "https://authors.library.caltech.edu/records/gvbcy-72522/files/nihms505534.pdf" } ], "resource_type": "article", "pub_year": "2013", "author_list": "von Loeffelholz, Ottilie; Knoops, K\u00e8vin; et el." }, { "id": "https://authors.library.caltech.edu/records/900ta-xz941", "eprint_id": 39320, "eprint_status": "archive", "datestamp": "2023-08-19 19:17:56", "lastmod": "2023-10-24 16:40:51", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Arslan-S", "name": { "family": "Arslan", "given": "Sinan" } }, { "id": "Akopian-D", "name": { "family": "Akopian", "given": "David" } }, { "id": "Ha-Taekjip", "name": { "family": "Ha", "given": "Taekjip" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Activated GTPase movement on an RNA scaffold drives cotranslational protein targeting", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2013 FASEB.", "abstract": "Signal Recognition Particle (SRP) and its receptor (SR) are GTPases that translocate\nribosome-nascent chain complexes (RNCs) from cytosol to cellular membranes.\nSRP recognizes the RNC, and through interactions with SR, brings the RNC to the\nmembrane. Subsequent rearrangements activate GTP hydrolysis and load the RNC\nonto the translocon. During this process, the SRP RNA plays an indispensible role\nin stimulating both SRP-SR complex formation and the following GTP hydrolysis.\nPrevious studies hinted that the SRP RNA is a bi-functional molecule with two\ndistinct functional ends, the tetraloop end and the distal end, that each stimulates\nthe SRP-SR complex formation and GTP hydrolysis steps. Here we used single\nmolecule technique to directly visualize the global rearrangement of the GTPase\ncomplex from the RNA's tetraloop end to the distal end, traveling over 100\nAngstrom in the targeting reaction. Moreover, we showed that the rearrangement\nis tightly controlled by the RNC and the translocon. This large-scale movement of\nthe GTPase complex provides a molecular mechanism of coupling GTPase\nactivation to the transfer of RNC, thereby ensuring productive protein targeting.", "date": "2013-04", "date_type": "published", "publication": "FASEB Journal", "volume": "27", "publisher": "Federation of American Societies for Experimental Biology", "pagerange": "Art. No. 556.3", "id_number": "CaltechAUTHORS:20130711-133423197", "issn": "0892-6638", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130711-133423197", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "resource_type": "article", "pub_year": "2013", "author_list": "Shen, Kuang; Arslan, Sinan; et el." }, { "id": "https://authors.library.caltech.edu/records/dt67m-q4c35", "eprint_id": 39321, "eprint_status": "archive", "datestamp": "2023-08-19 19:18:02", "lastmod": "2023-10-24 16:40:56", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "GTPase and ATPase tangos during intracellular protein targeting", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2013 FASEB.", "abstract": "Rougly one third of the proteome are destined for the cellular membrane, whose\nproper localization is essential for the structure and function of all cells. The signal\nrecognition particle (SRP) is a universally conserved cellular machinery that couples\nthe synthesis of membrane and secretory proteins to their proper cellular\nlocalization, and has served as a paradigm to understand the molecular basis of\nprotein localization. Using a combination of chemical, biophysical, structural and\ncellular approaches, our work has established a quantitative framework for how\ntwo highly homologous GTPases in the SRP and SRP receptor use a novel GTPase\ncycle to drive this fundamental cellular pathway, and elucidated how fidelity of\nprotein localization by the SRP is achieved through a combination of binding,\ninduced fit, and kinetic proofreading mechanisms. These findings also define a\nnovel class of 'dimerization-activated' GTPases, represented by the SRP and SRP\nreceptor, whose regulatory principles may extend to a growing number of\nnucleotide hydrolases that drive diverse cellular processes.", "date": "2013-04", "date_type": "published", "publication": "FASEB Journal", "volume": "27", "publisher": "Federation of American Societies for Experimental Biology", "pagerange": "Art. No. 198.1", "id_number": "CaltechAUTHORS:20130711-133855161", "issn": "0892-6638", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130711-133855161", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "resource_type": "article", "pub_year": "2013", "author_list": "Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/e121d-8e263", "eprint_id": 39310, "eprint_status": "archive", "datestamp": "2023-08-19 19:17:38", "lastmod": "2023-10-24 16:40:11", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Rome-M-E", "name": { "family": "Rome", "given": "Michael" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "The ATPase Cycle of the Tail-Anchored Protein Chaperone Get3", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2013 FASEB.", "abstract": "The cytosolic ATPase Get3/TRC40 mediates targeting of tail-anchored (TA)\nmembrane proteins to the endoplasmic reticulum (ER). Get3 functions as a\nmolecular chaperone by binding to transmembrane domains of newly synthesized\nTA proteins and delivering them to a membrane docking complex in the ER.\nPrevious work have shown that ATP binding and hydrolysis can regulate Get3s\nfunction and are essential for the targeting of TA proteins. However, the precise\nmechanisms by which this is accomplished remain unclear. Using a combination of\nmechanistic enzymology and biophysical methods, we delineate Get3s ATPase\ncycle and show that it is required at multiple stages during TA protein targeting.\nOur data complements previous structural studies and contributes to our\nunderstanding of how Get3 harnesses the energy of ATP hydrolysis to spatially and\ntemporally coordinate the delivery of TA proteins to the ER membrane.", "date": "2013-04", "date_type": "published", "publication": "FASEB Journal", "volume": "27", "publisher": "Federation of American Societies for Experimental Biology", "pagerange": "Art. No. 542.4", "id_number": "CaltechAUTHORS:20130711-103448215", "issn": "0892-6638", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130711-103448215", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "resource_type": "article", "pub_year": "2013", "author_list": "Rome, Michael and Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/9seds-wpp69", "eprint_id": 38552, "eprint_status": "archive", "datestamp": "2023-08-19 19:08:24", "lastmod": "2023-10-23 20:34:22", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Pierce-N-W", "name": { "family": "Pierce", "given": "Nathan W." } }, { "id": "Lee-J-Eugene", "name": { "family": "Lee", "given": "J. Eugene" } }, { "id": "Liu-Xing", "name": { "family": "Liu", "given": "Xing" }, "orcid": "0000-0002-0506-6821" }, { "id": "Sweredoski-M-J", "name": { "family": "Sweredoski", "given": "Michael J." }, "orcid": "0000-0003-0878-3831" }, { "id": "Graham-R-L-J", "name": { "family": "Graham", "given": "Robert L. J." } }, { "id": "Larimore-E-A", "name": { "family": "Larimore", "given": "Elizabeth A." } }, { "id": "Rome-M-E", "name": { "family": "Rome", "given": "Michael" } }, { "id": "Zheng-Ning", "name": { "family": "Zheng", "given": "Ning" } }, { "id": "Clurman-B-E", "name": { "family": "Clurman", "given": "Bruce E." } }, { "id": "Hess-S", "name": { "family": "Hess", "given": "Sonja" }, "orcid": "0000-0002-5904-9816" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Deshaies-R-J", "name": { "family": "Deshaies", "given": "Raymond J." }, "orcid": "0000-0002-3671-9354" } ] }, "title": "Cand1 Promotes Assembly of New SCF Complexes through Dynamic Exchange of F Box Proteins", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2013 Elsevier Inc. \n\nReceived: December 10, 2012. Revised: January 24, 2013. Accepted: February 12, 2013. Published: February 28, 2013. \n\nWe thank J. Vielmetter of the Caltech Protein Expression Facility for providing Fbxw7-Skp1, b-TrCP-Skp1, and ubiquitin E1. We thank B. Schulman, L. Busino, M. Pagano, F. Bassermann, O. Schneewind, A. Saha, and W. den Besten for gifts of reagents. We thank G. Smith for assistance with mass spectrometry analyses. We thank all the members of the Deshaies and Shan lab for support and helpful discussions. We thank D. Wolf and T. Kurz for communicating unpublished results. N.W.P. was supported by the Gordon Ross Fellowship, and an NIH Training Grant. J.E.L. was supported by the Ruth L. Kirschstein NRSA Fellowship (CA138126) from the NIH. R.J.D. is an Investigator of the HHMI. This work was supported in part by NIH GM065997 to R.J.D. The Proteome Exploration lab is supported in part by grants from the Gordon and Betty Moore Foundation and the Beckman Institute and an instrumentation grant from NIH (10565784).\n\nAccepted Version - nihms463502.pdf
Supplemental Material - mmc1.xlsx
Supplemental Material - mmc2.xlsx
Supplemental Material - mmc3.xlsx
Supplemental Material - mmc4.xlsx
Supplemental Material - mmc5.xlsx
Supplemental Material - mmc6.pdf
", "abstract": "The modular SCF (Skp1, cullin, and F box) ubiquitin ligases feature a large family of F box protein substrate receptors that enable recognition of diverse targets. However, how the repertoire of SCF complexes is sustained remains unclear. Real-time measurements of formation and disassembly indicate that SCF^(Fbxw7) is extraordinarily stable, but, in the Nedd8-deconjugated state, the cullin-binding protein Cand1 augments its dissociation by one-million-fold. Binding and ubiquitylation assays show that Cand1 is a protein exchange factor that accelerates the rate at which Cul1-Rbx1 equilibrates with multiple F box protein-Skp1 modules. Depletion of Cand1 from cells impedes recruitment of new F box proteins to pre-existing Cul1 and profoundly alters the cellular landscape of SCF complexes. We suggest that catalyzed protein exchange may be a general feature of dynamic macromolecular machines and propose a hypothesis for how substrates, Nedd8, and Cand1 collaborate to regulate the cellular repertoire of SCF complexes.", "date": "2013-03-28", "date_type": "published", "publication": "Cell", "volume": "153", "number": "1", "publisher": "Elsevier", "pagerange": "206-215", "id_number": "CaltechAUTHORS:20130517-081333288", "issn": "0092-8674", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130517-081333288", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Gordon Ross Fellowship" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "CA138126" }, { "agency": "NIH", "grant_number": "GM065997" }, { "agency": "Gordon and Betty Moore Foundation" }, { "agency": "Caltech Beckman Institute" }, { "agency": "NIH", "grant_number": "10565784" } ] }, "doi": "10.1016/j.cell.2013.02.024", "pmcid": "PMC3656483", "primary_object": { "basename": "mmc4.xlsx", "url": "https://authors.library.caltech.edu/records/9seds-wpp69/files/mmc4.xlsx" }, "related_objects": [ { "basename": "mmc5.xlsx", "url": "https://authors.library.caltech.edu/records/9seds-wpp69/files/mmc5.xlsx" }, { "basename": "mmc6.pdf", "url": "https://authors.library.caltech.edu/records/9seds-wpp69/files/mmc6.pdf" }, { "basename": "nihms463502.pdf", "url": "https://authors.library.caltech.edu/records/9seds-wpp69/files/nihms463502.pdf" }, { "basename": "mmc1.xlsx", "url": "https://authors.library.caltech.edu/records/9seds-wpp69/files/mmc1.xlsx" }, { "basename": "mmc2.xlsx", "url": "https://authors.library.caltech.edu/records/9seds-wpp69/files/mmc2.xlsx" }, { "basename": "mmc3.xlsx", "url": "https://authors.library.caltech.edu/records/9seds-wpp69/files/mmc3.xlsx" } ], "resource_type": "article", "pub_year": "2013", "author_list": "Pierce, Nathan W.; Lee, J. Eugene; et el." }, { "id": "https://authors.library.caltech.edu/records/x6zts-gwn66", "eprint_id": 37578, "eprint_status": "archive", "datestamp": "2023-08-19 14:43:52", "lastmod": "2023-10-23 17:48:47", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Akopian-D", "name": { "family": "Akopian", "given": "David" } }, { "id": "Dalal-Kush", "name": { "family": "Dalal", "given": "Kush" } }, { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Duong-Franck", "name": { "family": "Duong", "given": "Franck" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "SecYEG activates GTPases to drive the completion of cotranslational protein targeting", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2013 Akopian et al. This article is distributed under the terms of an Attribution\u2013Noncommercial\u2013Share Alike\u2013No Mirror Sites license for the first six months after the publication\ndate (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution\u2013Noncommercial\u2013Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/). \n\nSubmitted: 8 August 2012; accepted: 16 January 2013.\n\nWe thank Oded Lewinson for help with liposome experiments, Ishu Saraogi for help in cotranslational targeting assays and valuable discussions, and members of the Shan group for comments on the manuscript. This work was supported by National Institutes of Health (NIH) grant GM078024, and career awards from the Henry and Camille Dreyfus foundation\nand the David and Lucile Packard foundation to S. Shan. D. Akopian was supported by NIH/National Research Service Award training grant 5T32GM07616.\n\nPublished - J_Cell_Biol-2013-Akopian-397-405.pdf
", "abstract": "Signal recognition particle (SRP) and its receptor (SR) comprise a highly conserved cellular machine that cotranslationally targets proteins to a protein-conducting channel, the bacterial SecYEG or eukaryotic Sec61p complex, at the target membrane. Whether SecYEG is a passive recipient of the translating ribosome or actively regulates this targeting machinery remains unclear. Here we show that SecYEG drives conformational changes in the cargo-loaded SRP\u2013SR targeting complex that activate it for GTP hydrolysis and for handover of the translating ribosome. These results provide the first evidence that SecYEG actively drives the efficient delivery and unloading of translating ribosomes at the target membrane.", "date": "2013-02-18", "date_type": "published", "publication": "Journal of Cell Biology", "volume": "200", "number": "4", "publisher": "Rockeller University Press", "pagerange": "397-405", "id_number": "CaltechAUTHORS:20130320-140650051", "issn": "0021-9525", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130320-140650051", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "5T32GM07616" } ] }, "doi": "10.1083/jcb.201208045", "pmcid": "PMC3575545", "primary_object": { "basename": "J_Cell_Biol-2013-Akopian-397-405.pdf", "url": "https://authors.library.caltech.edu/records/x6zts-gwn66/files/J_Cell_Biol-2013-Akopian-397-405.pdf" }, "resource_type": "article", "pub_year": "2013", "author_list": "Akopian, David; Dalal, Kush; et el." }, { "id": "https://authors.library.caltech.edu/records/z85dx-06z52", "eprint_id": 38146, "eprint_status": "archive", "datestamp": "2023-08-19 14:28:19", "lastmod": "2023-10-23 19:51:50", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Arslan-S", "name": { "family": "Arslan", "given": "Sinan" } }, { "id": "Akopian-D", "name": { "family": "Akopian", "given": "David" } }, { "id": "Ha-Taekjip", "name": { "family": "Ha", "given": "Taekjip" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Activated GTPase Movement on SRP RNA Drives Cotranslational Protein Targeting", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2013 Biophysical Society. Published by Elsevier Inc.\n\n Available online 29 January 2013.\n 2152-Pos B171", "abstract": "Signal Recognition Particle (SRP) and its receptor (SR) are co-translational protein targeting machineries responsible for delivering ribosome-nascent chain-complexes (RNCs) from cytosol to cellular membranes. Loaded with its cargo (RNC), SRP forms a complex with SR and bring the RNC to the membrane. Extensive rearrangements in this complex activate GTP hydrolysis and unload the RNC onto the translocon. During this process, the SRP RNA plays an essential role in accelerating both SRP-SR complex formation and GTP hydrolysis step. Here we show that the SRP RNA is a bi-functional molecule with its two ends, the tetraloop end and the distal end, stimulating different stages of SRP-SR interaction. using single molecule techniques to direct visualize the global relocalization along the SRP RNA, we demonstrate that the SRP-SR GTPase complex travels over 100A in the targeting reaction, from the RNA's tetraloop end during initial complex assembly to the distal end during GTPase activation. Moreover, this rearrangement is tightly regulated by the RNC and the translocon. The large-scale movement of the GTPase complex provides an attractive mechanism for coupling GTPase activation to the transfer of RNC from SRP to translocon, thereby ensuring productive protein targeting.", "date": "2013-01-29", "date_type": "published", "publication": "Biophysical Journal", "volume": "104", "number": "2", "publisher": "Biophysical Society", "pagerange": "419A", "id_number": "CaltechAUTHORS:20130429-084737150", "issn": "0006-3495", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130429-084737150", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1016/j.bpj.2012.11.2334", "resource_type": "article", "pub_year": "2013", "author_list": "Shen, Kuang; Arslan, Sinan; et el." }, { "id": "https://authors.library.caltech.edu/records/6fcv2-c8j07", "eprint_id": 38197, "eprint_status": "archive", "datestamp": "2023-08-19 14:28:32", "lastmod": "2023-10-23 19:55:21", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Rao-M", "name": { "family": "Rao", "given": "Meera" } }, { "id": "Rome-M-E", "name": { "family": "Rome", "given": "Michael E." } }, { "id": "Clemons-W-M-Jr", "name": { "family": "Clemons", "given": "William M." }, "orcid": "0000-0002-0021-889X" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Precise Timing of ATPase Activation Drives Targeting of Tail-Anchored Proteins", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2013 Biophysical Society. Published by Elsevier Inc.\n\nPublished - 1-s2.0-S0006349512044244-main.pdf
", "abstract": "Hundreds of proteins are anchored on cellular membranes by a transmembrane domain (TM) at their extreme C-terminus. These 'tail-anchored' (TA) proteins perform numerous essential cellular functions, yet their unique topology poses fundamental challenges to their proper localization. In eukaryotic cells, the highly conserved ATPase Get3 coordinates the efficient capture and delivery of TA proteins to the endoplasmic reticulum (ER). How Get3 uses its ATPase cycles to drive this fundamental process remains elusive. Here we define the Get3 ATPase cycle and show that ATP specifically induces conformational changes in Get3 that culminate in its ATPase activation through tetramerization. This activation is inhibited by the TA protein loading complex Get4/5 and is activated by the TA substrate, ensuring the precise timing of ATP hydrolysis during targeting. Our results provide an explicit model for how Get3 harnesses the energy from ATP to spatially and temporally coordinate the membrane localization of TA proteins.", "date": "2013-01-29", "date_type": "published", "publication": "Biophysical Journal", "volume": "104", "number": "2", "publisher": "Biophysical Society", "pagerange": "572A", "id_number": "CaltechAUTHORS:20130501-084952283", "issn": "0006-3495", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130501-084952283", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1016/j.bpj.2012.11.3178", "primary_object": { "basename": "1-s2.0-S0006349512044244-main.pdf", "url": "https://authors.library.caltech.edu/records/6fcv2-c8j07/files/1-s2.0-S0006349512044244-main.pdf" }, "resource_type": "article", "pub_year": "2013", "author_list": "Rao, Meera; Rome, Michael E.; et el." }, { "id": "https://authors.library.caltech.edu/records/53sh0-0cj59", "eprint_id": 39488, "eprint_status": "archive", "datestamp": "2023-08-19 14:22:04", "lastmod": "2023-10-24 16:51:32", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Ariosa-Aileen-R", "name": { "family": "Ariosa", "given": "Aileen R." } }, { "id": "Duncan-Stacy-S", "name": { "family": "Duncan", "given": "Stacy S." } }, { "id": "Saraogi-Ishu", "name": { "family": "Saraogi", "given": "Ishu" } }, { "id": "Lu-Xiaodong", "name": { "family": "Lu", "given": "Xiaodong" } }, { "id": "Brown-April", "name": { "family": "Brown", "given": "April" }, "orcid": "0000-0003-2631-3905" }, { "id": "Phillips-Gregory-J", "name": { "family": "Phillips", "given": "Gregory J." } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Fingerloop activates cargo delivery and unloading during cotranslational protein targeting", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2013 Ariosa et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution\u2013Noncommercial\u2013Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).\n\nReceived: Jun 12, 2012; Revised: Oct 25, 2012; Accepted: Nov 1, 2012.\n\nWe thank members of the Shan group for helpful comments on the manuscript. This work was supported by National Institutes of Health (NIH) grant GM078024, the Beckman Young Investigator Award, the David and Lucile Packard Fellowship in Science and Engineering, and the Henry Dreyfus Teacher-Scholar Award to S.-O.S. G.J.P. was supported by NIH grant R01 GM069628. A.R.A. was supported by NIH/NRSA training grant 5T32GM07616 and by the Betty and Gordon Moore Foundation.\n\nPublished - Mol._Biol._Cell-2013-Ariosa-63-73.pdf
Supplemental Material - CombinedSupMats.pdf
", "abstract": "During cotranslational protein targeting by the signal recognition particle (SRP), information about signal sequence binding in the SRP's M domain must be effectively communicated to its GTPase domain to turn on its interaction with the SRP receptor (SR) and thus deliver the cargo proteins to the membrane. A universally conserved \"fingerloop\" lines the signal sequence\u2013binding groove of SRP; the precise role of this fingerloop in protein targeting has remained elusive. In this study, we show that the fingerloop plays important roles in SRP function by helping to induce the SRP into a more active conformation that facilitates multiple steps in the pathway, including efficient recruitment of SR, GTPase activation in the SRP\u2022SR complex, and most significantly, the unloading of cargo onto the target membrane. On the basis of these results and recent structural work, we propose that the fingerloop is the first structural element to detect signal sequence binding; this information is relayed to the linker connecting the SRP's M and G domains and thus activates the SRP and SR for carrying out downstream steps in the pathway.", "date": "2013-01-15", "date_type": "published", "publication": "Molecular Biology of the Cell", "volume": "24", "number": "2", "publisher": "American Society for Cell Biology", "pagerange": "63-73", "id_number": "CaltechAUTHORS:20130722-092857106", "issn": "1059-1524", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130722-092857106", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "NIH", "grant_number": "R01 GM069628" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "5T32GM07616" }, { "agency": "Gordon and Betty Moore Foundation" } ] }, "doi": "10.1091/mbc.E12-06-0434", "pmcid": "PMC3541965", "primary_object": { "basename": "CombinedSupMats.pdf", "url": "https://authors.library.caltech.edu/records/53sh0-0cj59/files/CombinedSupMats.pdf" }, "related_objects": [ { "basename": "Mol._Biol._Cell-2013-Ariosa-63-73.pdf", "url": "https://authors.library.caltech.edu/records/53sh0-0cj59/files/Mol._Biol._Cell-2013-Ariosa-63-73.pdf" } ], "resource_type": "article", "pub_year": "2013", "author_list": "Ariosa, Aileen R.; Duncan, Stacy S.; et el." }, { "id": "https://authors.library.caltech.edu/records/t4k5m-s5529", "eprint_id": 36150, "eprint_status": "archive", "datestamp": "2023-08-22 07:56:35", "lastmod": "2023-10-20 22:18:06", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Arslan-S", "name": { "family": "Arslan", "given": "Sinan" } }, { "id": "Akopian-D", "name": { "family": "Akopian", "given": "David" } }, { "id": "Ha-Taekjip", "name": { "family": "Ha", "given": "Taekjip" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Activated GTPase movement on an RNA scaffold drives co-translational protein targeting", "ispublished": "pub", "full_text_status": "public", "keywords": "Enzyme mechanisms; Protein translocation; Single-molecule biophysics; RNA", "note": "\u00a9 2012 Macmillan Publishers Limited.\n\nReceived 31 July; accepted 26 October 2012.\nPublished online 12 December 2012.\nWe thank N. Ban and members of the Shan group for comments\non the manuscript, C. Richards, L. Cai, T. Zhiyentayev, K. Lee and R. Zhou for help with RNC labelling and the instrument and software setup, and C. L. Guo, S. Kouand H. Lester for discussions. This work is supported by National Institutes of Health (NIH) grant GM078024 to S.-o.S., an NIH instrument supplement to grant GM45162 to D.C. Rees,\nand Caltech matching fund 350270 for the single-molecule instruments. S.-o.S. was supported by the Beckman Young Investigator award, the David and Lucile Packard\nFellowship in Science and Engineering, and the Henry Dreyfus Teacher-Scholar award. T. H. was supported by National Science FoundationPhysics Frontiers Centers program\n(08222613) and NIH grant GM065367.\nAuthor Contributions: K.S., S.A., T.H. and S.-o.S. conceived the experiments. K.S. purified and labelled Ffh, FtsY, DNA, RNA and RNC. D.A. purified SecYEG and performed the GTPase assay in Fig. 4a. K.S. and S.A. carried out smFRET\nmeasurements under the direction of T.H. K.S. and S.A. analysed the data. K.S. and S.-o.S. wrote the paper with inputs from all other authors.\n\nAccepted Version - nihms418620.pdf
Supplemental Material - nature11726-s1.pdf
", "abstract": "Approximately one-third of the proteome is initially destined for the eukaryotic endoplasmic reticulum or the bacterial plasma membrane. The proper localization of these proteins is mediated by a universally conserved protein-targeting machinery, the signal recognition particle (SRP), which recognizes ribosomes carrying signal sequences and, through interactions with the SRP receptor delivers them to the protein-translocation machinery on the target membrane. The SRP is an ancient ribonucleoprotein particle containing an essential, elongated SRP RNA for which precise functions have remained elusive. Here we used single-molecule fluorescence microscopy to show that the Escherichia coli SRP\u2013SRP receptor GTPase complex, after initial assembly at the tetraloop end of SRP RNA, travels over 100\u2009\u00c5 to the distal end of this RNA, where rapid GTP hydrolysis occurs. This movement is negatively regulated by the translating ribosome and, at a later stage, positively regulated by the SecYEG translocon, providing an attractive mechanism for ensuring the productive exchange of the targeting and translocation machineries at the ribosome exit site with high spatial and temporal accuracy. Our results show that large RNAs can act as molecular scaffolds that enable the easy exchange of distinct factors and precise timing of molecular events in a complex cellular process; this concept may be extended to similar phenomena in other ribonucleoprotein complexes.", "date": "2012-12-13", "date_type": "published", "publication": "Nature", "volume": "492", "number": "7428", "publisher": "Nature Publishing Group", "pagerange": "271-275", "id_number": "CaltechAUTHORS:20130103-133200806", "issn": "0028-0836", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130103-133200806", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "NIH", "grant_number": "GM45162" }, { "agency": "Caltech Matching Fund", "grant_number": "350270" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "NSF", "grant_number": "PHY-08222613" }, { "agency": "NIH", "grant_number": "GM065367" } ] }, "doi": "10.1038/nature11726", "pmcid": "PMC3531814", "primary_object": { "basename": "nature11726-s1.pdf", "url": "https://authors.library.caltech.edu/records/t4k5m-s5529/files/nature11726-s1.pdf" }, "related_objects": [ { "basename": "nihms418620.pdf", "url": "https://authors.library.caltech.edu/records/t4k5m-s5529/files/nihms418620.pdf" } ], "resource_type": "article", "pub_year": "2012", "author_list": "Shen, Kuang; Arslan, Sinan; et el." }, { "id": "https://authors.library.caltech.edu/records/s1d7w-v7j98", "eprint_id": 31266, "eprint_status": "archive", "datestamp": "2023-08-19 10:26:57", "lastmod": "2023-10-17 15:43:20", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Liu-Ming", "name": { "family": "Liu", "given": "Ming" } }, { "id": "Lara-Lemus-R", "name": { "family": "Lara-Lemus", "given": "Roberto" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Wright-Jordan", "name": { "family": "Wright", "given": "Jordan" } }, { "id": "Haataja-L", "name": { "family": "Haataja", "given": "Leena" } }, { "id": "Barbetti-F", "name": { "family": "Barbetti", "given": "Fabrizio" } }, { "id": "Guo-Huan", "name": { "family": "Guo", "given": "Huan" } }, { "id": "Larkin-Dennis", "name": { "family": "Larkin", "given": "Dennis" } }, { "id": "Arvan-P", "name": { "family": "Arvan", "given": "Peter" } } ] }, "title": "Impaired Cleavage of Preproinsulin Signal Peptide Linked to Autosomal-Dominant Diabetes", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2012 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered. See http://creativecommons.org/licenses/by-nc-nd/3.0/ for details. \n\nReceived June 24, 2011. Accepted December 19, 2011. \n\nThis work was supported primarily by National Institutes of Health (NIH) grants R01-DK-48280 (to P.A.) and R01-DK-088856 and by March of Dimes 6-FY11-357 (to M.L.) as well as National Nature Science Foundation of China Grant 81070629 (to M.L.). We also acknowledge assistance from the Molecular Biology and DNA Sequencing Core of the NIH-funded Diabetes Research and Training Center (P60-DK-20572). \n\nNo potential conflicts of interest relevant to this article were reported. \n\nAuthor Contributions: M.L., R.L.-L., J.W., L.H., H.G., and D.L. researched data. M.L., S.-o.S., J.W., L.H., F.B., and P.A. contributed to discussion. M.L. and P.A. wrote the manuscript. M.L., J.W., F.B., and P.A. reviewed and edited the manuscript. P.A. is the guarantor of this work and, as such, had full access to all of the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis. \n\nThe authors thank Bill and Dee Brehm for helping to create the Brehm Center for Diabetes Research to advance diabetes research at the University of Michigan.\n\nPublished - Liu2012p17855Diabetes.pdf
", "abstract": "Recently, missense mutations upstream of preproinsulin's signal peptide (SP) cleavage site were reported to cause mutant INS gene-induced diabetes of youth (MIDY). Our objective was to understand the molecular pathogenesis using metabolic labeling and assays of proinsulin export and insulin and C-peptide production to examine the earliest events of insulin biosynthesis, highlighting molecular mechanisms underlying \u03b2-cell failure plus a novel strategy that might ameliorate the MIDY syndrome. We find that whereas preproinsulin-A(SP23)S is efficiently cleaved, producing authentic proinsulin and insulin, preproinsulin-A(SP24)D is inefficiently cleaved at an improper site, producing two subpopulations of molecules. Both show impaired oxidative folding and are retained in the endoplasmic reticulum (ER). Preproinsulin-A(SP24)D also blocks ER exit of coexpressed wild-type proinsulin, accounting for its dominant-negative behavior. Upon increased expression of ER\u2013oxidoreductin-1, preproinsulin-A(SP24)D remains blocked but oxidative folding of wild-type proinsulin improves, accelerating its ER export and increasing wild-type insulin production. We conclude that the efficiency of SP cleavage is linked to the oxidation of (pre)proinsulin. In turn, impaired (pre)proinsulin oxidation affects ER export of the mutant as well as that of coexpressed wild-type proinsulin. Improving oxidative folding of wild-type proinsulin may provide a feasible way to rescue insulin production in patients with MIDY.", "date": "2012-04", "date_type": "published", "publication": "Diabetes", "volume": "61", "number": "4", "publisher": "American Diabetes Association", "pagerange": "828-837", "id_number": "CaltechAUTHORS:20120502-085929510", "issn": "0012-1797", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20120502-085929510", "rights": "Readers may use this article as\nlong as the work is properly cited, the use is educational and not for profit,\nand the work is not altered. See http://creativecommons.org/licenses/by\n-nc-nd/3.0/ for details.", "funders": { "items": [ { "agency": "NIH", "grant_number": "R01-DK-48280" }, { "agency": "NIH", "grant_number": "R01-DK-088856" }, { "agency": "March of Dimes", "grant_number": "6-FY11-357" }, { "agency": "National Nature Science Foundation of China", "grant_number": "81070629" }, { "agency": "NIH", "grant_number": "P60-DK-20572" } ] }, "doi": "10.2337/db11-0878", "pmcid": "PMC3314357", "primary_object": { "basename": "Liu2012p17855Diabetes.pdf", "url": "https://authors.library.caltech.edu/records/s1d7w-v7j98/files/Liu2012p17855Diabetes.pdf" }, "resource_type": "article", "pub_year": "2012", "author_list": "Liu, Ming; Lara-Lemus, Roberto; et el." }, { "id": "https://authors.library.caltech.edu/records/42cxe-ewa08", "eprint_id": 36084, "eprint_status": "archive", "datestamp": "2023-08-19 10:33:13", "lastmod": "2023-10-20 22:13:29", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Lu-Zeyu-Mike", "name": { "family": "Lu", "given": "Zeyu Mike" } }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "The cpSRP54-cpFtsY Interaction in the Chloroplast SRP Pathway", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2012 FASEB.", "abstract": "The universally conserved signal recognition particle (SRP) and SRP receptor (SR) mediate the cotranslational targeting of proteins to cellular membranes. In contrast, a recently discovered unique chloroplast SRP in green plants is primarily dedicated to the post-translational targeting of light harvesting chlorophyll a/b binding (LHC) proteins to the thylakoid membranes. The pathway involves cpSRP54 and its receptor cpFtsY, two GTPases that are similar to the cytosolic SRP54 and SR GTPases. Intriguingly, the otherwise universally conserved SRP RNA which accelerates complex formation between the two GTPases has not been found in the chloroplast pathway. Instead, a new 43-kDa protein, cpSRP43, forms the chloroplast SRP together with cpSRP54. Kinetic analyses have shown that cpSRP43 does not significantly affect the rate of the cpSRP54\u00b7cpFtsY complex formation or the GTP hydrolysis rate of the complex. Instead, cpSRP43 facilitates the substrate recognition through interactions with the cargo proteins, LHCPs. This project seeks to understand how cpSRP54 and cpFtsY are able to associate more efficiently than their bacterial homologues in the absence of the SRP RNA by studying their complex structure using crystallography and lysine footprinting mass spectrometry. To date, 2-dimensional crystal plates have been obtained, and progress continues toward the determination of a cpSRP54\u00b7cpFtsY structure.", "date": "2012-04", "date_type": "published", "publication": "FASEB Journal", "volume": "26", "publisher": "Federation of American Societies for Experimental Biology", "pagerange": "Art. No. 973.3", "id_number": "CaltechAUTHORS:20121220-150130013", "issn": "0892-6638", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20121220-150130013", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "resource_type": "article", "pub_year": "2012", "author_list": "Lu, Zeyu Mike; Chandrasekar, Sowmya; et el." }, { "id": "https://authors.library.caltech.edu/records/hmwqm-zs530", "eprint_id": 29966, "eprint_status": "archive", "datestamp": "2023-08-19 10:12:17", "lastmod": "2023-10-24 22:37:54", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Zhang-Dawei", "name": { "family": "Zhang", "given": "Dawei" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Translation Elongation Regulates Substrate Selection by the Signal Recognition Particle", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2012 by The American Society for Biochemistry and Molecular Biology, Inc. Received for publication, November 17, 2011, and in revised form, January 2, 2012. Published, JBC Papers in Press, January 6, 2012. This work was supported, in whole or in part, by National Institutes of Health Grant GM078024 (to S. S.). \nThis article contains supplemental Figs. S1\u2013S3.\nSupported by career awards from the Beckman Young Investigator Award, the Packard and Lucile Award in Science and Engineering, and the Henry Dreyfus Teacher-Scholar Award. \n\nWe thank R. Hegde for RRL and microsomal membranes, H. Bernstein for the strain HDB52, J. Beckwith for antibodies, and X. Zhang and members of the Shan group for helpful comments on the manuscript.\n\nPublished - Zhang2012p17580J_Biol_Chem.pdf
Supplemental Material - jbc.M111.325001-1.doc
", "abstract": "The signal recognition particle (SRP) is a universally conserved cellular machinery responsible for delivering membrane and secretory proteins to the proper cellular destination. The precise mechanism by which fidelity is achieved by the SRP pathway within the in vivo environment is yet to be understood. Previous studies have focused on the SRP pathway in isolation. Here we describe another important factor that modulates substrate selection by the SRP pathway: the ongoing synthesis of the nascent polypeptide chain by the ribosome. A slower translation elongation rate rescues the targeting defect of substrate proteins bearing mutant, suboptimal signal sequences both in vitro and in vivo. Consistent with a kinetic origin of this effect, similar rescue of protein targeting was also observed with mutant SRP receptors or SRP RNAs that specifically compromise the kinetics of SRP-receptor interaction during protein targeting. These data are consistent with a model in which ongoing protein translation is in constant kinetic competition with the targeting of the nascent proteins by the SRP and provides an important factor to regulate the fidelity of substrate selection by the SRP.", "date": "2012-03-02", "date_type": "published", "publication": "Journal of Biological Chemistry", "volume": "287", "number": "10", "publisher": "American Society for Biochemistry and Molecular Biology", "pagerange": "7652-7660", "id_number": "CaltechAUTHORS:20120404-070455923", "issn": "0021-9258", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20120404-070455923", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "NIH", "grant_number": "GM078024" } ] }, "doi": "10.1074/jbc.M111.325001", "pmcid": "PMC3293578", "primary_object": { "basename": "Zhang2012p17580J_Biol_Chem.pdf", "url": "https://authors.library.caltech.edu/records/hmwqm-zs530/files/Zhang2012p17580J_Biol_Chem.pdf" }, "related_objects": [ { "basename": "jbc.M111.325001-1.doc", "url": "https://authors.library.caltech.edu/records/hmwqm-zs530/files/jbc.M111.325001-1.doc" } ], "resource_type": "article", "pub_year": "2012", "author_list": "Zhang, Dawei and Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/qk2kb-hkw10", "eprint_id": 29582, "eprint_status": "archive", "datestamp": "2023-08-19 09:46:14", "lastmod": "2023-10-24 22:18:59", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Zhang-Dawei", "name": { "family": "Zhang", "given": "Dawei" } }, { "id": "Sweredoski-M-J", "name": { "family": "Sweredoski", "given": "Michael J." }, "orcid": "0000-0003-0878-3831" }, { "id": "Graham-R-L-J", "name": { "family": "Graham", "given": "Robert L. J." } }, { "id": "Hess-S", "name": { "family": "Hess", "given": "Sonja" }, "orcid": "0000-0002-5904-9816" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Novel Proteomic Tools Reveal Essential Roles of SRP and Importance of Proper Membrane Protein Biogenesis", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2012 by The American Society for Biochemistry and Molecular Biology, Inc. Received May 27, 2011, and in revised form, September 24, 2011. Published, MCP Papers in Press, October 26, 2011. This work was supported by National Institutes of Health grant\nGM078024 to S.S. S.S. was supported by career awards from the\nBurroughs Welcome Foundation, a Beckman Young Investigator\naward, a Packard and Lucile award in science and engineering, and a\nHenry Dreyfus teacher-scholar award. S. H., R. L. J. G. and M. S. were supported by the Betty and Gordon Moore Foundation and the Beckman Institute.\n\nPublished - Zhang2012p17341Mol_Cell_Proteomics.pdf
Supplemental Material - mcp.M111.011585-1.doc
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Supplemental Material - mcp.M111.011585-2.doc
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Supplemental Material - mcp.M111.011585-7.pdf
Supplemental Material - mcp.M111.011585-8.xlsx
Supplemental Material - mcp.M111.011585-9.xlsx
", "abstract": "The signal recognition particle (SRP), which mediates cotranslational protein targeting to cellular membranes, is universally conserved and essential for bacterial and mammalian cells. However, the current understanding of the role of SRP in cell physiology and pathology is still poor, and the reasons behind its essential role in cell survival remain unclear. Here, we systematically analyzed the consequences of SRP loss in E. coli using time-resolved quantitative proteomic analyses. A series of snapshots of the steady-state and newly synthesized proteome unveiled three stages of cellular responses to SRP depletion, and demonstrated essential roles of SRP in metabolism, membrane potential, and protein and energy homeostasis in both the membrane and cytoplasm. We also identified a group of periplasmic proteins, including key molecular chaperones, whose localization was impaired by the loss of SRP; this and additional results showed that SRP is crucial for protein homeostasis in the bacterial envelope. These results reveal the extensive roles that SRP plays in bacterial physiology, emphasize the importance of proper membrane protein biogenesis, and demonstrate the ability of time-resolved quantitative proteomic analysis to provide new biological insights.", "date": "2012-02", "date_type": "published", "publication": "Molecular and Cellular Proteomics", "volume": "11", "number": "2", "publisher": "American Society for Biochemistry and Molecular Biology", "pagerange": "Art. No. M111.011585", "id_number": "CaltechAUTHORS:20120306-072520214", "issn": "1535-9476", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20120306-072520214", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Burroughs Welcome Foundation" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "Gordon and Betty Moore Foundation" }, { "agency": "Caltech Beckman Institute" } ] }, "doi": "10.1074/mcp.M111.011585", "pmcid": "PMC3277757", "primary_object": { "basename": "mcp.M111.011585-1.doc", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-1.doc" }, "related_objects": [ { "basename": "mcp.M111.011585-4.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-4.xlsx" }, { "basename": "mcp.M111.011585-11.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-11.xlsx" }, { "basename": "mcp.M111.011585-12.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-12.xlsx" }, { "basename": "mcp.M111.011585-16.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-16.xlsx" }, { "basename": "mcp.M111.011585-19.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-19.xlsx" }, { "basename": "mcp.M111.011585-21.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-21.xlsx" }, { "basename": "mcp.M111.011585-5.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-5.xlsx" }, { "basename": "mcp.M111.011585-3.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-3.xlsx" }, { "basename": "mcp.M111.011585-6.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-6.xlsx" }, { "basename": "Zhang2012p17341Mol_Cell_Proteomics.pdf", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/Zhang2012p17341Mol_Cell_Proteomics.pdf" }, { "basename": "mcp.M111.011585-10.pdf", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-10.pdf" }, { "basename": "mcp.M111.011585-15.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-15.xlsx" }, { "basename": "mcp.M111.011585-17.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-17.xlsx" }, { "basename": "mcp.M111.011585-18.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-18.xlsx" }, { "basename": "mcp.M111.011585-22.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-22.xlsx" }, { "basename": "mcp.M111.011585-9.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-9.xlsx" }, { "basename": "mcp.M111.011585-13.doc", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-13.doc" }, { "basename": "mcp.M111.011585-14.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-14.xlsx" }, { "basename": "mcp.M111.011585-2.doc", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-2.doc" }, { "basename": "mcp.M111.011585-20.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-20.xlsx" }, { "basename": "mcp.M111.011585-7.pdf", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-7.pdf" }, { "basename": "mcp.M111.011585-8.xlsx", "url": "https://authors.library.caltech.edu/records/qk2kb-hkw10/files/mcp.M111.011585-8.xlsx" } ], "resource_type": "article", "pub_year": "2012", "author_list": "Zhang, Dawei; Sweredoski, Michael J.; et el." }, { "id": "https://authors.library.caltech.edu/records/93n0n-rft26", "eprint_id": 27796, "eprint_status": "archive", "datestamp": "2023-08-22 04:01:18", "lastmod": "2023-10-24 17:25:57", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Saraogi-Ishu", "name": { "family": "Saraogi", "given": "Ishu" } }, { "id": "Akopian-D", "name": { "family": "Akopian", "given": "David" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "A tale of two GTPases in cotranslational protein targeting", "ispublished": "pub", "full_text_status": "restricted", "keywords": "GTPases; molecular recognition and regulation; signal recognition particle; protein targeting and translocation; protein interaction dynamics", "note": "\u00a9 2011 The Protein Society. Published by Wiley-Blackwell. \n\nReceived 11 August 2011; Revised 12 August 2011; Accepted 15 August 2011. Published online 6 September 2011. \n\nWe thank Shen K for comments on the manuscript. This work was supported by NIH grant GM078024, and career awards from the Burroughs Welcome Foundation, the Henry and Camille Dreyfus foundation, the Arnold and Mabel Beckman foundation, and the David and Lucile Packard foundation to S.S. D. A. was supported by NIH/NRSA training grant 5T32GM07616.", "abstract": "Guanosine triphosphatases (GTPases) comprise a superfamily of proteins that provide molecular switches to regulate numerous cellular processes. The \"GTPase switch\" paradigm, in which a GTPase acts as a bimodal switch that is turned \"on\" and \"off\" by external regulatory factors, has been used to interpret the regulatory mechanism of many GTPases. Recent work on a pair of GTPases in the signal recognition particle (SRP) pathway has revealed a distinct mode of GTPase regulation. Instead of the classical GTPase switch, the two GTPases in the SRP and SRP receptor undergo a series of conformational changes during their dimerization and reciprocal activation. Each conformational rearrangement provides a point at which these GTPases can communicate with and respond to their upstream and downstream biological cues, thus ensuring the spatial and temporal precision of all the molecular events in the SRP pathway. We suggest that the SRP and SRP receptor represent an emerging class of \"multistate\" regulatory GTPases uniquely suited to provide exquisite control over complex cellular pathways that require multiple molecular events to occur in a highly coordinated fashion.", "date": "2011-11", "date_type": "published", "publication": "Protein Science", "volume": "20", "number": "11", "publisher": "Wiley", "pagerange": "1790-1795", "id_number": "CaltechAUTHORS:20111116-085157884", "issn": "0961-8368", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20111116-085157884", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Burroughs Wellcome Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "5T32GM07616" } ] }, "doi": "10.1002/pro.729", "pmcid": "PMC3267943", "resource_type": "article", "pub_year": "2011", "author_list": "Saraogi, Ishu; Akopian, David; et el." }, { "id": "https://authors.library.caltech.edu/records/jw47x-jqv96", "eprint_id": 27517, "eprint_status": "archive", "datestamp": "2023-08-19 08:10:09", "lastmod": "2023-10-24 17:12:53", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Saraogi-Ishu", "name": { "family": "Saraogi", "given": "Ishu" } }, { "id": "Zhang-Dawei", "name": { "family": "Zhang", "given": "Dawei" } }, { "id": "Chandrasekaran-Sandhya", "name": { "family": "Chandrasekaran", "given": "Sandhya" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Site-Specific Fluorescent Labeling of Nascent Proteins on the Translating Ribosome", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2011 American Chemical Society. \n\nReceived: July 15, 2011. Publication Date (Web): August 26, 2011. \n\nWe thank Prof. Swartz (Stanford University) for help with the generation of S30 extracts; Prof. Schultz (TSRI) for providing the pEVOL plasmid containing the tRNACUA/aaRS pair; Prof. Chapman (TSRI) for a sample of Cm; Profs. Tirrell, Barton, and Dougherty (Caltech) for use of their facilities; Prof. Sando(Kyushu University) for the RF-1 inhibitor aptamer sequence; and D. Dougherty and members of the Shan group for helpful comments on the manuscript. This work was supported by NIH Grant GM078024 to S.S. S.S. was supported by the Henry Dreyfus Teacher-Scholar Award, the Beckman Young Investigator Award, and the Packard and Lucile Award in Science and Engineering.\n\nAccepted Version - nihms323508.pdf
Supplemental Material - ja206626g_si_001.pdf
", "abstract": "As newly synthesized proteins emerge from the ribosome, they interact with a variety of cotranslational cellular machineries that facilitate their proper folding, maturation, and localization. These interactions are essential for proper function of the cell, and the ability to study these events is crucial to understanding cellular protein biogenesis.\nTo this end, we have developed a highly efficient\nmethod to generate ribosomenascent chain complexes\n(RNCs) site-specifically labeled with a fluorescent dye on\nthe nascent polypeptide. The fluorescent RNC provides\nreal-time, quantitative information on its cotranslational\ninteraction with the signal recognition particle and will be\na valuable tool in elucidating the role of the translating\nribosome in numerous biochemical pathways.", "date": "2011-09-28", "date_type": "published", "publication": "Journal of the American Chemical Society", "volume": "133", "number": "38", "publisher": "American Chemical Society", "pagerange": "14936-14939", "id_number": "CaltechAUTHORS:20111031-102635621", "issn": "0002-7863", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20111031-102635621", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" } ] }, "doi": "10.1021/ja206626g", "pmcid": "PMC3189723", "primary_object": { "basename": "ja206626g_si_001.pdf", "url": "https://authors.library.caltech.edu/records/jw47x-jqv96/files/ja206626g_si_001.pdf" }, "related_objects": [ { "basename": "nihms323508.pdf", "url": "https://authors.library.caltech.edu/records/jw47x-jqv96/files/nihms323508.pdf" } ], "resource_type": "article", "pub_year": "2011", "author_list": "Saraogi, Ishu; Zhang, Dawei; et el." }, { "id": "https://authors.library.caltech.edu/records/hz8g6-g4f85", "eprint_id": 25224, "eprint_status": "archive", "datestamp": "2023-08-19 07:47:11", "lastmod": "2023-10-24 15:42:53", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Nguyen-Thang-X", "name": { "family": "Nguyen", "given": "Thang X." } }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Neher-Saskia", "name": { "family": "Neher", "given": "Saskia" } }, { "id": "Walter-Peter", "name": { "family": "Walter", "given": "Peter" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Concerted Complex Assembly and GTPase Activation in the Chloroplast Signal Recognition Particle", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2011 American Chemical Society. \n\nReceived: May 12, 2011. Revised: July 21, 2011. Publication Date (Web): July 22, 2011. \n\nWe thank X. Zhang, N. Pierce, and members of the Shan lab for helpful comments on the manuscript. This work was supported by NIH grant GM078024 to S.S. S.S. was supported by the Beckman Young Investigator award, the Packard and Lucile Award in Science and Engineering, and the Henry Dreyfus Teacher-Scholar Award.\n\nAccepted Version - nihms-1001159.pdf
Supplemental Material - bi200742a_si_001.pdf
", "abstract": "The universally conserved signal recognition particle (SRP) and SRP receptor (SR) mediate the cotranslational targeting of proteins to cellular membranes. In contrast, a unique chloroplast SRP in green plants is primarily dedicated to the post-translational targeting of light harvesting chlorophyll a/b binding (LHC) proteins. In both pathways, dimerization and activation between the SRP and SR GTPases mediate the delivery of cargo; whether and how the GTPase cycle in each system adapts to its distinct substrate proteins were unclear. Here, we show that interactions at the active site essential for GTPase activation in the chloroplast SRP and SR play key roles in the assembly of the GTPase complex. In contrast to their cytosolic homologues, GTPase activation in the chloroplast SRP\u2013SR complex contributes marginally to the targeting of LHC proteins. These results demonstrate that complex assembly and GTPase activation are highly coupled in the chloroplast SRP and SR and suggest that the chloroplast GTPases may forego the GTPase activation step as a key regulatory point. These features may reflect adaptations of the chloroplast SRP to the delivery of their unique substrate protein.", "date": "2011-08-23", "date_type": "published", "publication": "Biochemistry", "volume": "50", "number": "33", "publisher": "American Chemical Society", "pagerange": "7208-7217", "id_number": "CaltechAUTHORS:20110906-083414953", "issn": "0006-2960", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20110906-083414953", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" } ] }, "doi": "10.1021/bi200742a", "pmcid": "PMC6309729", "primary_object": { "basename": "bi200742a_si_001.pdf", "url": "https://authors.library.caltech.edu/records/hz8g6-g4f85/files/bi200742a_si_001.pdf" }, "related_objects": [ { "basename": "nihms-1001159.pdf", "url": "https://authors.library.caltech.edu/records/hz8g6-g4f85/files/nihms-1001159.pdf" } ], "resource_type": "article", "pub_year": "2011", "author_list": "Nguyen, Thang X.; Chandrasekar, Sowmya; et el." }, { "id": "https://authors.library.caltech.edu/records/bvhx2-kjk82", "eprint_id": 23451, "eprint_status": "archive", "datestamp": "2023-08-22 02:35:46", "lastmod": "2023-10-23 19:35:51", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Saraogi-Ishu", "name": { "family": "Saraogi", "given": "Ishu" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Molecular Mechanism of Co-translational Protein Targeting by the Signal Recognition Particle", "ispublished": "pub", "full_text_status": "public", "keywords": "GTPases; molecular regulation; protein targeting; protein translocation; signal recognition particle; signal sequence", "note": "\u00a9 2011 John Wiley & Sons A/S. \n\nReceived 6 June 2010, revised and accepted for publication 1 February 2011; uncorrected manuscript published online 3 February 2011; published online 25 February 2011. \n\nWe thank members of the Shan Laboratory and expert reviewers for helpful comments on the manuscript. Our work is supported by the National Institute of Health (GM078024), the Burroughs Wellcome Fund, the Arnold and Mabel Beckman Foundation, the David and Lucile Packard Foundation and the Camile and Henry Dreyfus Foundation.\n\nAccepted Version - nihms-285827.pdf
", "abstract": "The signal recognition particle (SRP) is a key component of the cellular machinery that couples the ongoing synthesis of proteins to their proper localization, and has often served as a paradigm for understanding the molecular basis of protein localization within the cell. The SRP pathway exemplifies several key molecular events required for protein targeting to cellular membranes: the specific recognition of signal sequences on cargo proteins, the efficient delivery of cargo to the target membrane, the productive unloading of cargo to the translocation machinery and the precise spatial and temporal coordination of these molecular events. Here we highlight recent advances in our understanding of the molecular mechanisms underlying this pathway, and discuss new questions raised by these findings.", "date": "2011-05", "date_type": "published", "publication": "Traffic", "volume": "12", "number": "5", "publisher": "Wiley", "pagerange": "535-542", "id_number": "CaltechAUTHORS:20110425-113304294", "issn": "1398-9219", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20110425-113304294", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Burroughs Wellcome Fund" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camile and Henry Dreyfus Foundation" } ] }, "doi": "10.1111/j.1600-0854.2011.01171.x", "pmcid": "PMC3077218", "primary_object": { "basename": "nihms-285827.pdf", "url": "https://authors.library.caltech.edu/records/bvhx2-kjk82/files/nihms-285827.pdf" }, "resource_type": "article", "pub_year": "2011", "author_list": "Saraogi, Ishu and Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/dgex9-jj749", "eprint_id": 23525, "eprint_status": "archive", "datestamp": "2023-08-22 02:36:19", "lastmod": "2023-10-23 19:43:36", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Zhang-Xin", "name": { "family": "Zhang", "given": "Xin" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Synergistic actions between the SRP RNA and translating ribosome allow efficient delivery of the correct cargos during cotranslational protein targeting", "ispublished": "pub", "full_text_status": "public", "keywords": "Signal Recognition Particle; protein targeting; catalytic RNA; fluorescence; GTPases", "note": "\u00a9 2011 RNA Society. Published by Cold Spring Harbor Laboratory Press. \n\nReceived December 21, 2010; accepted February 22, 2011. Published in Advance April 1, 2011. \n\nWe thank members of the Shan group for helpful comments on\nthe manuscript. This work was supported by NIH grant\nGM078024 to S.S. S.S. was supported by a career award from\nthe Burroughs Wellcome Foundation, the Beckman Young Investigator award, the Packard and Lucile award in science and engineering, and the Henry Dreyfus teacher-scholar award.\n\nPublished - Shen2011p13641Rna.pdf
Supplemental Material - Supplementary_figure_1.doc
", "abstract": "During cotranslational protein targeting by the Signal Recognition Particle (SRP), the correct cargo accelerates stable complex assembly between the SRP and SRP receptor (FtsY) by several orders of magnitude, thus enabling rapid and faithful cargo delivery to the target membrane. The molecular mechanism underlying this cargo-induced rate acceleration has been unclear. Here we show that the SRP RNA allows assembly of the SRP\u2013FtsY complex to be specifically stimulated by a correct cargo, and, reciprocally, a correct cargo enables the SRP RNA to optimize its electrostatic interactions with FtsY. These results combined with recent structural work led us to suggest a \"conformational selection\" model that explains the synergistic action of the SRP RNA with the cargo in accelerating complex assembly. In addition to its previously proposed role in preventing the premature dissociation of SRP and FtsY, we found that the SRP RNA also plays an active role in ensuring the formation of productive assembly intermediates, thus guiding the SRP and FtsY through the most efficient pathway of assembly.", "date": "2011-05", "date_type": "published", "publication": "RNA", "volume": "17", "number": "5", "publisher": "RNA Society", "pagerange": "892-902", "id_number": "CaltechAUTHORS:20110502-112759851", "issn": "1355-8382", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20110502-112759851", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Burroughs Wellcome Foundation" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" } ] }, "doi": "10.1261/rna.2610411", "pmcid": "PMC3078738", "primary_object": { "basename": "Shen2011p13641Rna.pdf", "url": "https://authors.library.caltech.edu/records/dgex9-jj749/files/Shen2011p13641Rna.pdf" }, "related_objects": [ { "basename": "Supplementary_figure_1.doc", "url": "https://authors.library.caltech.edu/records/dgex9-jj749/files/Supplementary_figure_1.doc" } ], "resource_type": "article", "pub_year": "2011", "author_list": "Shen, Kuang; Zhang, Xin; et el." }, { "id": "https://authors.library.caltech.edu/records/jgha3-6rh36", "eprint_id": 23632, "eprint_status": "archive", "datestamp": "2023-08-22 02:32:47", "lastmod": "2023-10-23 19:48:59", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Zhang-Xin", "name": { "family": "Zhang", "given": "Xin" } }, { "id": "Lam-Vinh-Q", "name": { "family": "Lam", "given": "Vinh Q." } }, { "id": "Mou-Yun", "name": { "family": "Mou", "given": "Yun" } }, { "id": "Kimura-Tetsunari", "name": { "family": "Kimura", "given": "Tetsunari" } }, { "id": "Chung-Jaeyoon", "name": { "family": "Chung", "given": "Jaeyoon" } }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Winkler-J-R", "name": { "family": "Winkler", "given": "Jay R." }, "orcid": "0000-0002-4453-9716" }, { "id": "Mayo-S-L", "name": { "family": "Mayo", "given": "Stephen L." }, "orcid": "0000-0002-9785-5018" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Direct visualization reveals dynamics of a transient intermediate during protein assembly", "ispublished": "pub", "full_text_status": "public", "keywords": "EPR spectroscopy; fluorescence spectroscopy; molecular recognition; protein targeting; GTPases", "note": "\u00a9 2011 National Academy of Sciences. \n\nEdited by Jos\u00e9 N. Onuchic, University of California, San Diego, La Jolla, CA, and approved February 25, 2011 (received for review December 17, 2010). Published online before print April 4, 2011. \n\nWe thank B.S.P. Araujo for modeling the effect of fluorophore linkers on distant measurements; H.B. Gray and P.E. Wright for insightful discussions; and D.C. Rees, T.F. Miller III, and members of the Shan laboratory for comments on the manuscript. This work was supported by National Institutes of Health Grants GM078024 to S.-o.S. and GM068041 to J.R.W., DARPA Protein Design Processes to S.L.M., and career awards from the Burroughs Welcome Foundation, the Henry and Camille Dreyfus Foundation, the Arnold and Mabel Beckman Foundation, and the David and Lucile Packard Foundation to S.-o.S. \n\nAuthor contributions: X.Z., V.Q.L., J.R.W., and S.-o.S. designed research; X.Z., V.Q.L., Y.M., T.K., J.C., S.C., and S.-o.S. performed research; X.Z. and V.Q.L. contributed new reagents/analytic tools; X.Z., V.Q.L., Y.M., T.K., J.R.W., S.L.M., and S.-o.S. analyzed data; and X.Z. and S.-o.S. wrote the paper.\n\nPublished - Zhang2011p13713P_Natl_Acad_Sci_Usa.pdf
Supplemental Material - SM01.mov
Supplemental Material - pnas.1019051108_SI.pdf
", "abstract": "Interactions between proteins underlie numerous biological functions. Theoretical work suggests that protein interactions initiate with formation of transient intermediates that subsequently relax to specific, stable complexes. However, the nature and roles of these transient intermediates have remained elusive. Here, we characterized the global structure, dynamics, and stability of a transient, on-pathway intermediate during complex assembly between the Signal Recognition Particle (SRP) and its receptor. We show that this intermediate has overlapping but distinct interaction interfaces from that of the final complex, and it is stabilized by long-range electrostatic interactions. A wide distribution of conformations is explored by the intermediate; this distribution becomes more restricted in the final complex and is further regulated by the cargo of SRP. These results suggest a funnel-shaped energy landscape for protein interactions, and they provide a framework for understanding the role of transient intermediates in protein assembly and biological regulation.", "date": "2011-04-19", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "108", "number": "16", "publisher": "National Academy of Sciences", "pagerange": "6450-6455", "id_number": "CaltechAUTHORS:20110511-090620791", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20110511-090620791", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "NIH", "grant_number": "GM068041" }, { "agency": "Burroughs Welcome Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Defense Advanced Research Projects Agency (DARPA)" } ] }, "doi": "10.1073/pnas.1019051108", "pmcid": "PMC3081034", "primary_object": { "basename": "SM01.mov", "url": "https://authors.library.caltech.edu/records/jgha3-6rh36/files/SM01.mov" }, "related_objects": [ { "basename": "Zhang2011p13713P_Natl_Acad_Sci_Usa.pdf", "url": "https://authors.library.caltech.edu/records/jgha3-6rh36/files/Zhang2011p13713P_Natl_Acad_Sci_Usa.pdf" }, { "basename": "pnas.1019051108_SI.pdf", "url": "https://authors.library.caltech.edu/records/jgha3-6rh36/files/pnas.1019051108_SI.pdf" } ], "resource_type": "article", "pub_year": "2011", "author_list": "Zhang, Xin; Lam, Vinh Q.; et el." }, { "id": "https://authors.library.caltech.edu/records/8zc9t-arx28", "eprint_id": 22848, "eprint_status": "archive", "datestamp": "2023-08-19 05:37:11", "lastmod": "2023-10-23 17:15:38", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Ataide-S-F", "name": { "family": "Ataide", "given": "Sandro F." } }, { "id": "Schmitz-N", "name": { "family": "Schmitz", "given": "Nikolaus" } }, { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Ke-Ailong", "name": { "family": "Ke", "given": "Ailong" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Doudna-J-A", "name": { "family": "Doudna", "given": "Jennifer A." } }, { "id": "Ban-Nenad", "name": { "family": "Ban", "given": "Nenad" }, "orcid": "0000-0002-9527-210X" } ] }, "title": "The Crystal Structure of the Signal Recognition Particle in Complex with Its Receptor", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2011 American Association for the Advancement of Science. \n\nReceived for publication 13 August 2010. Accepted for publication 18 January 2011. \n\nWe thank K. Zhou for excellent technical assistance\nand help with crystal preparation during the early stages\nof the project. Initial crystallographic analysis was\nperformed at beamline 8.2.2 at the Advanced Light\nSource (ALS), Lawrence Berkeley National Laboratory;\nwe acknowledge C. Ralston for outstanding technical\nassistance at the ALS. Crystallographic data were\ncollected at the beamline X06SA at the Swiss Light Source\n(SLS). We thank A. Brunger for the prerelease version\nof CNS and for helpful comments on the refinement,\nC. Schulze-Briese and T. Tomizaki for their outstanding\nsupport at the SLS, T. Maier and S. Klinge for critical\ndiscussion and reading of the manuscript, and T. Maier\nand M. Leibundgut for help and assistance with data\ncollection and solving the structure. S.F.A. was funded\ninitially by the Howard Hughes Medical Institute and\ncurrently by an ETH postdoctoral fellowship, N.S. is\nfunded by Boehringer Ingelheim Fonds, and K.S. is\nfunded by NIH grant GM078024 to S.S. This work was\nsupported in part by the Howard Hughes Medical Institute\n(J.A.D.) and by the Swiss National Science Foundation\n(SNSF) and the National Center of Excellence in Research\n(NCCR) Structural Biology program of the SNSF. Atomic\ncoordinates and structure factors for the SRP:SR crystal\nstructure have been deposited with the Protein Data Bank\nunder accession code 2xxa.\n\nAccepted Version - nihms505518.pdf
Supplemental Material - Ataide.SOM.pdf
", "abstract": "Cotranslational targeting of membrane and secretory proteins is mediated by the universally conserved signal recognition particle (SRP). Together with its receptor (SR), SRP mediates the guanine triphosphate (GTP)\u2013dependent delivery of translating ribosomes bearing signal sequences to translocons on the target membrane. Here, we present the crystal structure of the SRP:SR complex at 3.9 angstrom resolution and biochemical data revealing that the activated SRP:SR guanine triphosphatase (GTPase) complex binds the distal end of the SRP hairpin RNA where GTP hydrolysis is stimulated. Combined with previous findings, these results suggest that the SRP:SR GTPase complex initially assembles at the tetraloop end of the SRP RNA and then relocalizes to the opposite end of the RNA. This rearrangement provides a mechanism for coupling GTP hydrolysis to the handover of cargo to the translocon.", "date": "2011-02-18", "date_type": "published", "publication": "Science", "volume": "331", "number": "6019", "publisher": "American Association for the Advancement of Science", "pagerange": "881-886", "id_number": "CaltechAUTHORS:20110314-100418858", "issn": "0036-8075", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20110314-100418858", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Howard Hughes Medical Institute (HHMI)" }, { "agency": "ETH Zurich" }, { "agency": "Boehringer Ingelheim Fonds" }, { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Swiss National Science Foundation (SNSF)" } ] }, "doi": "10.1126/science.1196473", "pmcid": "PMC3758919", "primary_object": { "basename": "Ataide.SOM.pdf", "url": "https://authors.library.caltech.edu/records/8zc9t-arx28/files/Ataide.SOM.pdf" }, "related_objects": [ { "basename": "nihms505518.pdf", "url": "https://authors.library.caltech.edu/records/8zc9t-arx28/files/nihms505518.pdf" } ], "resource_type": "article", "pub_year": "2011", "author_list": "Ataide, Sandro F.; Schmitz, Nikolaus; et el." }, { "id": "https://authors.library.caltech.edu/records/xkhjt-0c174", "eprint_id": 22157, "eprint_status": "archive", "datestamp": "2023-08-19 05:05:19", "lastmod": "2023-10-23 15:45:12", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Estrozi-L-F", "name": { "family": "Estrozi", "given": "Leandro F." } }, { "id": "Boehringer-D", "name": { "family": "Boehringer", "given": "Daniel" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Ban-Nenad", "name": { "family": "Ban", "given": "Nenad" }, "orcid": "0000-0002-9527-210X" }, { "id": "Schaffitzel-C", "name": { "family": "Schaffitzel", "given": "Christiane" } } ] }, "title": "Cryo-EM structure of the E. coli translating ribosome in complex with SRP and its receptor", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2010 Nature Publishing Group, a division of Macmillan Publishers Limited. \n\nReceived 25 April; accepted 8 October; published online 12 December 2010.\nWe would like to thank G. Schoehn for collecting EM data, I. Berger for designing scSRP, C. Frick for technical assistance and X. Zhang for helpful suggestions.\nW. Wintermeyer (Max Planck Institute for Biophysical Chemistry) kindly provided pET24a-Ffh. J. Luirink (Vrije Universiteit Amsterdam) is acknowledged for providing pET9-FtsY. This work was supported by the Swiss National Science Foundation (SNSF) and the National Center of Excellence in Research Structural Biology program of the SNSF. The authors acknowledge support by the Electron Microscopy of ETH Zurich (EMEZ) and from the infrastructure of the Partnership for Structural Biology in Grenoble.\nAuthor Contributions: C.S. conceived the scSRP construct and performed sample preparations; S.-o.S. performed activity assays; D.B. did the electron microscopy; D.B., L.F.E. and C.S. performed the image analysis and model building; C.S., N.B., D.B. and S.-o.S. prepared the manuscript.\n\nAccepted Version - nihms505492.pdf
Supplemental Material - nsmb.1952-S1.pdf
", "abstract": "We report the 'early' conformation of the Escherichia coli signal recognition particle (SRP) and its receptor FtsY bound to the translating ribosome, as determined by cryo-EM. FtsY binds to the tetraloop of the SRP RNA, whereas the NG domains of the SRP protein and FtsY interact weakly in this conformation. Our results suggest that optimal positioning of the SRP RNA tetraloop and the Ffh NG domain leads to FtsY recruitment.", "date": "2011-01", "date_type": "published", "publication": "Nature Structural & Molecular Biology", "volume": "18", "number": "1", "publisher": "Nature Publishing Group", "pagerange": "88-90", "id_number": "CaltechAUTHORS:20110214-091234951", "issn": "1545-9985", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20110214-091234951", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Swiss National Science Foundation (SNSF)" }, { "agency": "Electron Microscopy of ETH Zurich (EMEZ)" }, { "agency": "Partnership for Structural Biology in Grenoble" } ] }, "doi": "10.1038/nsmb.1952", "pmcid": "PMC3764645", "primary_object": { "basename": "nihms505492.pdf", "url": "https://authors.library.caltech.edu/records/xkhjt-0c174/files/nihms505492.pdf" }, "related_objects": [ { "basename": "nsmb.1952-S1.pdf", "url": "https://authors.library.caltech.edu/records/xkhjt-0c174/files/nsmb.1952-S1.pdf" } ], "resource_type": "article", "pub_year": "2011", "author_list": "Estrozi, Leandro F.; Boehringer, Daniel; et el." }, { "id": "https://authors.library.caltech.edu/records/mzksm-d8894", "eprint_id": 20033, "eprint_status": "archive", "datestamp": "2023-08-19 03:35:57", "lastmod": "2023-10-20 22:09:29", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Lam-Vinh-Q", "name": { "family": "Lam", "given": "Vinh Q." } }, { "id": "Akopian-D", "name": { "family": "Akopian", "given": "David" } }, { "id": "Rome-M", "name": { "family": "Rome", "given": "Michael" } }, { "id": "Henningsen-D", "name": { "family": "Henningsen", "given": "Doug" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Lipid activation of the signal recognition particle receptor provides spatial coordination of protein targeting", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2010 Lam et al. This article is distributed under the terms of an Attribution\u2013Noncommercial\u2013Share Alike\u2013No Mirror Sites license for the first six months after the publication date (see\nhttp://www.rupress.org/terms). After six months it is available under a Creative Commons\nLicense (Attribution\u2013Noncommercial\u2013Share Alike 3.0 Unported license, as described at\nhttp://creativecommons.org/licenses/by-nc-sa/3.0/).\n\nSubmitted: 26 April 2010.\nAccepted: 26 July 2010.\nWe thank Oded Lewinson in the Rees group for help and advice on liposome\nexperiments, Jost Vielmetter at the Protein Expression Center (PEC) at Caltech and\nJ. Van Deventer of the Tirrel group for help in SPR experiments, and Ray Deshaies\nand members of the Shan laboratory for comments on the manuscript.\nThis work was supported by National Institutes of Health (NIH) grant\nGM078024, and career awards from the Burroughs Welcome Foundation,\nthe Henry and Camille Dreyfus foundation, the Arnold and Mabel Beckman\nfoundation, and the David and Lucile Packard foundation to S. Shan. D. Akopian\nand M. Rome were supported by NIH/National Research Service Award\ntraining grant 5T32GM07616. The Biacore instrument at the PEC is funded\nby the Moore Foundation grant for \"Center for Integrative Study of Cell Regulation\"\nand in part by Caltech's Beckman Institute Funds.\n\nPublished - Lam2010p11400J_Cell_Biol.pdf
Supplemental Material - JCB_201004129_sm.pdf
", "abstract": "The signal recognition particle (SRP) and SRP receptor comprise the major cellular machinery that mediates the cotranslational targeting of proteins to cellular membranes. It remains unclear how the delivery of cargos to the target membrane is spatially coordinated. We show here that phospholipid binding drives important conformational rearrangements that activate the bacterial SRP receptor FtsY and the SRP\u2013FtsY complex. This leads to accelerated SRP\u2013FtsY complex assembly, and allows the SRP\u2013FtsY complex to more efficiently unload cargo proteins. Likewise, formation of an active SRP\u2013FtsY GTPase complex exposes FtsY's lipid-binding helix and enables stable membrane association of the targeting complex. Thus, membrane binding, complex assembly with SRP, and cargo unloading are inextricably linked to each other via conformational changes in FtsY. These allosteric communications allow the membrane delivery of cargo proteins to be efficiently coupled to their subsequent unloading and translocation, thus providing spatial coordination during protein targeting.", "date": "2010-08-23", "date_type": "published", "publication": "Journal of Cell Biology", "volume": "190", "number": "4", "publisher": "Rockefeller University Press", "pagerange": "623-635", "id_number": "CaltechAUTHORS:20100920-093243413", "issn": "0021-9525", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20100920-093243413", "rights": "This article is distributed under the terms of an Attribution\u2013Noncommercial\u2013Share Alike\u2013No Mirror Sites license for the first six months after the publication date (see\nhttp://www.rupress.org/terms). After six months it is available under a Creative Commons\nLicense (Attribution\u2013Noncommercial\u2013Share Alike 3.0 Unported license, as described at\nhttp://creativecommons.org/licenses/by-nc-sa/3.0/).", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Burroughs Welcome Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "Arnold and Mabel Beckman foundation" }, { "agency": "David and Lucile Packard foundation" }, { "agency": "NIH Predoctoral Fellowship", "grant_number": "5T32GM07616" }, { "agency": "Gordon and Betty Moore Foundation" }, { "agency": "Caltech Beckman Institute" } ] }, "doi": "10.1083/jcb.201004129", "pmcid": "PMC2928010", "primary_object": { "basename": "JCB_201004129_sm.pdf", "url": "https://authors.library.caltech.edu/records/mzksm-d8894/files/JCB_201004129_sm.pdf" }, "related_objects": [ { "basename": "Lam2010p11400J_Cell_Biol.pdf", "url": "https://authors.library.caltech.edu/records/mzksm-d8894/files/Lam2010p11400J_Cell_Biol.pdf" } ], "resource_type": "article", "pub_year": "2010", "author_list": "Lam, Vinh Q.; Akopian, David; et el." }, { "id": "https://authors.library.caltech.edu/records/e5hxv-02911", "eprint_id": 18783, "eprint_status": "archive", "datestamp": "2023-08-19 02:39:42", "lastmod": "2023-10-20 16:45:47", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Jaru-Ampornpan-P", "name": { "family": "Jaru-Ampornpan", "given": "Peera" } }, { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Lam-Vinh-Q", "name": { "family": "Lam", "given": "Vinh Q." } }, { "id": "Ali-M", "name": { "family": "Ali", "given": "Mona" } }, { "id": "Doniach-S", "name": { "family": "Doniach", "given": "Sebastian" } }, { "id": "Jia-Tony-Z", "name": { "family": "Jia", "given": "Tony Z." } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "ATP-independent reversal of a membrane protein aggregate by a chloroplast SRP subunit", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2010 Nature Publishing Group, a division of Macmillan Publishers Limited. \n\nReceived 18 March; accepted 20 April; published online 27 April 2010. \n\nWe thank R. Henry (Univ. of Arkansas) for expression plasmids of cpSRP43 and\nLHCP, C. Robinson (Univ. of Warwick) for plasmids encoding LHCA1 and LHCB5,\nZ. Liu for help and advice with LHCP reconstitution, A. Sim and V. Chu of the\nDoniach group and D. Herschlag for help and advice with the SAXS experiments\nand D.C. Rees, W.M. Clemons, A. Varshavsky, N. Pierce and members of the\nShan laboratory for comments on the manuscript. This work was supported by\nUS National Institutes of Health grant GM078024 and career awards from the\nBurroughs Welcome Foundation, the Henry and Camille Dreyfus foundation, the\nBeckman foundation and the Packard foundation to S.-o.S. and by US National\nInstitutes of Health program project grant P01-GM-66275 to S.D. and D.H.\nAuthor Contributions: P. J.-A. and S.-o.S. designed experiments; P.J.-A. and T.Z.J. performed the\nbiochemical experiments and analyzed data; V.Q.L. and M.A. performed the SAXS\nexperiment and analyzed data; K.S., M.A. and S.D. carried out molecular dynamics\nsimulations of SAXS data; P. J.-A. and S.-o.S. wrote the paper.\n\nAccepted Version - nihms223254.pdf
Supplemental Material - nsmb.1836-S1.pdf
", "abstract": "Membrane proteins impose enormous challenges to cellular protein homeostasis during their post-translational targeting, and they require chaperones to keep them soluble and translocation competent. Here we show that a novel targeting factor in the chloroplast signal recognition particle (cpSRP), cpSRP43, is a highly specific molecular chaperone that efficiently reverses the aggregation of its substrate proteins. In contrast to 'ATPases associated with various cellular activities' (AAA+) chaperones, cpSRP43 uses specific binding interactions with its substrate to mediate its 'disaggregase' activity. This disaggregase capability can allow targeting machineries to more effectively capture their protein substrates and emphasizes a close connection between protein folding and trafficking processes. Moreover, cpSRP43 provides the first example to our knowledge of an ATP-independent disaggregase and shows that efficient reversal of protein aggregation can be attained by specific binding interactions between a chaperone and its substrate.", "date": "2010-06", "date_type": "published", "publication": "Nature Structural & Molecular Biology", "volume": "17", "number": "6", "publisher": "Nature Publishing Group", "pagerange": "696-703", "id_number": "CaltechAUTHORS:20100623-160406505", "issn": "1545-9985", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20100623-160406505", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "NIH", "grant_number": "P01-GM-66275" }, { "agency": "Burroughs Welcome Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" } ] }, "doi": "10.1038/nsmb.1836", "pmcid": "PMC2917185", "primary_object": { "basename": "nihms223254.pdf", "url": "https://authors.library.caltech.edu/records/e5hxv-02911/files/nihms223254.pdf" }, "related_objects": [ { "basename": "nsmb.1836-S1.pdf", "url": "https://authors.library.caltech.edu/records/e5hxv-02911/files/nsmb.1836-S1.pdf" } ], "resource_type": "article", "pub_year": "2010", "author_list": "Jaru-Ampornpan, Peera; Shen, Kuang; et el." }, { "id": "https://authors.library.caltech.edu/records/cghp0-sen46", "eprint_id": 18403, "eprint_status": "archive", "datestamp": "2023-08-19 02:30:58", "lastmod": "2023-10-20 16:11:09", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Zhang-Xin", "name": { "family": "Zhang", "given": "Xin" } }, { "id": "Rashid-R", "name": { "family": "Rashid", "given": "Rumana" } }, { "id": "Wang-Kai", "name": { "family": "Wang", "given": "Kai" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Sequential Checkpoints Govern Substrate Selection During Cotranslational Protein Targeting", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2010 American Association for the Advancement of Science.\n\nReceived 6 January 2010; accepted 18 March 2010.\n\nWe thank C. Schaffitzel and N. Ban for help with the\npurification of RNCs and Trigger factor; J. Luirink for\nplasmids encoding the phoA signal sequence variants;\nL. Randall for the plasmid encoding luciferase; B. Bukau\nand E. Deuerling for the plasmid encoding Trigger factor;\nF. Nataro for the plasmid encoding EspP; H. D. Bernstein for\nthe strain HDB51; and R. J. Deshaies, A. Varshavsky,\nW. Zhong, N. Pierce, and the Shan laboratory for comments\non the manuscript. This work was supported by NIH grant\nGM078024, and career awards from the Burroughs\nWellcome Foundation, the Henry and Camille Dreyfus\nFoundation, the Arnold and Mabel Beckman Foundation,\nand the Packard Foundation to S.S.\n\nAccepted Version - nihms505487.pdf
Supplemental Material - 1.pdf
", "abstract": "Proper protein localization is essential for all cells. However, the precise mechanism by which high fidelity is achieved is not well understood for any protein-targeting pathway. To address this fundamental question, we investigated the signal recognition particle (SRP) pathway in Escherichia coli, which delivers proteins to the bacterial inner membrane through recognition of signal sequences on cargo proteins. Fidelity was thought to arise from the inability of SRP to bind strongly to incorrect cargos. Using biophysical assays, we found that incorrect cargos were also rejected through a series of checkpoints during subsequent steps of targeting. Thus, high fidelity of substrate selection is achieved through the cumulative effect of multiple checkpoints; this principle may be generally applicable to other pathways involving selective signal recognition.", "date": "2010-05-07", "date_type": "published", "publication": "Science", "volume": "328", "number": "5979", "publisher": "American Association for the Advancement of Science", "pagerange": "757-760", "id_number": "CaltechAUTHORS:20100521-152717223", "issn": "0036-8075", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20100521-152717223", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Burroughs Wellcome Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" } ] }, "doi": "10.1126/science.1186743", "pmcid": "PMC3760334", "primary_object": { "basename": "nihms505487.pdf", "url": "https://authors.library.caltech.edu/records/cghp0-sen46/files/nihms505487.pdf" }, "related_objects": [ { "basename": "1.pdf", "url": "https://authors.library.caltech.edu/records/cghp0-sen46/files/1.pdf" } ], "resource_type": "article", "pub_year": "2010", "author_list": "Zhang, Xin; Rashid, Rumana; et el." }, { "id": "https://authors.library.caltech.edu/records/4xcw9-62c93", "eprint_id": 18469, "eprint_status": "archive", "datestamp": "2023-08-21 23:49:05", "lastmod": "2023-10-20 16:25:09", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shen-Kuang", "name": { "family": "Shen", "given": "Kuang" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Transient tether between the SRP RNA and SRP receptor ensures efficient cargo delivery during cotranslational protein targeting", "ispublished": "pub", "full_text_status": "public", "keywords": "catalytic RNA; macromolecular interaction; signal recognition particle; GTPases fluroescence", "note": "\u00a9 2010 the National Academy of Sciences. Freely available online through the PNAS open access option. \n\nCommunicated by Douglas C. Rees, Caltech/Howard Hughes Medical Institute, Pasadena, CA, March 16, 2010 (received for review October 9, 2009). Published online before print April 12, 2010. \n\nWe thank William M. Clemons, Nenad Ban, Christiane\nSchaffitzel, and members of the Shan group for helpful comments on the\nmanuscript. This work was supported by National Institutes of Health Grant\nGM078024 (S.S.). S.S. was supported by a career award from the Burroughs\nWelcome Foundation, the Beckman Young Investigator award, the Packard\nand Lucile Award in Science and Engineering, and the Henry Dreyfus Teacher-\nScholar Award.\nAuthor contributions: K.S. and S.-o.S. designed research; K.S. performed research; K.S.\nanalyzed data; and K.S. and S.-o.S. wrote the paper.\n\nPublished - Shen2010p10034P_Natl_Acad_Sci_Usa.pdf
Supplemental Material - pnas.1002968107_SI.pdf
", "abstract": "Kinetic control of macromolecular interactions plays key roles in biological regulation. An example of such control occurs in cotranslational protein targeting by the signal recognition particle (SRP), during which the SRP RNA and the cargo both accelerate complex assembly between the SRP and SRP receptor FtsY 10^2-fold. The molecular mechanism underlying these rate accelerations was unclear. Here we show that a highly conserved basic residue, Lys399, on the lateral surface of FtsY provides a novel RNA tetraloop receptor to mediate the SRP RNA- and cargo-induced acceleration of SRP\u2013FtsY complex assembly. We propose that the SRP RNA, by using its tetraloop to interact with FtsY\u2013Lys399, provides a transient tether to stabilize the early stage and transition state of complex formation; this accelerates the assembly of a stable SRP\u2013FtsY complex and allows the loading of cargo to be efficiently coupled to its membrane delivery. The use of a transient tether to increase the lifetime of collisional intermediates and reduce the dimension of diffusional search represents a novel and effective mechanism to accelerate macromolecular interactions.", "date": "2010-04-27", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "107", "number": "17", "publisher": "National Academy of Sciences", "pagerange": "7698-7703", "id_number": "CaltechAUTHORS:20100527-075322402", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20100527-075322402", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Burroughs Welcome Foundation" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" } ] }, "doi": "10.1073/pnas.1002968107", "pmcid": "PMC2867919", "primary_object": { "basename": "Shen2010p10034P_Natl_Acad_Sci_Usa.pdf", "url": "https://authors.library.caltech.edu/records/4xcw9-62c93/files/Shen2010p10034P_Natl_Acad_Sci_Usa.pdf" }, "related_objects": [ { "basename": "pnas.1002968107_SI.pdf", "url": "https://authors.library.caltech.edu/records/4xcw9-62c93/files/pnas.1002968107_SI.pdf" } ], "resource_type": "article", "pub_year": "2010", "author_list": "Shen, Kuang and Shan, Shu-ou" }, { "id": "https://authors.library.caltech.edu/records/zazg2-s3b34", "eprint_id": 17010, "eprint_status": "archive", "datestamp": "2023-08-19 00:48:02", "lastmod": "2023-10-19 22:43:40", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Pierce-N-W", "name": { "family": "Pierce", "given": "Nathan W." } }, { "id": "Kleiger-G", "name": { "family": "Kleiger", "given": "Gary" }, "orcid": "0000-0003-3924-1680" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Deshaies-R-J", "name": { "family": "Deshaies", "given": "Raymond J." }, "orcid": "0000-0002-3671-9354" } ] }, "title": "Detection of sequential polyubiquitylation on a millisecond timescale", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2009 Macmillan Publishers Limited. \n\nReceived 25 September 2009; Accepted 19 October 2009. \n\nWe thank J. Vielmetter for providing SCFCdc4, b-TrCP-Skp1\nand human E1; S. Hess, R. L. J. Graham and the Proteome Exploration Laboratory for providing assistance with mass spectrometry of CYCE and Cdc34 thioester. We thank S. Schwarz, B. Schulman and G. Wu for gifts of reagents. We thank D. Sprinzak and all the members of the Deshaies and Shan laboratories for support and discussions. N.W.P. was supported by the Gordon Ross Fellowship and a National Institutes of Health Training Grant. R.J.D. is an Investigator of the Howard Hughes Medical Institute. This work was supported in part by National Institutes of Health GM065997. \n\nAuthor Contributions N.W.P. performed all computational modelling and experiments, except G.K. performed the mass spectrometry experiments in Fig. 1g. N.W.P., R.J.D. and S.-o.S. conceived the experiments. N.W.P. and R.J.D. wrote the manuscript with editorial input from the other authors.\nSupplementary Information is linked to the online version of the paper at www.nature.com/nature.\n\nAccepted Version - nihms153662.pdf
Supplemental Material - nature08595-s1.pdf
", "abstract": "The pathway by which ubiquitin chains are generated on substrate through a cascade of enzymes consisting of an E1, E2 and E3 remains unclear. Multiple distinct models involving chain assembly on E2 or substrate have been proposed. However, the speed and complexity of the reaction have precluded direct experimental tests to distinguish between potential pathways. Here we introduce new theoretical and experimental methodologies to address both limitations. A quantitative framework based on product distribution predicts that the really interesting new gene (RING) E3 enzymes SCF^(Cdc4) and SCF^(\u03b2-TrCP) work with the E2 Cdc34 to build polyubiquitin chains on substrates by sequential transfers of single ubiquitins. Measurements with millisecond time resolution directly demonstrate that substrate polyubiquitylation proceeds sequentially. Our results present an unprecedented glimpse into the mechanism of RING ubiquitin ligases and illuminate the quantitative parameters that underlie the rate and pattern of ubiquitin chain assembly.", "date": "2009-12-03", "date_type": "published", "publication": "Nature", "volume": "462", "number": "7273", "publisher": "Nature Publishing Group", "pagerange": "615-620", "id_number": "CaltechAUTHORS:20091221-110503492", "issn": "0028-0836", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20091221-110503492", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Gordon Ross Fellowship" }, { "agency": "NIH Predoctoral Fellowship" }, { "agency": "Howard Hughes Medical Institute (HHMI)" }, { "agency": "NIH", "grant_number": "GM065997" } ] }, "doi": "10.1038/nature08595", "pmcid": "PMC2791906", "primary_object": { "basename": "nihms153662.pdf", "url": "https://authors.library.caltech.edu/records/zazg2-s3b34/files/nihms153662.pdf" }, "related_objects": [ { "basename": "nature08595-s1.pdf", "url": "https://authors.library.caltech.edu/records/zazg2-s3b34/files/nature08595-s1.pdf" } ], "resource_type": "article", "pub_year": "2009", "author_list": "Pierce, Nathan W.; Kleiger, Gary; et el." }, { "id": "https://authors.library.caltech.edu/records/pyex9-nx640", "eprint_id": 15801, "eprint_status": "archive", "datestamp": "2023-08-19 00:01:23", "lastmod": "2023-10-19 17:15:44", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Jaru-Ampornpan-P", "name": { "family": "Jaru-Ampornpan", "given": "Peera" } }, { "id": "Nguyen-Thang-X", "name": { "family": "Nguyen", "given": "Thang X." } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "A Distinct Mechanism to Achieve Efficient Signal Recognition Particle (SRP)-SRP Receptor Interaction by the Chloroplast SRP Pathway", "ispublished": "pub", "full_text_status": "public", "note": "Copyright \u00a9 2009 by The American Society for Cell Biology. Under the License and Publishing Agreement, authors grant to the general public, effective two months after publication of (i.e.,. the appearance of) the edited manuscript in an online issue of MBoC, the nonexclusive right to copy, distribute, or display the manuscript subject to the terms of the Creative Commons\u2013Noncommercial\u2013Share Alike 3.0 Unported license (http://creativecommons.org/licenses/by-nc-sa/3.0). \n\nSubmitted October 2, 2008; Revised June 23, 2009; Accepted June 25, 2009. This was published online ahead of print in MBC in Press (http://www.molbiolcell.org/cgi/doi/10.1091/mbc.E08-10-0989) on July 8, 2009. \n\nWe thank the members of the Shan laboratory for helpful comments on the manuscript. This work was supported by National Institutes of Health grant GM-078024 (to S. S.). S. S. was supported by the Burroughs Wellcome Fund career award, the Beckman Young Investigator award, and the Packard and Lucile award in science and engineering. P.J.-A. was supported by a fellowship from the Brey Endowment foundation.\n\nPublished - JaruAmpornpan2009p5866Mol_Biol_Cell.pdf
Supplemental Material - E08-10-0989_Shan.doc
", "abstract": "Cotranslational protein targeting by the signal recognition particle (SRP) requires the SRP RNA, which accelerates the interaction between the SRP and SRP receptor 200-fold. This otherwise universally conserved SRP RNA is missing in the chloroplast SRP (cpSRP) pathway. Instead, the cpSRP and cpSRP receptor (cpFtsY) by themselves can interact 200-fold faster than their bacterial homologues. Here, cross-complementation analyses revealed the molecular origin underlying their efficient interaction. We found that cpFtsY is 5- to 10-fold more efficient than Escherichia coli FtsY at interacting with the GTPase domain of SRP from both chloroplast and bacteria, suggesting that cpFtsY is preorganized into a conformation more conducive to complex formation. Furthermore, the cargo-binding M-domain of cpSRP provides an additional 100-fold acceleration for the interaction between the chloroplast GTPases, functionally mimicking the effect of the SRP RNA in the cotranslational targeting pathway. The stimulatory effect of the SRP RNA or the M-domain of cpSRP is specific to the homologous SRP receptor in each pathway. These results strongly suggest that the M-domain of SRP actively communicates with the SRP and SR GTPases and that the cytosolic and chloroplast SRP pathways have evolved distinct molecular mechanisms (RNA vs. protein) to mediate this communication.", "date": "2009-09-01", "date_type": "published", "publication": "Molecular Biology of the Cell", "volume": "20", "number": "17", "publisher": "American Society for Cell Biology", "pagerange": "3965-3973", "id_number": "CaltechAUTHORS:20090911-153600633", "issn": "1059-1524", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20090911-153600633", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM-078024" }, { "agency": "Burroughs Wellcome Fund" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Brey Endowment Foundation" } ] }, "doi": "10.1091/mbc.E08-10-0989", "pmcid": "PMC2735494", "primary_object": { "basename": "JaruAmpornpan2009p5866Mol_Biol_Cell.pdf", "url": "https://authors.library.caltech.edu/records/pyex9-nx640/files/JaruAmpornpan2009p5866Mol_Biol_Cell.pdf" }, "related_objects": [ { "basename": "E08-10-0989_Shan.doc", "url": "https://authors.library.caltech.edu/records/pyex9-nx640/files/E08-10-0989_Shan.doc" } ], "resource_type": "article", "pub_year": "2009", "author_list": "Jaru-Ampornpan, Peera; Nguyen, Thang X.; et el." }, { "id": "https://authors.library.caltech.edu/records/sfmg0-jdt72", "eprint_id": 15891, "eprint_status": "archive", "datestamp": "2023-08-20 02:21:29", "lastmod": "2023-10-19 17:20:14", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Schmid-S-L", "name": { "family": "Schmid", "given": "Sandra L." } }, { "id": "Zhang-Xin", "name": { "family": "Zhang", "given": "Xin" } } ] }, "title": "Signal Recognition Particle (SRP) and SRP Receptor: A New Paradigm for Multistate Regulatory GTPases", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2009 American Chemical Society. \n\nReceived April 23, 2009; Revised Manuscript Received May 25, 2009. \n\nWe thank Thomas Pucadyil, Marcel Mettlen, and members of\nthe Shan laboratory for helpful comments on the manuscript.\n\nAccepted Version - nihms204160.pdf
", "abstract": "The GTP-binding proteins or GTPases comprise a superfamily of proteins that provide molecular switches in numerous cellular processes. The \"GTPase switch\" paradigm, in which a GTPase acts as a bimodal switch that is turned \"on\" and \"off\" by external regulatory factors, has been used to interpret the regulatory mechanism of many GTPases for more than two decades. Nevertheless, recent work has unveiled an emerging class of \"multistate\" regulatory GTPases that do not adhere to this classical paradigm. Instead of relying on external nucleotide exchange factors or GTPase activating proteins to switch between the on and off states, these GTPases have the intrinsic ability to exchange nucleotides and to sense and respond to upstream and downstream factors. In contrast to the bimodal nature of the GTPase switch, these GTPases undergo multiple conformational rearrangements, allowing multiple regulatory points to be built into a complex biological process to ensure the efficiency and fidelity of the pathway. We suggest that these multistate regulatory GTPases are uniquely suited to provide spatial and temporal control of complex cellular pathways that require multiple molecular events to occur in a highly coordinated fashion.", "date": "2009-07-28", "date_type": "published", "publication": "Biochemistry", "volume": "48", "number": "29", "publisher": "American Chemical Society", "pagerange": "6696-6704", "id_number": "CaltechAUTHORS:20090916-153459576", "issn": "0006-2960", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20090916-153459576", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1021/bi9006989", "pmcid": "PMC2883566", "primary_object": { "basename": "nihms204160.pdf", "url": "https://authors.library.caltech.edu/records/sfmg0-jdt72/files/nihms204160.pdf" }, "resource_type": "article", "pub_year": "2009", "author_list": "Shan, Shu-ou; Schmid, Sandra L.; et el." }, { "id": "https://authors.library.caltech.edu/records/2bkpp-x2t57", "eprint_id": 13862, "eprint_status": "archive", "datestamp": "2023-08-21 20:46:55", "lastmod": "2023-10-18 14:31:31", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Zhang-Xin", "name": { "family": "Zhang", "given": "Xin" } }, { "id": "Schaffitzel-C", "name": { "family": "Schaffitzel", "given": "Christiane" } }, { "id": "Ban-N", "name": { "family": "Ban", "given": "Nenad" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Multiple conformational switches in a GTPase complex control co-translational protein targeting", "ispublished": "pub", "full_text_status": "public", "keywords": "conformational change; fluorescence spectroscopy; protein targeting and translocation; signal recognition particle", "note": "\u00a9 2009 by the National Academy of Sciences. \n\nEdited by Jennifer A. Doudna, University of California, Berkeley, CA, and approved December 22, 2008 (received for review August 28, 2008). \n\nWe thank Sandra Schmid, Douglas C. Rees, Raymond\nDeshaies, Nathan Pierce, and members of the Shan laboratory for comments on\nthe manuscript. This work was supported by a National Institutes of Health grant\nGM078024 to S.S., and by the Swiss National Science Foundation (SNSF) and the\nNCCR Structural Biology program of the SNSF to N.B. S.S. was supported by a\ncareer award from the Burroughs Welcome Foundation, the Henry and Camille\nDreyfus foundation, the Beckman Young Investigator award, and the Packard\nand Lucile award in science and engineering. X.Z. was supported by a fellowship\nfrom the Ulric B. and Evelyn L. Bray Endowment Fund.\n\nPublished - ZHApnas09.pdf
Supplemental Material - ZHApnas09supp.pdf
", "abstract": "The \"GTPase switch\" paradigm, in which a GTPase switches between an active, GTP-bound state and an inactive, GDP-bound state through the recruitment of nucleotide exchange factors (GEFs) or GTPase activating proteins (GAPs), has been used to interpret the regulatory mechanism of many GTPases. A notable exception to this paradigm is provided by two GTPases in the signal recognition particle (SRP) and the SRP receptor (SR) that control the co-translational targeting of proteins to cellular membranes. Instead of the classical \"GTPase switch,\" both the SRP and SR undergo a series of discrete conformational rearrangements during their interaction with one another, culminating in their reciprocal GTPase activation. Here, we show that this series of rearrangements during SRP-SR binding and activation provide important control points to drive and regulate protein targeting. Using real-time fluorescence, we showed that the cargo for SRP\u2014ribosomes translating nascent polypeptides with signal sequences\u2014accelerates SRP\u00b7SR complex assembly over 100-fold, thereby driving rapid delivery of cargo to the membrane. A series of subsequent rearrangements in the SRP\u00b7SR GTPase complex provide important driving forces to unload the cargo during late stages of protein targeting. Further, the cargo delays GTPase activation in the SRP\u00b7SR complex by 8\u201312 fold, creating an important time window that could further improve the efficiency and fidelity of protein targeting. Thus, the SRP and SR GTPases, without recruiting external regulatory factors, constitute a self-sufficient system that provides exquisite spatial and temporal control of a complex cellular process.", "date": "2009-02-10", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "106", "number": "6", "publisher": "National Academy of Sciences", "pagerange": "1754-1759", "id_number": "CaltechAUTHORS:20090407-091223238", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20090407-091223238", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Swiss National Science Foundation (SNSF)" }, { "agency": "Burroughs Welcome Foundation" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Ulric B. and Evelyn L. Bray Endowment Fund" } ] }, "doi": "10.1073/pnas.0808573106", "pmcid": "PMC2644110", "primary_object": { "basename": "ZHApnas09supp.pdf", "url": "https://authors.library.caltech.edu/records/2bkpp-x2t57/files/ZHApnas09supp.pdf" }, "related_objects": [ { "basename": "ZHApnas09.pdf", "url": "https://authors.library.caltech.edu/records/2bkpp-x2t57/files/ZHApnas09.pdf" } ], "resource_type": "article", "pub_year": "2009", "author_list": "Zhang, Xin; Schaffitzel, Christiane; et el." }, { "id": "https://authors.library.caltech.edu/records/wv18h-z2c87", "eprint_id": 32731, "eprint_status": "archive", "datestamp": "2023-08-22 13:00:35", "lastmod": "2023-10-18 16:01:34", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Zhang-Xin", "name": { "family": "Zhang", "given": "Xin" } }, { "id": "Kung-Simon", "name": { "family": "Kung", "given": "Simon" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Demonstration of a Multistep Mechanism for Assembly of the SRP\u00b7SRP Receptor Complex: Implications for the Catalytic Role of SRP RNA", "ispublished": "pub", "full_text_status": "public", "keywords": "signal recognition particle; protein targeting and translocation; GTPases; SRP RNA; fluorescence resonance energy transfer", "note": "\u00a9 2008 Elsevier Ltd. \n\nReceived 16 April 2008. Revised 20 May 2008. Accepted 20 May 2008. Available online 29 May 2008. \n\nEdited by J. Doudna. \n\nThis work was supported by the National Institutes of Health through grant GM078024 to S.S. S.S. was supported by a Burroughs Wellcome Fund Career Award, the Henry and Camille Dreyfus Foundation, a Beckman Young Investigator Award, and a Packard and Lucile Award in Science and Engineering. X.Z. was supported by a fellowship from the Ulric B. and Evelyn L. Bray Endowment Fund. We thank members of the Shan laboratory for their comments on the manuscript.\n\nAccepted Version - nihms65232.pdf
Supplemental Material - mmc1.doc
", "abstract": "Two GTPases in the signal recognition particle (SRP) and its receptor (SR) control the delivery of newly synthesized proteins to the endoplasmic reticulum or plasma membrane. During the protein targeting reaction, the 4.5S SRP RNA accelerates the association between the two GTPases by 400-fold. Using fluorescence resonance energy transfer, we demonstrate here that formation of a stable SRP\u00b7SR complex involves two distinct steps: a fast initial association between SRP and SR to form a GTP-independent early complex and then a GTP-dependent conformational rearrangement to form the stable final complex. We also found that the 4.5S SRP RNA significantly stabilizes the early GTP-independent intermediate. Furthermore, mutational analyses show that there is a strong correlation between the ability of the mutant SRP RNAs to stabilize the early intermediate and their ability to accelerate SRP\u00b7SR complex formation. We propose that the SRP RNA, by stabilizing the early intermediate, can give this transient intermediate a longer life time and therefore a higher probability to rearrange to the stable final complex. This provides a coherent model that explains how the 4.5S RNA exerts its catalytic role in SRP\u00b7SR complex assembly.", "date": "2008-09-05", "date_type": "published", "publication": "Journal of Molecular Biology", "volume": "381", "number": "3", "publisher": "Elsevier", "pagerange": "581-593", "id_number": "CaltechAUTHORS:20120726-073902314", "issn": "0022-2836", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20120726-073902314", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM078024" }, { "agency": "Burroughs Wellcome Fund" }, { "agency": "Camille and Henry Dreyfus Foundation" }, { "agency": "Arnold and Mabel Beckman Foundation" }, { "agency": "David and Lucile Packard Foundation" }, { "agency": "Ulric B. and Evelyn L. Bray Endowment Fund" } ] }, "doi": "10.1016/j.jmb.2008.05.049", "pmcid": "PMC2630804", "primary_object": { "basename": "mmc1.doc", "url": "https://authors.library.caltech.edu/records/wv18h-z2c87/files/mmc1.doc" }, "related_objects": [ { "basename": "nihms65232.pdf", "url": "https://authors.library.caltech.edu/records/wv18h-z2c87/files/nihms65232.pdf" } ], "resource_type": "article", "pub_year": "2008", "author_list": "Zhang, Xin; Kung, Simon; et el." }, { "id": "https://authors.library.caltech.edu/records/rae6w-ext41", "eprint_id": 32733, "eprint_status": "archive", "datestamp": "2023-08-22 10:58:09", "lastmod": "2023-10-18 16:01:41", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Chartron-Justin-W", "name": { "family": "Chartron", "given": "Justin" } }, { "id": "Jaru-Ampornpan-Peera", "name": { "family": "Jaru-Ampornpan", "given": "Peera" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Structure of the Chloroplast Signal Recognition Particle (SRP) Receptor: Domain Arrangement Modulates SRP\u2013Receptor Interaction", "ispublished": "pub", "full_text_status": "public", "keywords": "protein sorting and translocation; signal recognition particle; GTPases; conformational change; protein targeting in chloroplast", "note": "\u00a9 2007 Elsevier Ltd. \n\nReceived 24 May 2007. Revised 24 August 2007. Accepted 20 September 2007. Available online 28 September 2007. \n\nEdited by J. Doudna. \n\nWe thank D. C. Rees, P. J. Bjorkman, W. M. Clemons and their laboratory members for extensive assistance throughout this work and for valuable comments on the manuscript; B. Sankaran at the conformation from its E. coli and Taq homologues, and thus further contribute to the efficient association kinetics between cpSRP54 and cpFtsY. Together, the biochemical and crystallographic analyses of cpFtsY emphasize the importance of proper N\u2013G domain arrangement as one of the key factors that modulate the efficiency of interaction between the SRP and SR GTPases and help explain, in part, why the chloroplast SRP pathway can bypass the requirement for an SRP RNA, which is used to accelerate complex formation between the two GTPases\nin most bacterial SRP pathways.\n\nSupplemental Material - mmc1.doc
", "abstract": "The signal recognition particle (SRP) pathway mediates co-translational targeting of nascent proteins to membranes. Chloroplast SRP is unique in that it does not contain the otherwise universally conserved SRP RNA, which accelerates the association between the SRP guanosine-5\u2032-triphosphate (GTP) binding protein and its receptor FtsY in classical SRP pathways. Recently, we showed that the SRP and SRP receptor (SR) GTPases from chloroplast (cpSRP54 and cpFtsY, respectively) can interact with one another 400-fold more efficiently than their bacterial homologues, thus providing an explanation as to why this novel chloroplast SRP pathway bypasses the requirement for the SRP RNA. Here we report the crystal structure of cpFtsY from Arabidopsis thaliana at 2.0 \u00c5 resolution. In this chloroplast SR, the N-terminal \"N\" domain is more tightly packed, and a more extensive interaction surface is formed between the GTPase \"G\" domain and the N domain than was previously observed in many of its bacterial homologues. As a result, the overall conformation of apo-cpFtsY is closer to that found in the bacterial SRP\u2022FtsY complex than in free bacterial FtsY, especially with regard to the relative orientation of the N and G domains. In contrast, active-site residues in the G domain are mispositioned, explaining the low basal GTP binding and hydrolysis activity of free cpFtsY. This structure emphasizes proper N\u2013G domain arrangement as a key factor in modulating the efficiency of SRP\u2013receptor interaction and helps account, in part, for the faster kinetics at which the chloroplast SR interacts with its binding partner in the absence of an SRP RNA.", "date": "2008-01-11", "date_type": "published", "publication": "Journal of Molecular Biology", "volume": "375", "number": "2", "publisher": "Elsevier", "pagerange": "425-436", "id_number": "CaltechAUTHORS:20120726-074454154", "issn": "0022-2836", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20120726-074454154", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1016/j.jmb.2007.09.061", "primary_object": { "basename": "mmc1.doc", "url": "https://authors.library.caltech.edu/records/rae6w-ext41/files/mmc1.doc" }, "resource_type": "article", "pub_year": "2008", "author_list": "Chandrasekar, Sowmya; Chartron, Justin; et el." }, { "id": "https://authors.library.caltech.edu/records/w3ve8-ktc16", "eprint_id": 8529, "eprint_status": "archive", "datestamp": "2023-08-22 09:45:02", "lastmod": "2023-10-16 21:32:14", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Walter-Peter", "name": { "family": "Walter", "given": "Peter" } } ] }, "title": "Conformational changes in the GTPase modules of the signal reception particle and its receptor drive initiation of protein translocation", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 The Rockefeller University Press, 2007. \n\nSubmitted: 5 February 2007. Accepted: 1 July 2007. Published online 6 August 2007. \n\nWe thank Ted Powers for help in reconstituting the heterologous protein targeting reaction. \n\nThis work was supported by National Institutes of Health grant GM 32384 to P. Walter and career awards from the Burroughs Wellcome Fund and the Camille and Henry Dreyfus Foundation to S. Shan. P. Walter is an investigator at the Howard Hughes Medical Institute. \n\nFig. S1 shows that the targeting and translocation of pPL occurs on a faster time scale than completion of protein synthesis. Fig. S2 shows that FtsY(47\u2013497) is reduced by about half in translocation of pPL compared with full-length FtsY. Fig. S3 and Tables S1\u2013S7 show additional data for repetitions of the experiments shown in Figs. 2\u20135. Online supplemental material is available at http://www.jcb.org/cgi/content/full/jcb.200702018/DC1.\n\nPublished - SHAjcb07.pdf
Published - SHAjcb07figS1.jpg
Supplemental Material - SHAjcb07figS2.jpg
Supplemental Material - SHAjcb07figS3.jpg
Supplemental Material - SHAjcb07tables.pdf
", "abstract": "During cotranslational protein targeting, two guanosine triphosphatase (GTPase) in the signal recognition particle (SRP) and its receptor (SR) form a unique complex in which hydrolyses of both guanosine triphosphates (GTP) are activated in a shared active site. It was thought that GTP hydrolysis drives the recycling of SRP and SR, but is not crucial for protein targeting. Here, we examined the translocation efficiency of mutant GTPases that block the interaction between SRP and SR at specific stages. Surprisingly, mutants that allow SRP\u2013SR complex assembly but block GTPase activation severely compromise protein translocation. These mutations map to the highly conserved insertion box domain loops that rearrange upon complex formation to form multiple catalytic interactions with the two GTPs. Thus, although GTP hydrolysis is not required, the molecular rearrangements that lead to GTPase activation are essential for protein targeting. Most importantly, our results show that an elaborate rearrangement within the SRP\u2013SR GTPase complex is required to drive the unloading and initiate translocation of cargo proteins.", "date": "2007-08-13", "date_type": "published", "publication": "Journal of Cell Biology", "volume": "178", "number": "4", "publisher": "Rockefeller University Press", "pagerange": "611-620", "id_number": "CaltechAUTHORS:SHAjcb07", "issn": "0021-9525", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:SHAjcb07", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1083/jcb.200702018", "pmcid": "PMC2064468", "primary_object": { "basename": "SHAjcb07.pdf", "url": "https://authors.library.caltech.edu/records/w3ve8-ktc16/files/SHAjcb07.pdf" }, "related_objects": [ { "basename": "SHAjcb07figS1.jpg", "url": "https://authors.library.caltech.edu/records/w3ve8-ktc16/files/SHAjcb07figS1.jpg" }, { "basename": "SHAjcb07figS2.jpg", "url": "https://authors.library.caltech.edu/records/w3ve8-ktc16/files/SHAjcb07figS2.jpg" }, { "basename": "SHAjcb07figS3.jpg", "url": "https://authors.library.caltech.edu/records/w3ve8-ktc16/files/SHAjcb07figS3.jpg" }, { "basename": "SHAjcb07tables.pdf", "url": "https://authors.library.caltech.edu/records/w3ve8-ktc16/files/SHAjcb07tables.pdf" }, { "basename": "medium.png", "url": "https://authors.library.caltech.edu/records/w3ve8-ktc16/files/medium.png" }, { "basename": "small.png", "url": "https://authors.library.caltech.edu/records/w3ve8-ktc16/files/small.png" } ], "resource_type": "article", "pub_year": "2007", "author_list": "Shan, Shu-ou; Chandrasekar, Sowmya; et el." }, { "id": "https://authors.library.caltech.edu/records/qs54e-bvv73", "eprint_id": 8673, "eprint_status": "archive", "datestamp": "2023-08-22 09:23:07", "lastmod": "2023-10-16 21:38:31", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Jaru-Ampornpan-Peera", "name": { "family": "Jaru-Ampornpan", "given": "Peera" } }, { "id": "Chandrasekar-Sowmya", "name": { "family": "Chandrasekar", "given": "Sowmya" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" } ] }, "title": "Efficient Interaction between Two GTPases Allows the Chloroplast SRP Pathway to Bypass the Requirement for an SRP RNA", "ispublished": "pub", "full_text_status": "public", "note": "Copyright \u00a9 2007 by The American Society for Cell Biology. Under the License and Publishing Agreement, authors grant to the general public, effective two months after publication of (i.e.,. the appearance of) the edited manuscript in an online issue of MBoC, the nonexclusive right to copy, distribute, or display the manuscript subject to the terms of the Creative Commons\u2013Noncommercial\u2013Share Alike 3.0 Unported license (http://creativecommons.org/licenses/by-nc-sa/3.0). \n\nSubmitted January 17, 2007; Revised April 18, 2007; Accepted April 20, 2007. Originally published as MBC in Press, 10.1091/mbc.E07-01-0037 on May 2, 2007. \n\nWe thank Dr. Ralph Henry (University of Arkansas) for the bacterial expression vectors for cpSRP54 and cpFtsY, Dr. Judith Campbell and the rest of the Shan laboratory for comments on the manuscript, and Dr. Peter Walter for insightful discussions and intellectual support. S.S. was supported by career awards from the Burroughs Wellcome Fund and the Camile and Henry Dreyfus Foundation. P.J.A. is supported by the Bray fellowship. \n\nThe online version of this article contains supplemental material at MBC Online (http://www.molbiolcell.org).\n\nPublished - JARmbc07.pdf
Supplemental Material - JARmbc07suppmat.doc
", "abstract": "Cotranslational protein targeting to membranes is regulated by two GTPases in the signal recognition particle (SRP) and the SRP receptor; association between the two GTPases is slow and is accelerated 400-fold by the SRP RNA. Intriguingly, the otherwise universally conserved SRP RNA is missing in a novel chloroplast SRP pathway. We found that even in the absence of an SRP RNA, the chloroplast SRP and receptor GTPases can interact efficiently with one another; the kinetics of interaction between the chloroplast GTPases is 400-fold faster than their bacterial homologues, and matches the rate at which the bacterial SRP and receptor interact with the help of SRP RNA. Biochemical analyses further suggest that the chloroplast SRP receptor is pre-organized in a conformation that allows optimal interaction with its binding partner, so that conformational changes during complex formation are minimized. Our results highlight intriguing differences between the classical and chloroplast SRP and SRP receptor GTPases, and help explain how the chloroplast SRP pathway can mediate efficient targeting of proteins to the thylakoid membrane in the absence of the SRP RNA, which plays an indispensable role in all the other SRP pathways.", "date": "2007-07", "date_type": "published", "publication": "Molecular Biology of the Cell", "volume": "18", "number": "7", "publisher": "American Society for Cell Biology", "pagerange": "2636-2645", "id_number": "CaltechAUTHORS:JARmbc07", "issn": "1059-1524", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:JARmbc07", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1091/mbc.E07-01-0037", "pmcid": "PMC1924832", "primary_object": { "basename": "JARmbc07.pdf", "url": "https://authors.library.caltech.edu/records/qs54e-bvv73/files/JARmbc07.pdf" }, "related_objects": [ { "basename": "JARmbc07suppmat.doc", "url": "https://authors.library.caltech.edu/records/qs54e-bvv73/files/JARmbc07suppmat.doc" } ], "resource_type": "article", "pub_year": "2007", "author_list": "Jaru-Ampornpan, Peera; Chandrasekar, Sowmya; et el." }, { "id": "https://authors.library.caltech.edu/records/3hnx3-tw207", "eprint_id": 73893, "eprint_status": "archive", "datestamp": "2023-08-19 15:41:30", "lastmod": "2023-10-24 21:05:04", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Walter-P", "name": { "family": "Walter", "given": "Peter" } } ] }, "title": "Molecular Crosstalk between the Nucleotide Specificity Determinant of the SRP GTPase and the SRP Receptor", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2005 American Chemical Society. \n\nReceived 17 January 2005. Published online 5 April 2005. Published in print 1 April 2005. \n\nThis work was supported by NIH Grant GM 32384 to P.W. P.W.\nis an Investigator of the Howard Hughes Medical Institute. S.S. was a Cancer Research Fund Fellow of the Damon Runyon-Walter Winchell Foundation when this work began and is now a Burroughs Wellcome Fund Fellow. \n\nWe thank Dr. Henry Bourne, Daniel Herschlag, Geeta J. Narlikar, and members of the Walter lab for helpful comments on the manuscript.\n\nSupplemental Material - bi0500980si20050117_065711.pdf
", "abstract": "In signal recognition particle (SRP)-dependent targeting of proteins to the bacterial plasma membrane, two GTPases, Ffh (the SRP GTPase) and FtsY (the receptor GTPase), form a complex in which both proteins reciprocally stimulate each other's GTPase activities. We mutated Asp251 in the Ffh active site to Asn (D251N), converting Ffh to a xanthosine 5'-triphosphate (XTP)-specific protein as has been observed in many other GTPases. Unexpectedly, mutant SRP(D251N) is severely compromised in the formation of an active SRP\u00b7FtsY complex when bound with cognate XTP, and even more surprisingly, mutant SRP(D251N) works better when bound with noncognate GTP. These paradoxical results are explained by a model in which Ffh Asp251 forms a bidentate interaction with not only the bound GTP but also the receptor FtsY across the dimer interface. These interactions form part of the network that seals the lateral entrance to the composite active site at the dimer interface, thereby ensuring the electrostatic and/or structural integrity of the active site and contributing to the formation of an active SRP\u00b7FtsY complex.", "date": "2005-04-26", "date_type": "published", "publication": "Biochemistry", "volume": "44", "number": "16", "publisher": "American Chemical Society", "pagerange": "6214-6222", "id_number": "CaltechAUTHORS:20170131-140941855", "issn": "0006-2960", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20170131-140941855", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM-32384" }, { "agency": "Howard Hughes Medical Institute (HHMI)" }, { "agency": "Damon Runyon-Walter Winchell Cancer Fund" }, { "agency": "Burroughs Wellcome Fund" } ] }, "doi": "10.1021/bi0500980", "primary_object": { "basename": "bi0500980si20050117_065711.pdf", "url": "https://authors.library.caltech.edu/records/3hnx3-tw207/files/bi0500980si20050117_065711.pdf" }, "resource_type": "article", "pub_year": "2005", "author_list": "Shan, Shu-ou and Walter, Peter" }, { "id": "https://authors.library.caltech.edu/records/5ah4e-ts431", "eprint_id": 97890, "eprint_status": "archive", "datestamp": "2023-08-19 15:21:03", "lastmod": "2023-10-18 16:51:04", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Walter-P", "name": { "family": "Walter", "given": "Peter" } } ] }, "title": "Co-translational protein targeting by the signal recognition particle", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2005 Federation of European Biochemical Societies. \n\nIssue Online: 28 November 2004; Version of Record online: 28 November 2004; Manuscript accepted: 22 November 2004; Manuscript revised: 02 November 2004; Manuscript received: 02 November 2004.", "abstract": "The signal recognition particle (SRP) mediates the co\u2010translational targeting of nascent proteins to the eukaryotic endoplasmic reticulum membrane, or the bacterial plasma membrane. During this process, two GTPases, one in the SRP and one in the SRP receptor (SR), form a complex in which both proteins reciprocally activate the GTPase reaction of one another. The recent crystal structures of the T. aquaticus SRP \u00b7 SR complex show that the two GTPases associate via an unusually extensive and highly cooperative interaction surface, and form a composite active site at the interface. GTPase activation proceeds through a unique mechanism, stimulated by both interactions between the twinned GTP molecules across the dimer interface and by conformational rearrangements that position catalytic residues in each active site with respect to the bound substrates. Distinct classes of mutations have been isolated that inhibit specific stages during SRP\u2013SR complex formation and activation, suggesting discrete conformational stages during formation of the active SRP \u00b7 SR complex. Each stage provides a potential control point in the targeting reaction at which regulation by additional components can be exerted, thus ensuring the binding and release of cargo at the appropriate time.", "date": "2005-02-07", "date_type": "published", "publication": "FEBS Letters", "volume": "579", "number": "4", "publisher": "Elsevier", "pagerange": "921-926", "id_number": "CaltechAUTHORS:20190814-113039620", "issn": "0014-5793", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190814-113039620", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "doi": "10.1016/j.febslet.2004.11.049", "resource_type": "article", "pub_year": "2005", "author_list": "Shan, Shu-ou and Walter, Peter" }, { "id": "https://authors.library.caltech.edu/records/tp9w3-1mv13", "eprint_id": 98053, "eprint_status": "archive", "datestamp": "2023-08-22 02:43:41", "lastmod": "2023-10-18 17:00:04", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Chu-Feixia", "name": { "family": "Chu", "given": "Feixia" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Moustakas-Demetri-T", "name": { "family": "Moustakas", "given": "Demetri T." } }, { "id": "Alber-Frank", "name": { "family": "Alber", "given": "Frank" } }, { "id": "Egea-Pascal-F", "name": { "family": "Egea", "given": "Pascal F." } }, { "id": "Stroud-Robert-M", "name": { "family": "Stroud", "given": "Robert M." } }, { "id": "Walter-Peter", "name": { "family": "Walter", "given": "Peter" } }, { "id": "Burlingame-Alma-L", "name": { "family": "Burlingame", "given": "Alma L." } } ] }, "title": "Unraveling the interface of signal recognition particle and its receptor by using chemical cross-linking and tandem mass spectrometry", "ispublished": "pub", "full_text_status": "public", "keywords": "computational modeling; protein\u2013protein interactions; protein targeting; GTPases", "note": "\u00a9 2004 The National Academy of Sciences. Freely available online through the PNAS open access option. \n\nContributed by Peter Walter, October 10, 2004. \n\nWe thank David Maltby and Drs. Lan Huang and Katalin M. Medzihradszky for helpful discussions on MS experiments. This work was supported by National Institutes of Health Grants RR 01614 (to A.L.B.), GM 32384 (to P.W.), GM 60641 (to R.M.S.), RR 12961 (to A.L.B.), and RR 15804 (to A.L.B.). F.C. was a Eugene Cota-Robles graduate student fellow. S.o.S. started as a Damon Runyon\u2013Walter Winchell Cancer Research fellow and now is a Burroughs Wellcome Fund fellow. P.W. is an Investigator of the Howard Hughes Medical Institute. \n\nAuthor contributions: F.C., S.-o.S., D.T.M., F.A., P.F.E., R.M.S., P.W., and A.L.B. designed research; F.C., S.-o.S., D.T.M., F.A., and P.F.E. performed research; F.C., S.-o.S., D.T.M., F.A., and P.F.E. contributed new reagents/analytic tools; F.C., D.T.M., F.A., S.-o.S., and P.F.E. analyzed data; and F.C., S.-o.S., D.T.M., F.A., P.F.E., P.W., R.M.S., and A.L.B. wrote the paper.\n\nPublished - pnas-0407456101.pdf
", "abstract": "Among the methods used to unravel protein interaction surfaces, chemical cross-linking followed by identification of the cross-linked peptides by mass spectrometry has proven especially useful in dynamic and complex systems. During the signal recognition particle (SRP)-dependent targeting of proteins to the bacterial plasma membrane, the specific interaction between Ffh (the protein component of SRP) and FtsY (the SRP receptor) is known to be essential for the efficiency and fidelity of this process. In this work, we studied the Escherichia coli and Thermus aquaticus Ffh\u00b7FtsY complexes by using chemical cross-linking and tandem mass spectrometry to identify nine intermolecular cross-linked peptides. This information was used in conjunction with a previously undescribed model-building approach that combines geometric restraint optimization with macromolecular docking. The resulting model of the Ffh\u00b7FtsY complex is in good agreement with the crystal structure solved shortly thereafter. Intriguingly, four of the cross-linked pairs involve the M domain of Ffh, which is absent from the crystal structure, providing previously undocumented experimental evidence that the M domain is positioned in close proximity to the Ffh\u00b7FtsY interface in the complex.", "date": "2004-11-23", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "101", "number": "47", "publisher": "National Academy of Sciences", "pagerange": "16454-16459", "id_number": "CaltechAUTHORS:20190820-160532216", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190820-160532216", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "RR 01614" }, { "agency": "NIH", "grant_number": "GM 32384" }, { "agency": "NIH", "grant_number": "GM 60641" }, { "agency": "NIH", "grant_number": "RR 12961" }, { "agency": "NIH", "grant_number": "RR 15804" }, { "agency": "Eugene V. Cota-Robles Fellowship" }, { "agency": "Damon Runyon-Walter Winchell Foundation" }, { "agency": "Burroughs Wellcome Fund" }, { "agency": "Howard Hughes Medical Institute (HHMI)" } ] }, "doi": "10.1073/pnas.0407456101", "pmcid": "PMC528904", "primary_object": { "basename": "pnas-0407456101.pdf", "url": "https://authors.library.caltech.edu/records/tp9w3-1mv13/files/pnas-0407456101.pdf" }, "resource_type": "article", "pub_year": "2004", "author_list": "Chu, Feixia; Shan, Shu-ou; et el." }, { "id": "https://authors.library.caltech.edu/records/9vrra-wne13", "eprint_id": 98050, "eprint_status": "archive", "datestamp": "2023-08-19 14:17:24", "lastmod": "2023-10-18 16:59:53", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Stroud-Robert-M", "name": { "family": "Stroud", "given": "Robert M." } }, { "id": "Walter-Peter", "name": { "family": "Walter", "given": "Peter" } } ] }, "title": "Mechanism of Association and Reciprocal Activation of Two GTPases", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2004 Shan et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. \n\nReceived June 9, 2004; Accepted July 26, 2004; Published September 21, 2004. \n\nWe thank Geeta J. Narlikar, Wallace Marshall, Pascal F. Egea, and Niels R. Bradshaw for helpful comments on the manuscript. This work was supported by National Institutes of Health grants to PW and RMS. PW is an investigator of the Howard Hughes Medical Institute. SS was a cancer research fund fellow of the Damon Runyon-Walter Winchell Foundation when this work began and is currently a Burroughs Wellcome Fund fellow. \n\nThe authors have declared that no conflicts of interest exist. \n\nAuthor Contributions: SS and PW conceived and designed the experiments. SS performed the experiments. SS analyzed the data. SS contributed reagents/materials/analysis tools. SS, RMS, and PW wrote the paper.\n\nPublished - pbio.0020320.pdf
", "abstract": "The signal recognition particle (SRP) mediates the cotranslational targeting of nascent proteins to the eukaryotic endoplasmic reticulum membrane or the bacterial plasma membrane. During this process, two GTPases, one in SRP and one in the SRP receptor (named Ffh and FtsY in bacteria, respectively), form a complex in which both proteins reciprocally activate the GTPase reaction of one another. Here, we explore by site-directed mutagenesis the role of 45 conserved surface residues in the Ffh-FtsY interaction. Mutations of a large number of residues at the interface impair complex formation, supporting the importance of an extensive interaction surface. Surprisingly, even after a stable complex is formed, single mutations in FtsY can block the activation of GTP hydrolysis in both active sites. Thus, activation requires conformational changes across the interface that coordinate the positioning of catalytic residues in both GTPase sites. A distinct class of mutants exhibits half-site reactivity and thus allows us to further uncouple the activation of individual GTPases. Our dissection of the activation process suggests discrete conformational stages during formation of the active SRP\u2022SRP receptor complex. Each stage provides a potential control point in the targeting reaction at which regulation by additional components can be exerted, thus ensuring the binding and release of cargo at the appropriate time.", "date": "2004-09-21", "date_type": "published", "publication": "PLoS Biology", "volume": "2", "number": "10", "publisher": "Public Library of Science", "pagerange": "Art. No. e320", "id_number": "CaltechAUTHORS:20190820-153826181", "issn": "1545-7885", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190820-153826181", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH" }, { "agency": "Howard Hughes Medical Institute (HHMI)" }, { "agency": "Damon Runyon-Walter Winchell Foundation" }, { "agency": "Burroughs Wellcome Fund" } ] }, "doi": "10.1371/journal.pbio.0020320", "pmcid": "PMC517823", "primary_object": { "basename": "pbio.0020320.pdf", "url": "https://authors.library.caltech.edu/records/9vrra-wne13/files/pbio.0020320.pdf" }, "resource_type": "article", "pub_year": "2004", "author_list": "Shan, Shu-ou; Stroud, Robert M.; et el." }, { "id": "https://authors.library.caltech.edu/records/sq0k6-c2372", "eprint_id": 98098, "eprint_status": "archive", "datestamp": "2023-08-19 12:58:33", "lastmod": "2023-10-18 17:02:54", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Egea-Pascal-F", "name": { "family": "Egea", "given": "Pascal F." }, "orcid": "0000-0003-4432-8625" }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Napetschnig-Johanna", "name": { "family": "Napetschnig", "given": "Johanna" } }, { "id": "Savage-David-F", "name": { "family": "Savage", "given": "David F." }, "orcid": "0000-0003-0042-2257" }, { "id": "Walter-Peter", "name": { "family": "Walter", "given": "Peter" } }, { "id": "Stroud-Robert-M", "name": { "family": "Stroud", "given": "Robert M." }, "orcid": "0000-0003-2083-5665" } ] }, "title": "Substrate twinning activates the signal recognition particle and its receptor", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2004 Nature Publishing Group. \n\nReceived 25 August 2003; Accepted 25 November 2003; Issue Date\n15 January 2004. \n\nWe thank C. Reyes for invaluable contributions to the initial FtsY mutant design and structure determination of T. aquaticus FtsY\u00b7GMPPNP, and R. Vale, H. Bourne and N. Bradshaw for comments on the manuscript. We acknowledge K. Slep and L. Rice for discussion and advice, and thank J. Holton and G. Meigg for support during data collection at the Advanced Light Source. D.F.S was supported by a Burroughs-Wellcome Fund graduate fellowship. S.S. is supported by a Damon Runyan/Walter Winchell Cancer research fellowship. This work was supported by NIH grants to R.M.S. and P.W. P.W is an Investigator of the Howard Hughes Medical Institute. \n\nThe authors declare that they have no competing financial interests.\n\nSupplemental Material - 41586_2004_BFnature02250_MOESM1_ESM.mov
Supplemental Material - 41586_2004_BFnature02250_MOESM2_ESM.mov
Supplemental Material - 41586_2004_BFnature02250_MOESM3_ESM.doc
", "abstract": "Signal sequences target proteins for secretion from cells or for integration into cell membranes. As nascent proteins emerge from the ribosome, signal sequences are recognized by the signal recognition particle (SRP), which subsequently associates with its receptor (SR). In this complex, the SRP and SR stimulate each other's GTPase activity, and GTP hydrolysis ensures unidirectional targeting of cargo through a translocation pore in the membrane. To define the mechanism of reciprocal activation, we determined the 1.9\u2009\u00c5 structure of the complex formed between these two GTPases. The two partners form a quasi-two-fold symmetrical heterodimer. Biochemical analysis supports the importance of the extensive interaction surface. Complex formation aligns the two GTP molecules in a symmetrical, composite active site, and the 3\u2032OH groups are essential for association, reciprocal activation and catalysis. This unique circle of twinned interactions is severed twice on hydrolysis, leading to complex dissociation after cargo delivery.", "date": "2004-01-14", "date_type": "published", "publication": "Nature", "volume": "427", "number": "6971", "publisher": "Nature Publishing Group", "pagerange": "215-221", "id_number": "CaltechAUTHORS:20190822-090051239", "issn": "0028-0836", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190822-090051239", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Burroughs-Wellcome Fund" }, { "agency": "Damon Runyan-Walter Winchell Cancer Research Fund" }, { "agency": "NIH" }, { "agency": "Howard Hughes Medical Institute (HHMI)" } ] }, "doi": "10.1038/nature02250", "primary_object": { "basename": "41586_2004_BFnature02250_MOESM1_ESM.mov", "url": "https://authors.library.caltech.edu/records/sq0k6-c2372/files/41586_2004_BFnature02250_MOESM1_ESM.mov" }, "related_objects": [ { "basename": "41586_2004_BFnature02250_MOESM2_ESM.mov", "url": "https://authors.library.caltech.edu/records/sq0k6-c2372/files/41586_2004_BFnature02250_MOESM2_ESM.mov" }, { "basename": "41586_2004_BFnature02250_MOESM3_ESM.doc", "url": "https://authors.library.caltech.edu/records/sq0k6-c2372/files/41586_2004_BFnature02250_MOESM3_ESM.doc" } ], "resource_type": "article", "pub_year": "2004", "author_list": "Egea, Pascal F.; Shan, Shu-ou; et el." }, { "id": "https://authors.library.caltech.edu/records/00j7p-wed05", "eprint_id": 98717, "eprint_status": "archive", "datestamp": "2023-08-19 11:20:04", "lastmod": "2023-10-18 17:35:32", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Walter-P", "name": { "family": "Walter", "given": "Peter" } } ] }, "title": "Induced nucleotide specificity in a GTPase", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 2003 The National Academy of Sciences. \n\nEdited by Jennifer A. Doudna, University of California, Berkeley, CA, and approved February 5, 2003 (received for review December 17, 2002). \n\nWe thank A. E. Johnson for advice on fluorescence experiments and T. Powers, R. Gilmore, H. Bourne, R. Stroud, and members of the Walter and Stroud laboratories for comments on the manuscript. This work was supported by National Institutes of Health Grant GM 32384 (to P.W.). P.W. is an Investigator of the Howard Hughes Medical Institute, and S.S. is a Cancer Research Fund Fellow of the Damon Runyon\u2013Walter Winchell Foundation. \n\nThis paper was submitted directly (Track II) to the PNAS office.\n\nPublished - pq0803004480.pdf
Supplemental Material - 7693Fig7.jpg
Supplemental Material - 7693SuppText.pdf
", "abstract": "In signal-recognition particle (SRP)-dependent protein targeting to the bacterial plasma membrane, two GTPases, Ffh (a subunit of the bacterial SRP) and FtsY (the bacterial SRP receptor), act as GTPase activating proteins for one another. The molecular mechanism of this reciprocal GTPase activation is poorly understood. In this work, we show that, unlike other GTPases, free FtsY exhibits only low preference for GTP over other nucleotides. On formation of the SRP\u22c5FtsY complex, however, the nucleotide specificity of FtsY is enhanced 10^3-fold. Thus, interactions with SRP must induce conformational changes that directly affect the FtsY GTP-binding site: in response to SRP binding, FtsY switches from a nonspecific \"open\" state to a \"closed\" state that provides discrimination between cognate and noncognate nucleotides. We propose that this conformational change leads to more accurate positioning of the nucleotide and thus could contribute to activation of FtsY's GTPase activity by a novel mechanism.", "date": "2003-04-15", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "100", "number": "8", "publisher": "National Academy of Sciences", "pagerange": "4480-4485", "id_number": "CaltechAUTHORS:20190918-102339005", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190918-102339005", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM 32384" }, { "agency": "Howard Hughes Medical Institute (HHMI)" }, { "agency": "Damon Runyon-Walter Winchell Foundation" } ] }, "doi": "10.1073/pnas.0737693100", "pmcid": "PMC153581", "primary_object": { "basename": "7693Fig7.jpg", "url": "https://authors.library.caltech.edu/records/00j7p-wed05/files/7693Fig7.jpg" }, "related_objects": [ { "basename": "7693SuppText.pdf", "url": "https://authors.library.caltech.edu/records/00j7p-wed05/files/7693SuppText.pdf" }, { "basename": "pq0803004480.pdf", "url": "https://authors.library.caltech.edu/records/00j7p-wed05/files/pq0803004480.pdf" } ], "resource_type": "article", "pub_year": "2003", "author_list": "Shan, Shu-ou and Walter, Peter" }, { "id": "https://authors.library.caltech.edu/records/62pkc-ptx15", "eprint_id": 98013, "eprint_status": "archive", "datestamp": "2023-08-21 23:29:49", "lastmod": "2023-10-18 16:57:38", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Herschlag-D", "name": { "family": "Herschlag", "given": "Daniel" } } ] }, "title": "Dissection of a metal-ion-mediated conformational change in Tetrahymena ribozyme catalysis", "ispublished": "pub", "full_text_status": "public", "keywords": "conformational change; mechanistic analysis; metal ion; ribozyme; RNA catalysis", "note": "\u00a9 2002 RNA Society. The Authors acknowledge that six months after the full-issue publication date, the Article will be distributed under a Creative Commons CC-BY-NC License (Attribution-NonCommercial 4.0 International License, http://creativecommons.org/licenses/by-nc/4.0/).\n\nPublished online by Cambridge University Press: 20 August 2002. \n\nWe thank Dr. F. Eckstein for the generous gift of 2'-aminoguanosine, L. Beigelman for oligonucleotides, and K. Karbstein and L. Bartley for permission to cite unpublished results. This work was supported by National Institutes of Health Grant GM49243 to D.H.\n\nPublished - 12166641.pdf
", "abstract": "Conformational changes are often required for the biological function of RNA molecules. In the Tetrahymena group I ribozyme reaction, a conformational change has been suggested to occur upon binding of the oligonucleotide substrate (S) or the guanosine nucleophile (G), leading to stronger binding of the second substrate. Recent work showed that the two substrates are bridged by a metal ion that coordinates both the nonbridging reactive phosphoryl oxygen of S and the 2\u2032-OH of G. These results suggest that the energy from the metal ion[bull ]substrate interactions is used to drive the proposed conformational change. In this work, we provide an experimental test for this model. The results provide strong support for the proposed conformational change and for a central role of the bridging metal ion in this change. The results from this work, combined with previous data, allow construction of a two-state model that quantitatively accounts for all of the observations in this and previous work. This model provides a conceptual and quantitative framework that will facilitate understanding and further probing of the energetic and structural features of this conformational change and its role in catalysis.", "date": "2002-07", "date_type": "published", "publication": "RNA", "volume": "8", "number": "7", "publisher": "Cambridge University Press", "pagerange": "861-872", "id_number": "CaltechAUTHORS:20190819-161842072", "issn": "1355-8382", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190819-161842072", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM49243" } ] }, "doi": "10.1017/s1355838202020216", "pmcid": "PMC1370303", "primary_object": { "basename": "12166641.pdf", "url": "https://authors.library.caltech.edu/records/62pkc-ptx15/files/12166641.pdf" }, "resource_type": "article", "pub_year": "2002", "author_list": "Shan, Shu-ou and Herschlag, Daniel" }, { "id": "https://authors.library.caltech.edu/records/c6pwc-74b80", "eprint_id": 99108, "eprint_status": "archive", "datestamp": "2023-08-19 08:39:59", "lastmod": "2023-10-18 17:51:51", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Peluso-P", "name": { "family": "Peluso", "given": "Paul" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Nock-S", "name": { "family": "Nock", "given": "Silke" } }, { "id": "Herschlag-D", "name": { "family": "Herschlag", "given": "Daniel" } }, { "id": "Walter-P", "name": { "family": "Walter", "given": "Peter" } } ] }, "title": "Role of SRP RNA in the GTPase Cycles of Ffh and FtsY", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 2001 American Chemical Society. \n\nReceived August 9, 2001; Revised Manuscript Received October 12, 2001. \n\nThis work is supported by NIH Grants GM 32384 to P.W. and\nGM 26494 to D.H. P.W. is an Investigator of the Howard Hughes Medical Institute, and S.S. is a Cancer Research Fund Fellow of the Damon Runyon-Walter Winchell Foundation. \n\nWe thank C. Murphy for help with setting up the stopped-flow experiments, A. E. Johnson for advice in fluorescence experiments and for communications of unpublished data, D. Freymann for the expression plasmid for Ffh, and members of the Walter lab for comments on the manuscript.", "abstract": "The bacterial homologues of the signal recognition particle (SRP) and its receptor, the Ffh\u20224.5S RNA ribonucleoprotein complex and the FtsY protein, respectively, form a unique complex in which both Ffh and FtsY act as GTPase activating proteins for one another, resulting in the mutual stimulation of GTP hydrolysis by both proteins. Previous work showed that 4.5S RNA enhances the GTPase activity in the presence of both Ffh and FtsY, but it was not clear how this was accomplished. In this work, kinetic and thermodynamic analyses of the GTPase reactions of Ffh and FtsY have provided insights into the role of 4.5S RNA in the GTPase cycles of Ffh and FtsY. We found that 4.5S RNA accelerates the association between Ffh and FtsY 400-fold in their GTP-bound form, analogous to its 200-fold catalytic effect on Ffh\u2022FtsY association previously observed with the GppNHp-bound form [Peluso, P., et al. (2000) Science 288, 1640\u22121643]. Further, Ffh\u2212FtsY association is rate-limiting for the observed GTPase reaction with subsaturating Ffh and FtsY, thereby accounting for the apparent stimulatory effect of 4.5S RNA on the GTPase activity observed previously. An additional step, GTP hydrolysis from the Ffh\u2022FtsY complex, is also moderately facilitated by 4.5S RNA. These results suggest that 4.5S RNA modulates the conformation of the Ffh\u2022FtsY complex and may, in turn, regulate its GTPase activity during the SRP functional cycle.", "date": "2001-12-01", "date_type": "published", "publication": "Biochemistry", "volume": "40", "number": "50", "publisher": "American Chemical Society", "pagerange": "15224-15233", "id_number": "CaltechAUTHORS:20191007-081920896", "issn": "0006-2960", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20191007-081920896", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM 32384" }, { "agency": "NIH", "grant_number": "GM 26494" }, { "agency": "Howard Hughes Medical Institute (HHMI)" }, { "agency": "Damon Runyon-Walter Winchell Foundation" } ] }, "doi": "10.1021/bi011639y", "resource_type": "article", "pub_year": "2001", "author_list": "Peluso, Paul; Shan, Shu-ou; et el." }, { "id": "https://authors.library.caltech.edu/records/km8yn-b3130", "eprint_id": 99103, "eprint_status": "archive", "datestamp": "2023-08-21 21:14:47", "lastmod": "2023-10-18 17:51:37", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Herschlag-D", "name": { "family": "Herschlag", "given": "Daniel" } } ] }, "title": "An unconventional origin of metal-ion rescue and inhibition in the Tetrahymena group I ribozyme reaction", "ispublished": "pub", "full_text_status": "public", "keywords": "chemical modification; enzymatic catalysis; metal ion; ribozyme", "note": "\u00a9 2000 RNA Society. The Authors acknowledge that six months after the full-issue publication date, the Article will be distributed under a Creative Commons CC-BY-NC License (Attribution-NonCommercial 4.0 International License, http://creativecommons.org/licenses/by-nc/4.0/). \n\nManuscript accepted without revision April 5, 2000. \n\nWe thank members of the Piccirilli group for modified oligonucleotides, L. Beigelman for oligonucleotide substrates, and members of the Herschlag group for comments on the manuscript. This work was supported by National Institutes of Health grant GM49243 to D.H.\n\nPublished - 10864040.pdf
", "abstract": "The presence of catalytic metal ions in RNA active sites has often been inferred from metal-ion rescue of modified substrates and sometimes from inhibitory effects of alternative metal ions. Herein we report that, in the Tetrahymena group I ribozyme reaction, the deleterious effect of a thio substitution at the pro-Sp position of the reactive phosphoryl group is rescued by Mn2+. However, analysis of the reaction of this thio substrate and of substrates with other modifications strongly suggest that this rescue does not stem from a direct Mn2+ interaction with the Sp sulfur. Instead, the apparent rescue arises from a Mn2+ ion interacting with the residue immediately 3' of the cleavage site, A(+1), that stabilizes the tertiary interactions between the oligonucleotide substrate (S) and the active site. This metal site is referred to as site D herein. We also present evidence that a previously observed Ca2+ ion that inhibits the chemical step binds to metal site D. These and other observations suggest that, whereas the interactions of Mn2+ at site D are favorable for the chemical reaction, the Ca2+ at site D exerts its inhibitory effect by disrupting the alignment of the substrates within the active site. These results emphasize the vigilance necessary in the design and interpretation of metal-ion rescue and inhibition experiments. Conversely, in-depth mechanistic analysis of the effects of site-specific substrate modifications can allow the effects of specific metal ion-RNA interactions to be revealed and the properties of individual metal-ion sites to be probed, even within the sea of metal ions bound to RNA.", "date": "2000-06", "date_type": "published", "publication": "RNA", "volume": "6", "number": "6", "publisher": "Cold Spring Harbor Laboratory Press", "pagerange": "795-813", "id_number": "CaltechAUTHORS:20191004-154123295", "issn": "1355-8382", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20191004-154123295", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM49243" } ] }, "pmcid": "PMC1369959", "primary_object": { "basename": "10864040.pdf", "url": "https://authors.library.caltech.edu/records/km8yn-b3130/files/10864040.pdf" }, "resource_type": "article", "pub_year": "2000", "author_list": "Shan, Shu-ou and Herschlag, Daniel" }, { "id": "https://authors.library.caltech.edu/records/ppq2q-dm426", "eprint_id": 99104, "eprint_status": "archive", "datestamp": "2023-08-19 04:48:23", "lastmod": "2023-10-18 17:51:39", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Yoshida-Aiichiro", "name": { "family": "Yoshida", "given": "Aiichiro" } }, { "id": "Sun-Sengen", "name": { "family": "Sun", "given": "Sengen" } }, { "id": "Piccirilli-J-A", "name": { "family": "Piccirilli", "given": "Joseph A." }, "orcid": "0000-0002-0541-6270" }, { "id": "Herschlag-D", "name": { "family": "Herschlag", "given": "Daniel" } } ] }, "title": "Three metal ions at the active site of the Tetrahymena group I ribozyme", "ispublished": "pub", "full_text_status": "public", "note": "\u00a9 1999 The National Academy of Sciences. \n\nCommunicated by Olke C. Uhlenbeck, University of Colorado, Boulder, CO, August 27, 1999 (received for review May 13, 1999). \n\nWe are grateful to F. Eckstein for the gift of 2\u2032-aminoguanosine, L. Beigelman for oligonucleotides, G. Narlikar for initial results and intellectual input, J. Brauman and members of the Herschlag laboratory for comments on the manuscript, and a reviewer for suggestions on the description of the experimental approach. This work was supported by grants from the National Institutes of Health to D. H. and the Howard Hughes Medical Institute to J.A.P.; S. Sun is a Research Associate of the Howard Hughes Medical Institute. \n\nThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked \"advertisement\" in accordance with 18 U.S.C. \u00a71734 solely to indicate this fact.\n\nPublished - pq012299.pdf
", "abstract": "Metal ions are critical for catalysis by many RNA and protein enzymes. To understand how these enzymes use metal ions for catalysis, it is crucial to determine how many metal ions are positioned at the active site. We report here an approach, combining atomic mutagenesis with quantitative determination of metal ion affinities, that allows individual metal ions to be distinguished. Using this approach, we show that at the active site of the Tetrahymena group I ribozyme the previously identified metal ion interactions with three substrate atoms, the 3\u2032-oxygen of the oligonucleotide substrate and the 3\u2032- and 2\u2032-moieties of the guanosine nucleophile, are mediated by three distinct metal ions. This approach provides a general tool for distinguishing active site metal ions and allows the properties and roles of individual metal ions to be probed, even within the sea of metal ions bound to RNA.", "date": "1999-10-26", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "96", "number": "22", "publisher": "National Academy of Sciences", "pagerange": "12299-12304", "id_number": "CaltechAUTHORS:20191004-154804302", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20191004-154804302", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH" }, { "agency": "Howard Hughes Medical Institute (HHMI)" } ] }, "doi": "10.1073/pnas.96.22.12299", "pmcid": "PMC22911", "primary_object": { "basename": "pq012299.pdf", "url": "https://authors.library.caltech.edu/records/ppq2q-dm426/files/pq012299.pdf" }, "resource_type": "article", "pub_year": "1999", "author_list": "Shan, Shu-ou; Yoshida, Aiichiro; et el." }, { "id": "https://authors.library.caltech.edu/records/7n5z3-g5d11", "eprint_id": 98010, "eprint_status": "archive", "datestamp": "2023-08-19 04:36:36", "lastmod": "2023-10-18 16:57:30", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Herschlag-D", "name": { "family": "Herschlag", "given": "Daniel" } } ] }, "title": "Probing the Role of Metal Ions in RNA Catalysis:\u00a0 Kinetic and Thermodynamic Characterization of a Metal Ion Interaction with the 2'-Moiety of the Guanosine Nucleophile in the Tetrahymena Group I Ribozyme", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 1999 American Chemical Society. \n\nReceived February 17, 1999; Revised Manuscript Received May 14, 1999. \n\nThis work was supported by NIH Grant GM49243 to D.H. \n\nWe are grateful to members of the Piccirilli group for helpful discussions and technical support, F. Eckstein for the gift of 2'-aminoguanosine, L. Beigelman for oligonucleotide substrates, G. Narlikar for initial results and intellectual input, and members of the Herschlag group for helpful comments on the manuscript.", "abstract": "Deciphering the role of individual metal ions in RNA catalysis is a tremendous challenge, as numerous metal ions coat the charged backbone of a folded RNA. Metal ion specificity switch experiments combined with quantitative analysis may provide a powerful tool for probing specific metal ion\u2212RNA interactions and for delineating the role of individual metal ions among the sea of metal ions bound to RNA. We show herein that Mn^(2+) rescues the deleterious effect of replacing the 2'-OH of the guanosine nucleophile (G) by \u2013NH_2 (G_(NH2)) in the reaction catalyzed by the Tetrahymena group I ribozyme (E), and the Mn^(2+) concentration dependence suggests that a single metal ion is responsible for rescue. This provides strong evidence for a metal ion interaction with the 2'-moiety of G in this ribozyme (referred to as MC), confirming and extending previous results in a bacteriophage group I intron [Sj\u00f6gren, A.-S., Pettersson, E., Sj\u00f6berg, B.-M., and Str\u00f6mberg, R. (1997) Nucleic Acids Res. 25, 648\u2212654]. Toward understanding the >10^6-fold catalytic contribution of the 2'-OH of G, we have determined the individual reaction steps affected by MC and quantitated these effects. Mn_C^(2+) has only a small effect on binding of G_(NH2) to the free ribozyme or ribozyme\u00b7oligonucleotide complexes that lack the reactive phosphoryl group. In contrast, Mn_C^(2+) increases the binding of G_(NH2) to the ribozyme\u00b7oligonucleotide substrate (E\u00b7S) complex 20-fold and increases the binding of S to the E\u00b7G_(NH2) complex by the same amount. These and other observations suggest that MC plays an integral role in the coupled binding of the oligonucleotide substrate and the guanosine nucleophile. This metal ion may be used to align the nucleophile within the active site, thereby facilitating the reaction. Alternatively or in addition, MC may act in concert with an additional metal ion to coordinate and activate the 3'-OH of G. Finally, these experiments have also allowed us to probe the properties of this metal ion site and isolate the energetic effects of the interaction of this specific metal ion with the 2'-moiety of G.", "date": "1999-08-01", "date_type": "published", "publication": "Biochemistry", "volume": "38", "number": "34", "publisher": "American Chemical Society", "pagerange": "10958-10975", "id_number": "CaltechAUTHORS:20190819-152022033", "issn": "0006-2960", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190819-152022033", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM49243" } ] }, "doi": "10.1021/bi990388e", "resource_type": "article", "pub_year": "1999", "author_list": "Shan, Shu-ou and Herschlag, Daniel" }, { "id": "https://authors.library.caltech.edu/records/0ppq8-xe564", "eprint_id": 99106, "eprint_status": "archive", "datestamp": "2023-08-19 04:36:44", "lastmod": "2023-10-18 17:51:46", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Narlikar-G-J", "name": { "family": "Narlikar", "given": "Geeta J." } }, { "id": "Herschlag-D", "name": { "family": "Herschlag", "given": "Daniel" } } ] }, "title": "Protonated 2'-Aminoguanosine as a Probe of the Electrostatic Environment of the Active Site of the Tetrahymena Group I Ribozyme", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 1999 American Chemical Society. \n\nReceived February 17, 1999; Revised Manuscript Received May 14, 1999. \n\nThis work was supported by NIH Grant GM49243 to D.H. \n\nWe are grateful to J. Piccirilli for helpful discussions, F. Eckstein for the gift of 2'-aminoguanosine, L. Beigelman for oligonucleotide substrates, and members of the Herschlag lab for comments on the manuscript.", "abstract": "We have probed the electrostatic environment of the active site of the Tetrahymena group I ribozyme (E) using protonated 2'-aminoguanosine (G_ NH+_3), in which the 2'-OH of the guanosine nucleophile (G) is replaced by an \u2013NH+_3 group. At low concentrations of divalent metal ion (2 mM Mg^(2+)), G_NH+_3 binds at least 200-fold stronger than G or G_(NH2), with a dissociation constant of \u22641 \u03bcM from the ribozyme\u00b7oligonucleotide substrate\u00b7G_NH+_3 complex (E\u2022S\u2022G_(NH+_3). This strong binding suggests that the -NH+_3 group interacts with negatively charged phosphoryl groups within the active site. Increasing the concentration of divalent metal ion weakens the binding of G_(NH+_3) to E\u00b7S more than 10^2-fold. The Mn^(2+) concentration dependence suggests that MC, the metal ion that interacts with the 2'-moiety of G in the normal reaction, is responsible for this effect. M_C and G_(NH+_3) compete for binding to the active site; this competition could arise from electrostatic repulsion between the positively charged -NH+_3 and M_C and, possibly, from their competition for interaction with active site phosphoryl groups. The reactive phosphoryl group of S increases the competition between M_C and G_(NH+_3), consistent with a network of interactions involving MC that help position the reactive phosphoryl group and the guanosine nucleophile with respect to one another. The chemical step with bound G_(NH+_3) is at least 10^4-fold slower than with G or G_(NH2). These results provide additional support for an integral role of M_C in catalysis by the Tetrahymena ribozyme, and demonstrate the utility of the -NH+_3 moiety as an electrostatic probe within a structured RNA.", "date": "1999-08-01", "date_type": "published", "publication": "Biochemistry", "volume": "38", "number": "34", "publisher": "American Chemical Society", "pagerange": "10976-10988", "id_number": "CaltechAUTHORS:20191004-161147327", "issn": "0006-2960", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20191004-161147327", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM49243" } ] }, "doi": "10.1021/bi9903897", "resource_type": "article", "pub_year": "1999", "author_list": "Shan, Shu-ou; Narlikar, Geeta J.; et el." }, { "id": "https://authors.library.caltech.edu/records/7qqas-b4g56", "eprint_id": 99105, "eprint_status": "archive", "datestamp": "2023-08-22 11:24:14", "lastmod": "2023-10-18 17:51:42", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Herschlag-D", "name": { "family": "Herschlag", "given": "Daniel" } } ] }, "title": "The change in hydrogen bond strength accompanying charge rearrangement: Implications for enzymatic catalysis", "ispublished": "pub", "full_text_status": "public", "keywords": "hydrogen bonding; active site environments", "note": "\u00a9 1996 The National Academy of Sciences of the USA. \n\nCommunicated by John I. Brauman, Stanford University, Stanford, CA, September 9, 1996 (received for review June 14, 1996). \n\nWe thank J. Brauman, W. Cleland, M. Kreevoy, and A. Williams for discussions; the Bordwell group for technical help; and P. Frey for permission to cite unpublished work. This work was supported by grants from the Lucille P. Markey Charitable Trust and the Chicago Community Trust to D.H. D.H. is a Lucille P. Markey Scholar and a Searle Scholar. \n\nThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked \"advertisement\" in accordance with 18 U.S.C. \u00a71734 solely to indicate this fact.\n\nPublished - pq014474.pdf
", "abstract": "The equilibrium for formation of the intramolecular hydrogen bond (K^(HB)) in a series of substituted salicylate monoanions was investigated as a function of \u0394pK_a, the difference between the pK_a values of the hydrogen bond donor and acceptor, in both water and dimethyl sulfoxide. The dependence of log K^(HB) upon \u0394pK_a is linear in both solvents, but is steeper in dimethyl sulfoxide (slope = 0.73) than in water (slope = 0.05). Thus, hydrogen bond strength can undergo substantially larger increases in nonaqueous media than aqueous solutions as the charge density on the donor or acceptor atom increases. These results support a general mechanism for enzymatic catalysis, in which hydrogen bonding to a substrate is strengthened as charge rearranges in going from the ground state to the transition state; the strengthening of the hydrogen bond would be greater in a nonaqueous enzymatic active site than in water, thus providing a rate enhancement for an enzymatic reaction relative to the solution reaction. We suggest that binding energy of an enzyme is used to fix the substrate in the low-dielectric active site, where the strengthening of the hydrogen bond in the course of a reaction is increased.", "date": "1996-12-10", "date_type": "published", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "volume": "93", "number": "25", "publisher": "National Academy of Sciences", "pagerange": "14474-14479", "id_number": "CaltechAUTHORS:20191004-160446800", "issn": "0027-8424", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20191004-160446800", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Lucille P. Markey Charitable Trust" }, { "agency": "Chicago Community Trust" }, { "agency": "Searle Scholars Program" } ] }, "doi": "10.1073/pnas.93.25.14474", "pmcid": "PMC26157", "primary_object": { "basename": "pq014474.pdf", "url": "https://authors.library.caltech.edu/records/7qqas-b4g56/files/pq014474.pdf" }, "resource_type": "article", "pub_year": "1996", "author_list": "Shan, Shu-ou and Herschlag, Daniel" }, { "id": "https://authors.library.caltech.edu/records/jsxt6-xsr98", "eprint_id": 97905, "eprint_status": "archive", "datestamp": "2023-08-20 07:28:54", "lastmod": "2023-10-18 16:52:04", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Loh-Stewart", "name": { "family": "Loh", "given": "Stewart" } }, { "id": "Herschlag-D", "name": { "family": "Herschlag", "given": "Daniel" } } ] }, "title": "The Energetics of Hydrogen Bonds in Model Systems: Implications for Enzymatic Catalysis", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 1996 American Association for the Advancement of Science. \n\nReceived 7 November 1995; accepted 14 February 1996. \n\nWe thank J. Brauman, J. Klinman, and J. Kirsch for discussions and helpful suggestions, the Bordwell group for technical help, and the Herschlag lab for comments on the manuscript. This work is supported by grants from the Lucille P. Markey Charitable Trust and the Chicago Community Trust to D.H. D.H. is a Lucille P. Markey Scholar and a Searle Scholar.", "abstract": "Low-barrier or short, strong hydrogen bonds have been proposed to contribute 10 to 20 kilocalories per mole to transition-state stabilization in enzymatic catalysis. The proposal invokes a large increase in hydrogen bond energy when the pK_a values of the donor and acceptor (where K_a is the acid constant) become matched in the transition state (\u0394pK_a = 0). This hypothesis was tested by investigating the energetics of hydrogen bonds as a function of \u0394pK_a for homologous series of compounds under nonaqueous conditions that are conducive to the formation of low-barrier hydrogen bonds. In all cases, there was a linear correlation between the increase in hydrogen-bond energy and the decrease in \u0394pK_a, as expected from simple electrostatic effects. However, no additional energetic contribution to the hydrogen bond was observed at \u0394pK_a = 0. These results and those of other model studies suggest alternative mechanisms by which hydrogen bonds can contribute to enzymatic catalysis, in accord with conventional electrostatic considerations.", "date": "1996-04-05", "date_type": "published", "publication": "Science", "volume": "272", "number": "5258", "publisher": "American Association for the Advancement of Science", "pagerange": "97-101", "id_number": "CaltechAUTHORS:20190814-161721487", "issn": "0036-8075", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190814-161721487", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "Lucille P. Markey Charitable Trust" }, { "agency": "Chicago Community Trust" }, { "agency": "Searle Scholars Program" } ] }, "doi": "10.1126/science.272.5258.97", "resource_type": "article", "pub_year": "1996", "author_list": "Shan, Shu-ou; Loh, Stewart; et el." }, { "id": "https://authors.library.caltech.edu/records/xkz07-p6n94", "eprint_id": 99034, "eprint_status": "archive", "datestamp": "2023-08-20 01:47:13", "lastmod": "2023-10-18 17:48:22", "type": "article", "metadata_visibility": "show", "creators": { "items": [ { "id": "Zhang-Pinghui", "name": { "family": "Zhang", "given": "Pinghui" } }, { "id": "Liu-Suxing", "name": { "family": "Liu", "given": "Suxing" } }, { "id": "Shan-S-O", "name": { "family": "Shan", "given": "Shu-ou" }, "orcid": "0000-0002-6526-1733" }, { "id": "Ji-Xinhua", "name": { "family": "Ji", "given": "Xinhua" } }, { "id": "Gilliland-G-L", "name": { "family": "Gilliland", "given": "Gary L." } }, { "id": "Armstrong-R-N", "name": { "family": "Armstrong", "given": "Richard N." } } ] }, "title": "Modular mutagenesis of exons 1, 2, and 8 of a glutathione S-transferase from the Mu class. Mechanistic and structural consequences for chimeras of isoenzyme 3-3", "ispublished": "pub", "full_text_status": "restricted", "note": "\u00a9 1992 American Chemical Society. \n\nReceived May 26, 1992; Revised Manuscript Received July 22, 1992. \n\nThis work was supported by a grant from the National Institutes of Health (GM 30910). \n\nThe authors thank Dr. Cecil Pickett and Professor David Tu and their research groups for generously providing the original cDNA clones from which the expression vectors were constructed and William W. Johnson for help with the fluorescence titrations.", "abstract": "Exons 1 and 2 and exon 8 of the mu class GSH transferases from rat encode sequence-variable regions 1 and 4 of mu class isoenzymes, respectively. These two of four variable regions are located at the N- and C-termini of this isoenzyme class and impinge on the active site. In order to assess the influence of these variable regions on the catalytic diversity of the class mu isoenzymes, seven chimeric isoenzymes were constructed by transplantation of the variable regions of the sequence of the type 4 subunit into the corresponding regions of the type 3 subunit. The chimeric isoenzymes exhibit unique catalytic properties. Replacement of all, or part, of variable region 4 of the type 3 subunit with that of the type 4 subunit results in chimeric catalysts with higher turnover numbers in nucleophilic aromatic substitution reactions. Analysis of the crystal structure of isoenzyme 3-3 [Ji, X., Zhang, P., Armstrong, R. N., & Gilliland, G. L. (1992) Biochemistry (preceding paper in this issue)] suggests that interaction of the flexible C-terminal tail with the N-terminal domain helps limit the rate of product release from the active site of isoenzyme 3-3 in this type of reaction. Substitution of all, or part, of the sequence-variable region 1 of subunit 3 with that of subunit 4 results in chimeric isoenzymes that mimic the high stereoselectivity but not the catalytic efficiency of isoenzyme 4-4 toward a,/3-unsaturated ketones. Modular mutagenesis of the N-terminal variable region together with linear free energy relationships using para-substituted 4-phenyl-3-buten-2-ones and the crystal structure of isoenzyme 3-3 has allowed the rapid location of a mutation, V9I, which appears to be crucial in determining the stereoselectivity of the chimeras and, by implication, isoenzyme 4-4 toward phenylbutenones and arene oxides.", "date": "1992-10-01", "date_type": "published", "publication": "Biochemistry", "volume": "31", "number": "42", "publisher": "American Chemical Society", "pagerange": "10185-10193", "id_number": "CaltechAUTHORS:20191002-154651520", "issn": "0006-2960", "official_url": "https://resolver.caltech.edu/CaltechAUTHORS:20191002-154651520", "rights": "No commercial reproduction, distribution, display or performance rights in this work are provided.", "funders": { "items": [ { "agency": "NIH", "grant_number": "GM 30910" } ] }, "doi": "10.1021/bi00157a005", "resource_type": "article", "pub_year": "1992", "author_list": "Zhang, Pinghui; Liu, Suxing; et el." } ]