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Profiling the Escherichia coli membrane protein interactome captured in Peptidisc libraries
Protein-correlation-profiling (PCP), in combination with quantitative proteomics, has emerged as a high-throughput method for the rapid identification of dynamic protein complexes in native conditions. While PCP has been successfully applied to soluble proteomes, characterization of the membrane int...
Autores principales: | , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
eLife Sciences Publications, Ltd
2019
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6697469/ https://www.ncbi.nlm.nih.gov/pubmed/31364989 http://dx.doi.org/10.7554/eLife.46615 |
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author | Carlson, Michael Luke Stacey, R Greg Young, John William Wason, Irvinder Singh Zhao, Zhiyu Rattray, David G Scott, Nichollas Kerr, Craig H Babu, Mohan Foster, Leonard J Duong Van Hoa, Franck |
author_facet | Carlson, Michael Luke Stacey, R Greg Young, John William Wason, Irvinder Singh Zhao, Zhiyu Rattray, David G Scott, Nichollas Kerr, Craig H Babu, Mohan Foster, Leonard J Duong Van Hoa, Franck |
author_sort | Carlson, Michael Luke |
collection | PubMed |
description | Protein-correlation-profiling (PCP), in combination with quantitative proteomics, has emerged as a high-throughput method for the rapid identification of dynamic protein complexes in native conditions. While PCP has been successfully applied to soluble proteomes, characterization of the membrane interactome has lagged, partly due to the necessary use of detergents to maintain protein solubility. Here, we apply the peptidisc, a ‘one-size fits all’ membrane mimetic, for the capture of the Escherichia coli cell envelope proteome and its high-resolution fractionation in the absence of detergent. Analysis of the SILAC-labeled peptidisc library via PCP allows generation of over 4900 possible binary interactions out of >700,000 random associations. Using well-characterized membrane protein systems such as the SecY translocon, the Bam complex and the MetNI transporter, we demonstrate that our dataset is a useful resource for identifying transient and surprisingly novel protein interactions. For example, we discover a trans-periplasmic supercomplex comprising subunits of the Bam and Sec machineries, including membrane-bound chaperones YfgM and PpiD. We identify RcsF and OmpA as bone fide interactors of BamA, and we show that MetQ association with the ABC transporter MetNI depends on its N-terminal lipid anchor. We also discover NlpA as a novel interactor of MetNI complex. Most of these interactions are largely undetected by standard detergent-based purification. Together, the peptidisc workflow applied to the proteomic field is emerging as a promising novel approach to characterize membrane protein interactions under native expression conditions and without genetic manipulation. |
format | Online Article Text |
id | pubmed-6697469 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2019 |
publisher | eLife Sciences Publications, Ltd |
record_format | MEDLINE/PubMed |
spelling | pubmed-66974692019-08-19 Profiling the Escherichia coli membrane protein interactome captured in Peptidisc libraries Carlson, Michael Luke Stacey, R Greg Young, John William Wason, Irvinder Singh Zhao, Zhiyu Rattray, David G Scott, Nichollas Kerr, Craig H Babu, Mohan Foster, Leonard J Duong Van Hoa, Franck eLife Biochemistry and Chemical Biology Protein-correlation-profiling (PCP), in combination with quantitative proteomics, has emerged as a high-throughput method for the rapid identification of dynamic protein complexes in native conditions. While PCP has been successfully applied to soluble proteomes, characterization of the membrane interactome has lagged, partly due to the necessary use of detergents to maintain protein solubility. Here, we apply the peptidisc, a ‘one-size fits all’ membrane mimetic, for the capture of the Escherichia coli cell envelope proteome and its high-resolution fractionation in the absence of detergent. Analysis of the SILAC-labeled peptidisc library via PCP allows generation of over 4900 possible binary interactions out of >700,000 random associations. Using well-characterized membrane protein systems such as the SecY translocon, the Bam complex and the MetNI transporter, we demonstrate that our dataset is a useful resource for identifying transient and surprisingly novel protein interactions. For example, we discover a trans-periplasmic supercomplex comprising subunits of the Bam and Sec machineries, including membrane-bound chaperones YfgM and PpiD. We identify RcsF and OmpA as bone fide interactors of BamA, and we show that MetQ association with the ABC transporter MetNI depends on its N-terminal lipid anchor. We also discover NlpA as a novel interactor of MetNI complex. Most of these interactions are largely undetected by standard detergent-based purification. Together, the peptidisc workflow applied to the proteomic field is emerging as a promising novel approach to characterize membrane protein interactions under native expression conditions and without genetic manipulation. eLife Sciences Publications, Ltd 2019-07-31 /pmc/articles/PMC6697469/ /pubmed/31364989 http://dx.doi.org/10.7554/eLife.46615 Text en © 2019, Carlson et al http://creativecommons.org/licenses/by/4.0/ http://creativecommons.org/licenses/by/4.0/This article is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use and redistribution provided that the original author and source are credited. |
spellingShingle | Biochemistry and Chemical Biology Carlson, Michael Luke Stacey, R Greg Young, John William Wason, Irvinder Singh Zhao, Zhiyu Rattray, David G Scott, Nichollas Kerr, Craig H Babu, Mohan Foster, Leonard J Duong Van Hoa, Franck Profiling the Escherichia coli membrane protein interactome captured in Peptidisc libraries |
title | Profiling the Escherichia coli membrane protein interactome captured in Peptidisc libraries |
title_full | Profiling the Escherichia coli membrane protein interactome captured in Peptidisc libraries |
title_fullStr | Profiling the Escherichia coli membrane protein interactome captured in Peptidisc libraries |
title_full_unstemmed | Profiling the Escherichia coli membrane protein interactome captured in Peptidisc libraries |
title_short | Profiling the Escherichia coli membrane protein interactome captured in Peptidisc libraries |
title_sort | profiling the escherichia coli membrane protein interactome captured in peptidisc libraries |
topic | Biochemistry and Chemical Biology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6697469/ https://www.ncbi.nlm.nih.gov/pubmed/31364989 http://dx.doi.org/10.7554/eLife.46615 |
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