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Complex-centric proteome profiling by SEC-SWATH-MS for the parallel detection of hundreds of protein complexes

Bludau, Isabell ; Heusel, Moritz LU ; Frank, Max ; Rosenberger, George ; Hafen, Robin ; Banaei-Esfahani, Amir ; van Drogen, Audrey ; Collins, Ben C. ; Gstaiger, Matthias and Aebersold, Ruedi (2020) In Nature Protocols 15(8). p.2341-2386
Abstract

Most catalytic, structural and regulatory functions of the cell are carried out by functional modules, typically complexes containing or consisting of proteins. The composition and abundance of these complexes and the quantitative distribution of specific proteins across different modules are therefore of major significance in basic and translational biology. However, detection and quantification of protein complexes on a proteome-wide scale is technically challenging. We have recently extended the targeted proteomics rationale to the level of native protein complex analysis (complex-centric proteome profiling). The complex-centric workflow described herein consists of size exclusion chromatography (SEC) to fractionate native protein... (More)

Most catalytic, structural and regulatory functions of the cell are carried out by functional modules, typically complexes containing or consisting of proteins. The composition and abundance of these complexes and the quantitative distribution of specific proteins across different modules are therefore of major significance in basic and translational biology. However, detection and quantification of protein complexes on a proteome-wide scale is technically challenging. We have recently extended the targeted proteomics rationale to the level of native protein complex analysis (complex-centric proteome profiling). The complex-centric workflow described herein consists of size exclusion chromatography (SEC) to fractionate native protein complexes, data-independent acquisition mass spectrometry to precisely quantify the proteins in each SEC fraction based on a set of proteotypic peptides and targeted, complex-centric analysis where prior information from generic protein interaction maps is used to detect and quantify protein complexes with high selectivity and statistical error control via the computational framework CCprofiler (https://github.com/CCprofiler/CCprofiler). Complex-centric proteome profiling captures most proteins in complex-assembled state and reveals their organization into hundreds of complexes and complex variants observable in a given cellular state. The protocol is applicable to cultured cells and can potentially also be adapted to primary tissue and does not require any genetic engineering of the respective sample sources. At present, it requires ~8 d of wet-laboratory work, 15 d of mass spectrometry measurement time and 7 d of computational analysis.

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author
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organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Nature Protocols
volume
15
issue
8
pages
46 pages
publisher
Nature Publishing Group
external identifiers
  • scopus:85088826886
  • pmid:32690956
ISSN
1754-2189
DOI
10.1038/s41596-020-0332-6
language
English
LU publication?
yes
id
379eef6d-074f-4089-b46d-c90c20be5ef8
date added to LUP
2020-08-10 08:46:06
date last changed
2024-06-26 20:07:48
@article{379eef6d-074f-4089-b46d-c90c20be5ef8,
  abstract     = {{<p>Most catalytic, structural and regulatory functions of the cell are carried out by functional modules, typically complexes containing or consisting of proteins. The composition and abundance of these complexes and the quantitative distribution of specific proteins across different modules are therefore of major significance in basic and translational biology. However, detection and quantification of protein complexes on a proteome-wide scale is technically challenging. We have recently extended the targeted proteomics rationale to the level of native protein complex analysis (complex-centric proteome profiling). The complex-centric workflow described herein consists of size exclusion chromatography (SEC) to fractionate native protein complexes, data-independent acquisition mass spectrometry to precisely quantify the proteins in each SEC fraction based on a set of proteotypic peptides and targeted, complex-centric analysis where prior information from generic protein interaction maps is used to detect and quantify protein complexes with high selectivity and statistical error control via the computational framework CCprofiler (https://github.com/CCprofiler/CCprofiler). Complex-centric proteome profiling captures most proteins in complex-assembled state and reveals their organization into hundreds of complexes and complex variants observable in a given cellular state. The protocol is applicable to cultured cells and can potentially also be adapted to primary tissue and does not require any genetic engineering of the respective sample sources. At present, it requires ~8 d of wet-laboratory work, 15 d of mass spectrometry measurement time and 7 d of computational analysis.</p>}},
  author       = {{Bludau, Isabell and Heusel, Moritz and Frank, Max and Rosenberger, George and Hafen, Robin and Banaei-Esfahani, Amir and van Drogen, Audrey and Collins, Ben C. and Gstaiger, Matthias and Aebersold, Ruedi}},
  issn         = {{1754-2189}},
  language     = {{eng}},
  number       = {{8}},
  pages        = {{2341--2386}},
  publisher    = {{Nature Publishing Group}},
  series       = {{Nature Protocols}},
  title        = {{Complex-centric proteome profiling by SEC-SWATH-MS for the parallel detection of hundreds of protein complexes}},
  url          = {{http://dx.doi.org/10.1038/s41596-020-0332-6}},
  doi          = {{10.1038/s41596-020-0332-6}},
  volume       = {{15}},
  year         = {{2020}},
}