An integrated workflow for crosslinking mass spectrometry

Marta L. Mendes; Lutz Fischer; Zhuo A. Chen; Marta Barbon; Francis J. O'Reilly; Sven H. Giese; Michael Bohlke-Schneider; Adam Belsom; Therese Dau; Colin W. Combe; Martin Graham; Markus R. Eisele; Wolfgang Baumeister; Christian Speck; Juri Rappsilber

doi:10.15252/msb.20198994

An integrated workflow for crosslinking mass spectrometry

Marta L. Mendes, Lutz Fischer, Zhuo A. Chen, Marta Barbon, Francis J. O'Reilly, Sven H. Giese, Michael Bohlke-Schneider, Adam Belsom, Therese Dau, Colin W. Combe, Martin Graham, Markus R. Eisele, Wolfgang Baumeister, Christian Speck, Juri Rappsilber^*

^*Corresponding author for this work

Proteomics of Cellular Signaling

Research output: Contribution to journal › Article › Research › peer-review

70 Citations (Scopus)

Abstract

We present a concise workflow to enhance the mass spectrometric detection of crosslinked peptides by introducing sequential digestion and the crosslink identification software xiSEARCH. Sequential digestion enhances peptide detection by selective shortening of long tryptic peptides. We demonstrate our simple 12-fraction protocol for crosslinked multi-protein complexes and cell lysates, quantitative analysis, and high-density crosslinking, without requiring specific crosslinker features. This overall approach reveals dynamic protein–protein interaction sites, which are accessible, have fundamental functional relevance and are therefore ideally suited for the development of small molecule inhibitors.

Original language	English
Article number	e8994
Journal	Molecular Systems Biology
Volume	15
Issue number	9
DOIs	https://doi.org/10.15252/msb.20198994
Publication status	Published - 1 Sept 2019

Keywords

crosslinking mass spectrometry
protein–protein interactions
proteomics
software
structural biology

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10.15252/msb.20198994

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title = "An integrated workflow for crosslinking mass spectrometry",

abstract = "We present a concise workflow to enhance the mass spectrometric detection of crosslinked peptides by introducing sequential digestion and the crosslink identification software xiSEARCH. Sequential digestion enhances peptide detection by selective shortening of long tryptic peptides. We demonstrate our simple 12-fraction protocol for crosslinked multi-protein complexes and cell lysates, quantitative analysis, and high-density crosslinking, without requiring specific crosslinker features. This overall approach reveals dynamic protein–protein interaction sites, which are accessible, have fundamental functional relevance and are therefore ideally suited for the development of small molecule inhibitors.",

keywords = "crosslinking mass spectrometry, protein–protein interactions, proteomics, software, structural biology",

author = "Mendes, {Marta L.} and Lutz Fischer and Chen, {Zhuo A.} and Marta Barbon and O'Reilly, {Francis J.} and Giese, {Sven H.} and Michael Bohlke-Schneider and Adam Belsom and Therese Dau and Combe, {Colin W.} and Martin Graham and Eisele, {Markus R.} and Wolfgang Baumeister and Christian Speck and Juri Rappsilber",

note = "Publisher Copyright: {\textcopyright} 2019 The Authors. Published under the terms of the CC BY 4.0 license",

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doi = "10.15252/msb.20198994",

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T1 - An integrated workflow for crosslinking mass spectrometry

AU - Mendes, Marta L.

AU - Fischer, Lutz

AU - Chen, Zhuo A.

AU - Barbon, Marta

AU - O'Reilly, Francis J.

AU - Giese, Sven H.

AU - Bohlke-Schneider, Michael

AU - Belsom, Adam

AU - Dau, Therese

AU - Combe, Colin W.

AU - Graham, Martin

AU - Eisele, Markus R.

AU - Baumeister, Wolfgang

AU - Speck, Christian

AU - Rappsilber, Juri

PY - 2019/9/1

Y1 - 2019/9/1

N2 - We present a concise workflow to enhance the mass spectrometric detection of crosslinked peptides by introducing sequential digestion and the crosslink identification software xiSEARCH. Sequential digestion enhances peptide detection by selective shortening of long tryptic peptides. We demonstrate our simple 12-fraction protocol for crosslinked multi-protein complexes and cell lysates, quantitative analysis, and high-density crosslinking, without requiring specific crosslinker features. This overall approach reveals dynamic protein–protein interaction sites, which are accessible, have fundamental functional relevance and are therefore ideally suited for the development of small molecule inhibitors.

AB - We present a concise workflow to enhance the mass spectrometric detection of crosslinked peptides by introducing sequential digestion and the crosslink identification software xiSEARCH. Sequential digestion enhances peptide detection by selective shortening of long tryptic peptides. We demonstrate our simple 12-fraction protocol for crosslinked multi-protein complexes and cell lysates, quantitative analysis, and high-density crosslinking, without requiring specific crosslinker features. This overall approach reveals dynamic protein–protein interaction sites, which are accessible, have fundamental functional relevance and are therefore ideally suited for the development of small molecule inhibitors.

KW - crosslinking mass spectrometry

KW - protein–protein interactions

KW - proteomics

KW - software

KW - structural biology

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An integrated workflow for crosslinking mass spectrometry

Abstract

Keywords

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