期刊
MOLECULAR SYSTEMS BIOLOGY
卷 15, 期 9, 页码 -出版社
WILEY
DOI: 10.15252/msb.20198994
关键词
crosslinking mass spectrometry; protein-protein interactions; proteomics; software; structural biology
资金
- International Post-Doc Initiative-IPODI
- European Union
- Einstein Foundation
- DFG [RA 2365/4-1]
- Wellcome Trust [103139, 107903/Z/15/Z, 108504, 203149]
- Biotechnology and Biological Sciences Research Council UK [BB/N000323/1]
- Medical Research Council UK [MC_U120085811]
- Wellcome Trust [107903/Z/15/Z] Funding Source: Wellcome Trust
- BBSRC [BB/S001387/1, BB/N000323/1] Funding Source: UKRI
- MRC [MC_U120085811] Funding Source: UKRI
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.
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