We present a methodology using in vivo crosslinking combined with HPLC-MS for the global analysis of endogenous protein complexes by protein correlation profiling. Formaldehyde crosslinked protein complexes were extracted with high yield using denaturing buffers that maintained complex solubility during chromatographic separation. We show this efficiently detects both integral membrane and membrane-associated protein complexes, in addition to soluble complexes, allowing identification and analysis of complexes not accessible in native extracts. We compare the protein complexes detected by HPLC-MS protein correlation profiling in both native and formaldehyde crosslinked U2OS cell extracts. These proteome-wide datasets of both in vivo crosslinked and native protein complexes from U2OS cells are freely available via a searchable online database (www.peptracker.com/epd).
[dataset license: CC0 1.0 Universal (CC0 1.0)]
Keywords: membrane protein ; interactome analysis ; proteomics ; protein complexes ; protein crosslinking ; mass spectrometry
Principal Investigators: (in alphabetical order) |
Dr Angus Lamond |
Submitting User: | ccms |
Larance M, Kirkwood KJ, Tinti M, Brenes Murillo A, Ferguson MA, Lamond AI.
Global Membrane Protein Interactome Analysis using In vivo Crosslinking and Mass Spectrometry-based Protein Correlation Profiling.
Mol. Cell Proteomics. 2016 Jul;15(7):2476-90. Epub 2016 Apr 25.
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