Global membrane protein interactome analysis using in vivo crosslinking and MS-based protein correlation profiling

Mark Larance, Kathryn J. Kirkwood, Michele Tinti, Alejandro Brenes Murillo, Michael Ferguson, Angus Lamond (Lead / Corresponding author)

Research output: Contribution to journalArticle

28 Citations (Scopus)
130 Downloads (Pure)

Abstract

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). Raw data are also available via ProteomeXchange (identifier PXD003754).
Original languageEnglish
Pages (from-to)2476-2490
Number of pages15
JournalMolecular & Cellular Proteomics
Volume15
Issue number7
Early online date25 Apr 2016
DOIs
Publication statusPublished - 1 Jul 2016

Keywords

  • membrane protein
  • interactome analysis
  • proteomics
  • protein complexes
  • protein cross-linking
  • mass spectrometry (MS)

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