Affinity proteomics to study endogenous protein complexes: Pointers, pitfalls, preferences and perspectives

John LaCava*, Kelly R. Molloy, Martin S. Taylor, Michal Domanski, Brian T. Chait, Michael P. Rout

*Corresponding author for this work

Research output: Contribution to journalReview articlepeer-review

45 Citations (Scopus)
47 Downloads (Pure)

Abstract

Dissecting and studying cellular systems requires the ability to specifically isolate distinct proteins along with the co-assembled constituents of their associated complexes. Affinity capture techniques leverage high affinity, high specificity reagents to target and capture proteins of interest along with specifically associated proteins from cell extracts. Affinity capture coupled to mass spectrometry (MS)-based proteomic analyses has enabled the isolation and characterization of a wide range of endogenous protein complexes. Here, we outline effective procedures for the affinity capture of protein complexes, highlighting best practices and common pitfalls.

Original languageEnglish
Pages (from-to)103-119
Number of pages14
JournalBiotechniques
Volume58
Issue number3
DOIs
Publication statusPublished - Mar-2015
Externally publishedYes

Keywords

  • protein complex
  • protein purification
  • affinity
  • proteomics
  • interactomics
  • NUCLEAR-PORE COMPLEX
  • PURIFICATION-MASS-SPECTROMETRY
  • INDUCIBLE GENE-EXPRESSION
  • EPSTEIN-BARR-VIRUS
  • MAMMALIAN-CELLS
  • SACCHAROMYCES-CEREVISIAE
  • MOLECULAR ARCHITECTURE
  • BAC TRANSGENEOMICS
  • ENHANCED DETECTION
  • RAPID PRODUCTION

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