title: Affinity proteomics to study endogenous protein complexes: pointers, pitfalls, preferences and perspectives.
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.
authors.affiliation: Laboratory of Cellular and Structural Biology, The Rockefeller University, New York.|Institute for Systems Genetics, New York University School of Medicine, New York, NY.
authors.forename: John
authors.initials: J
authors.lastname: LaCava
authors.affiliation: Laboratory of Mass Spectrometry and Gaseous Ion Chemistry, The Rockefeller University, New York, NY.
authors.forename: Kelly R
authors.initials: KR
authors.lastname: Molloy
authors.affiliation: High Throughput Biology Center and Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, Baltimore, MD.
authors.forename: Martin S
authors.initials: MS
authors.lastname: Taylor
authors.affiliation: Laboratory of Cellular and Structural Biology, The Rockefeller University, New York.|Centre for mRNP Biogenesis and Metabolism, Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark.
authors.forename: Michal
authors.initials: M
authors.lastname: Domanski
authors.affiliation: Laboratory of Mass Spectrometry and Gaseous Ion Chemistry, The Rockefeller University, New York, NY.
authors.forename: Brian T
authors.initials: BT
authors.lastname: Chait
authors.affiliation: Laboratory of Cellular and Structural Biology, The Rockefeller University, New York.
authors.forename: Michael P
authors.initials: MP
authors.lastname: Rout
mesh_terms: D002846:Chromatography, Affinity / Q000379:methods*; D006801:Humans; D013058:Mass Spectrometry; D046912:Multiprotein Complexes / Q000737:chemistry / Q000302:isolation & purification*; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification*; D040901:Proteomics
chemical_list: D046912:Multiprotein Complexes; D011506:Proteins
keywords: affinity; interactomics; protein complex; protein purification; proteomics