title: Simple and Effective Affinity Purification Procedures for Mass Spectrometry-Based Identification of Protein-Protein Interactions in Cell Signaling Pathways.
abstract: Identification of protein-protein interactions can be a critical step in understanding the function and regulation of a particular protein and for exploring intracellular signaling cascades. Affinity purification coupled to mass spectrometry (APMS) is a powerful method for isolating and characterizing protein complexes. This approach involves the tagging and subsequent enrichment of a protein of interest along with any stably associated proteins that bind to it, followed by the identification of the interacting proteins using mass spectrometry. The protocol described here offers a quick and simple method for routine sample preparation for APMS analysis of suitably tagged human cell lines.
authors.affiliation: Department of Molecular Genetics, Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, 160 College Street, Toronto, ON, Canada, M5S 3E1.
authors.forename: Julian H M
authors.initials: JHM
authors.lastname: Kwan
authors.affiliation: Department of Molecular Genetics, Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, 160 College Street, Toronto, ON, Canada, M5S 3E1. andrew.emili@utoronto.ca.
authors.forename: Andrew
authors.initials: A
authors.lastname: Emili
mesh_terms: D000818:Animals; D002460:Cell Line; D002846:Chromatography, Affinity; D002853:Chromatography, Liquid; D006801:Humans; D013058:Mass Spectrometry; D025941:Protein Interaction Mapping / Q000379:methods*; D011506:Proteins / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D011506:Proteins
keywords: Affinity purification-mass spectrometry; Cell signaling pathway; Liquid chromatography-mass spectrometry