Identifying Nuclear Protein–Protein Interactions Using GFP Affinity Purification and SILAC-Based Quantitative Mass Spectrometry

• DOI: 10.1007/978-1-4939-1142-4_15
• Publication Date: 2014-07-24T10:44:55Z
• Authors: H. Irem Baymaz; Cornelia G. Spruijt; Michiel Vermeulen
• Keywords: Cell Nucleus; Proteomics and Chromatin Biology; Isotope Labeling; Green Fluorescent Proteins; Protein Interaction Mapping; Proteolysis; Nuclear Proteins; Amino Acids; Peptides; Transfection; Chromatography, Affinity; Mass Spectrometry; Cell Line

Many cellular proteins assemble into macromolecular protein complexes. Therefore, identifying protein-protein interactions (PPIs) is essential to gain insight into the function of proteins. Recently established quantitative mass spectrometry-based techniques have significantly improved the unbiased search for PPIs. In this chapter, we describe a single-step GFP affinity purification method combined with SILAC-based quantitative mass spectrometry that can be used to identify nuclear PPIs in mammalian cells.