In Vivo Cross-Linking to Analyze Transient Protein–Protein Interactions

Abstract

Cross-linking converts noncovalent interactions between proteins into covalent bonds. The now artificially fused molecules are stable during purification steps (e.g., immunoprecipitation). In combination with a variety of techniques, including Western blotting, mass spectrometry (MS), and bioinformatics, this technology provides improved opportunities for modelling structural details of functional complexes in living cells and protein–protein interaction networks. The presented strategy of immunoaffinity purification and mass spectrometry (AP-MS) coupled with in vivo cross-linking can easily be adapted as a robust workflow in interactome analyses of various species, also nonmodel organisms.