Detection of structure-specific interactions of Rab GTPases

Protein–protein interactions (PPIs) mediate multiple essential functions and regulatory events in living organisms. The physical interactome of a given protein can be abnormally altered in response to external and internal cues, thus modulating cell physiology and contributing to human pathologies including neurodegenerative diseases. Despite substantial efforts, current approaches cannot pinpoint structure-specific PPIs or their interaction interfaces on a proteome-wide scale. To address this limitation, we have adapted the structural proteomics method known as limited proteolysis–mass spectrometry to identify PPIs by probing protein structural alterations within cellular extracts upon treatment with a protein of interest. Here, we added Rab GTPases, namely Rab5A and Rab2A in their GTP- and GDP-bound forms to a HEK293T cellular extract and probed their structure-specific interactomes. We identified already know interactors of these proteins bassed on previously published data as well as a number of candidate interactor proteins.