Ion Mobility-Mass Spectrometry to Evaluate the Effects of Protein Modification or Small Molecule Binding on Protein Dynamics

Abstract

Ion mobility-mass spectrometry (IM-MS) of intact protein complexes under native conditions is a powerful tool for the analysis of protein complexes and protein–ligand interactions, permitting insight into ligand-induced changes in protein conformation. Here we describe a procedure for analyzing the effects of phosphorylation and/or inhibitor binding on protein kinase conformational flexibility using Protein Kinase A (PKA) as a model system. By calculating the protein collision cross section (CCS) before and after inhibitor binding, and additionally by performing collision-induced unfolding (CIU), we can establish the effects of protein modification or small molecule binding on protein dynamics.