Profiling Native Kinases by Immuno‐Assisted Activity‐Based Profiling

Abstract The protocols in this unit describe efficient and cost‐effective approaches to determine the interaction of small‐molecule inhibitors with native kinases, and also analyze the interactions between kinases and their binding partners in a cellular setting. The combined attributes of activity‐based probes and western blotting procedures provide for quantitative measurement of inhibitor efficacy, isoform selectivity, and post‐translational modifications. We further demonstrate the ability to identify protein‐protein interactions between a probe‐labeled protein and its noncovalent binding partners. Curr. Protoc. Chem. Biol . 5:213‐226 © 2013 by John Wiley & Sons, Inc.