Harnessing Ionic Selectivity In Acetyltransferase Chemoproteomic Probes

Chemical proteomics (chemoproteomics) has emerged as a powerful strategy for the high-throughput annotation of protein function. This approach involves using an active site probe to capture an enzyme class of interest from cells, allowing the biological activity of multiple enzymes within that family to be assessed in parallel. Coenzyme A based chemoproteomic probes may have utility in studying the pharmacological interactions of additional enzyme classes, beyond histone acetyltransferases. However, one challenge in applying this approach is how to differentiate a ‘true hit’ from background. Many histone acetyltransferases utilize an ordered binding mechanism, with acetyl-CoA binding first, which allows their chemoproteomic capture to be competed by pre-incubating lysates with acetyl-CoA. However, for acetyltransferases that don’t exhibit this binding mode, or which bind to a different ligand, acetyl-CoA competition may not be evident, limiting our ability to discern selective chemoproteomic enrichment from noise. In this study, two physiochemically distinct capture probes were used to enrich proteins to distinguish true hits from background and identify new targets selectively enriched by CoA-based probes, thus expanding the scope of protein-ligand interactions open to chemoproteomic interrogation.