Chemoproteomic profiling and discovery of protein electrophiles in human cells.

Abstract

Activity-based protein profiling (ABPP) serves as a chemical proteomic platform to discover and characterize functional amino acids in proteins on the basis of their enhanced reactivity towards small-molecule probes. This approach, to date, has mainly targeted nucleophilic functional groups, such as the side chains of serine and cysteine, using electrophilic probes. We show here that "reverse-polarity" (RP)-ABPP using clickable, nucleophilic hydrazine probes can capture and identify protein-bound electrophiles in cells, including the pyruvoyl cofactor of S-adenosyl-l-methionine decarboxylase (AMD1), which we find is dynamically controlled by intracellular methionine concentrations, and a heretofore unknown modification – an N-terminally bound glyoxylyl group – in the poorly characterized protein secernin-3. RP-ABPP thus provides a versatile method to monitor the metabolic regulation of electrophilic cofactors and discover novel types of electrophilic modifications on proteins in human cells.