Carbon Electrophiles with Markedly Distinct Amino Acid Reactivity Profiles in Proteomes

Abstract

The field of activity-based protein profiling applies reactive chem. probes to monitor enzyme activities in complex proteomes. Insight into the proteome reactivity of various classes of electrophiles is crucial for the design of covalent modifiers of enzymes lacking cognate affinity labels. Here, we investigate the proteome-wide amino acid labeling profiles for two carbon electrophiles, an alpha -chloroacetamide and sulfonate ester through the application of click chem. and advanced mass spectrometry methods. Our studies demonstrate that the a-chloroacetamide shows near exclusive reactivity with cysteine residues in proteomes, whereas the sulfonate ester displays a very diverse labeling profile, forming adducts with aspartate, glutamate, tyrosine, and cysteine residues. These sites of labeling are biased toward functional residues known to play a variety of catalytic and regulatory roles. Remarkably, the different labeling profiles in proteomes for the alpha -chloroacetamide and sulfonate ester probes were not reflected in their inherent reactivity with isolated amino acids. Insights into proteome reactivity thus discriminates electrophiles with restricted and promiscuous amino acid labeling profiles to provide key information for the future design of activity-based probes for a wide range of enzyme classes.