Enzyme-enabled proteomics for decoding the functional proteome

Mass spectrometry (MS)-based proteomics has transformed our ability to measure protein abundance and modification state on a proteome-wide scale. However, many critical determinants of protein function, including post-translational modification (PTM) state, spatial localization, and enzyme activity, require strategies designed to specifically capture these biological features. Enzymes are uniquely suited for this challenge because their molecular recognition properties, catalytic activities, and genetic encodability enable highly selective transformations that are difficult to achieve using purely chemical approaches. In this review, we highlight recent advances in enzyme-enabled proteomics that exploit natural and engineered enzyme activity to expand the functional information accessible by MS-based proteomics analysis. We discuss approaches that harness endogenous enzymes for metabolic labeling, repurpose enzyme chemistry for selective PTM tagging, and engineer enzymes to record protein localization and synthesis. Finally, we consider how continued discovery and engineering of enzyme chemistry may shift the role of enzymes beyond enrichment toward molecular recording strategies that encode increasingly complex aspects of protein function.