Acrolein-Mediated Conversion of Lysine to Electrophilic Heterocycles for Protein Diversification and Toxicity Profiling

Abstract

Understanding protein interactions in the presence of biological metabolites is critical for unraveling biological processes and advancing therapeutic interventions. This study focuses on α,β-unsaturated carbonyls, particularly acrolein-derived protein modifications, unveiling a one-pot, four-step, selective chemistry that results in the formation of a heterocyclic α,β-unsaturated carbonyl, termed 3-formyl-3,4-dehydropiperidino (FDP), exclusively on lysine residues. Remarkably, this chemistry transforms lysine, a nucleophile, into an electrophilic warhead. We demonstrate its versatility in late-stage peptide diversification, precision protein engineering, and homogeneous protein labeling with diverse payloads. Additionally, FDP-lysine smoothly transforms into another heterocycle, 3-methylpyridinium (3-MP) lysine via deoxygenation and aromatization in reagentless conditions. This transformation facilitates late-stage peptide functionalization and homogeneous engineering of proteins, with MP-lysine acting as a mass booster. Leveraging this chemistry, we discovered hyperreactive sites responsible for acrolein-induced modification through chemoproteomic profiling of FDP- and MP-modified proteins. Our findings revealed changes in protein–protein interactions mediated by FDP-modified proteins and uncovered ∼1548 novel cross-linking partners of an FDP-modified protein.