A Protein Cleavage Platform Based on Selective Formylation at Cysteine Residues

Abstract

Site-selective cleavage of the peptide backbone in proteins is an important class of post-translational modification (PTM) in nature. However, the organic chemistry for such site-selective peptide bond cleavages has yet to be fully explored. Herein, we report cysteine S-formylation as a means of selective protein backbone cleavage. We developed N-formyl sulfonylanilide as a cysteine-selective formylation reagent for peptides and proteins. Upon S-formylation with the reagent, the amide bond adjacent to the S-formylated cysteine is cleaved by hydrolysis under neutral aqueous conditions. Formylation probes bearing a protein ligand enabled the affinity-based selective cleavage of the target proteins not only in the test tube but also under biorelevant conditions such as in crude cell lysate and on the cell surface. These results demonstrate the high biocompatibility of this protein cleavage technology. A proof-of-concept study of cleavage-induced protein activation further demonstrates its utility as a platform for the functional regulation of proteins by artificial PTM.