A Protein Cleavage Platform Based on Selective Formylation at Cysteine Residues

02 May 2024, Version 1
This content is a preprint and has not undergone peer review at the time of posting.


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 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 the cleavage-induced protein activation further demonstrates its utility as a platform for the functional regulation of proteins by artificial PTM.


protein cleavage
cysteine formylation
amide bond hydrolysis
affinity-based protein modification
post-translational modification

Supplementary materials

Supporting Information
Supplementary materials, figures, experimental details, general materials and methods for organic synthesis


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