Native chemical ligation through in situ O to S acyl shift

[reaction: see text] A novel strategy to generate thioester peptides compatible with Fmoc chemistry is presented. Peptide-C(alpha)oxy-(2-mercapto-1-carboxyamide)ethyl ester undergoes an O to S acyl shift during ligation and the newly formed thioester intermediate reacts with an N-terminal cysteine fragment generating a product with native amide bond at the ligation site.