Chemical Protein Synthesis by Hydroxylamine-Involved Ligation of Peptides

Chemical protein synthesis by native amide ligation between two unprotected peptides provides an effective protocol. It is well-known that the development of a ligation method that is not limited by the sites of specific amino acids is highly desired. Here, we report an efficient chemoselective ligation between C-terminal peptide thioesters and N-terminal aminoacyl-N-hydroxy peptides in water (pH 8) at room temperature. The ligation reaction undergoes sequential S,O-ester exchange of peptide thioesters with N-hydroxyl of aminoacyl-N-hydroxy peptides providing O-acyl isopeptides and O,N-acyl transfer across a six-membered ring yielding ligated N-hydroxyl peptides. Reduction of N-hydroxyl peptides with zinc in an aqueous medium (pH 3) affords long peptides with native amide bonds. This hydroxylamine-involved ligation (HAL) of peptides was successfully applied to the synthesis of ubiquitin.