Synthesis of peptides and proteins without cysteine residues by native chemical ligation combined with desulfurization

The highly chemoselective reaction between unprotected peptides bearing an N-terminal Cys residue and a C-terminal thioester enables the total and sem i-synthesis of complex polypeptides. Here we extend the utility of this nat ive chemical ligation approach to non-cysteine containing peptides. Since a lanine is a common amino acid in proteins, ligation at this residue would b e of great utility. To achieve this goal, a specific alanine residue in the parent protein is replaced with cysteine to facilitate synthesis by native chemical ligation. Following ligation, selective desulfurization of the re sulting unprotected polypeptide product with H-2/metal reagents converts th e cysteine residue to alanine. This approach, which provides a general meth od to prepare alanyl proteins from their cysteinyl forms, can be used to ch emically synthesize a variety of polypeptides, as demonstrated by the total chemical syntheses of the cyclic antibiotic microcin J25, the 56-amino aci d streptococcal protein G B1 domain, and a variant of the 110-amino acid ri bonuclease, barnase.