Iterative reducible ligation to form homogeneous penicillamine cross-linked polypeptides

The syntheses of homogeneous penicillamine disulfide cross-linked polypeptides are reported. Dodecapeptides containing N-terminal, C-terminal, or N- and C-terminal Pen were serially ligated into 36 amino acid polypeptides linked through Cys-Pen, Pen-Cys, or Pen–Pen disulfide bonds. Critical to the syntheses was the incorporation of thiazolidine masked Cys and Pen as the N-terminal residues and selective hydrolysis with silver trifluoromethanesulfonate in acidic aqueous conditions to generate a free thiol for subsequent ligation. This approach allows the synthesis of homogeneous disulfide cross-linked polypeptides that have different reductive stabilities and have application in gene delivery by undergoing a tempered reductive triggered release of DNA.