A novel strategy for a more efficient synthesis of difficult sequence-containing peptides, the S-acyl isopeptide method, was developed and successfully applied. A model pentapeptide Ac–Val–Val–Cys–Val–Val–NH2 was synthesized via its water-soluble S-acyl isopeptide using an S-acyl isodipeptide unit, Boc–Cys(Fmoc–Val)–OH. An S-acyl isopeptide possessing excellent water solubility could be readily and quantitatively converted to the native peptide via an SN intramolecular acyl migration reaction at pH 7.4. Thus, the S-acyl isopeptide method provides a useful tool in peptide chemistry. Copyright © 2008 European Peptide Society and John Wiley & Sons, Ltd.