Combination of Thiol-Additive-Free Native Chemical Ligation/Desulfurization and Intentional Replacement of Alanine with Cysteine.

We report a novel strategy for native chemical ligation (NCL). Alanines not located at a ligation site are temporarily replaced with cysteines, and this enables efficient thiol-additive-free NCL, with subsequent desulfurization to regenerate the target peptide. We synthesized stresscopin-related peptide and neuroendocrine regulatory peptide-2 (NERP-2) by this method. We confirmed that both conventional alkyl thioester and thioester-equivalent N-acyl-N'-methyl-benzimidazolinone (MeNbz) can be adopted as thioester components for thiol-additive-free NCL of multi-Cys-containing peptides.