Semisynthetic head-to-tail cyclized peptides obtained by combining protein trans-splicing and intramolecular expressed protein ligation.

A dual-intein approach for the preparation of head-to-tail macrocyclic peptides is reported, where synthetic and genetically encoded fragments are ligated by two native peptide bonds. A split intein ligates the synthetic and genetically encoded peptides via protein trans-splicing and is followed by intramolecular cyclization through an expressed protein ligation step mediated with a cis-intein. We identified a suitable pair of orthogonal inteins and optimized the conditions for a one-pot cyclization protocol. We report the semisynthesis of model macrocyles with various ring sizes and of the natural product sunflower trypsin inhibitor (SFTI) along with its ornithine analog.