In Cellulo Peptide Cyclization via Gene-Encoded Proximity-Induced Proline-Isothiocyanate Crosslinking

Cyclic peptides represent a very useful modality in modern drug discovery. However, there remain substantial technical challenges in the de novo biosynthesis of complex cyclic peptides particularly in an intracellular manner. Herein, we demonstrate that an isothiocyanate (ITC) group encoded via genetic code expansion could selectively and efficiently crosslink to proximal N-terminal Pro (P-ITC crosslinking) and affords cyclic peptides without obvious limitations related to amino acid composition (including Lys and Cys) or sequence length. Notably via P-ITC crosslinking, a disulfide-bridged 13-mer bicyclic peptide is facilely constructed in E. coli, thus lending us the ability to intracellularly construct and select complex cyclic peptides.