Genetically Encoding Propiolamide Warhead for the Construction of Phage Displayed Cyclic Peptide Library

Phage display is foundational to cyclic peptide‐based ligand discovery, and recently genetically encoded electrophilic noncanonical amino acids (ncAAs) have proven useful in constructing phage‐displayed cyclic peptide libraries via proximity‐induced crosslinking. However, electrophilic ncAAs exploited for this purpose remain quite rare, thus impeding the broad application of this emerging technology. Herein, we design and genetically encode propiolamide‐bearing ncAAs into proteins and demonstrate the capability of propiolamide warhead in conjugating to proximal Cys via a stable and rigid vinyl sulfide connectivity. Importantly, the encoded propiolamide warhead could be facilely displayed on phage surface and support large recombinant library construction. Via biopanning, cyclic peptide‐based inhibitor targeting SARS‐CoV‐2 main protease was discovered.