Cyclization provides a general strategy for improving the proteolytic stability, cell membrane permeability and target binding affinity of peptides. Disulphide bond formation between two free cysteine residues is an efficient and commonly used approach for cyclizing phage-displayed peptides.¹ However, these peptides have limited application in vivo as they cannot withstand reducing cellular environments. The disulphide bond within the peptide can be replaced with a stable, non-reducible linker. Among the vast collection of cysteine reactive linkers available, few provide the selectivity required to target the displayed peptide cysteine residues whilst sparing those on the phage capsid. This lack of selectivity negatively impacts phage infectivity and viability, which limits library diversity and disrupts the efficiency of selections.^{2, 3}

Here, we report the development of a novel chemical linker (linker CCA) that can efficiently cyclize synthetic peptides, peptides appended to a phage-coat protein and phage-displayed peptides in a site-specific manner. Unlike traditional cysteine reactive cyclization linkers, CCA does not affect the infectivity and / or viability of phage particles, nor does it require the use of disulphide-free pIII. We validate our cyclization approach through (A) the construction of a stable and highly diverse CCA-cyclized peptide phage display library and (B) the selection of novel binders to a range of biologically relevant targets.