De novo design of metal‐binding cleft in a Trp‐Trp stapled thermostable β‐hairpin peptide

Abstract

Metals are important molecules in protein biochemistry, as they are involved in various essential biochemical processes. Inspired by the coordination chemistry of metal-binding proteins, and to improvise the stability of recognition motifs, here we present the design of hyperstable stapled β-hairpin with a defined metal-binding cleft. We achieved this by establishing Trp-Trp covalent cross-linking in a long-chain 16-residue β-hairpin scaffold, and incorporating a stereospecific Cys₂-His₂ tetrad as the metal-binding cleft. This water-soluble peptide showed broad metal-binding properties, with Cu²⁺ specificity. Importance of the Cys-His tetrad in metal ion selectivity was established with Asp/Glu substitutions. We propose that such predefined hyperstable β-hairpins with recognition motifs are useful versatile tools for developing peptide-based catalysts, and in biomarker design.