Triple-helical ligands selectively targeting the closed αI domain of integrin α2β1.

Integrins α1β1, α2β1, α10β1, and α11β1 are known to recognize the GxxGEx motif of the collagen triple helix through their αI domains, with the Glu residue engaging the domain via a divalent metal cation. The binding amino acid sequences containing these motifs, identified from native collagen, exhibit low subtype selectivity. Here we identify novel triple-helical peptides that selectively bind the α2I domain independently of Glu and metal ions. Yeast two-hybrid screening of randomized triple-helical peptide libraries yielded non-natural sequences in which the canonical Glu was replaced by aliphatic residues such as Met. X-ray crystal structural analysis of the representative variant GFOGMR in complex with the α2I domain revealed a new binding mode. In this mode, peptides are recognized by the closed, inactive conformation of the α2I domain without metal coordination. This cryptic interaction, likely unused by native ligands, provides a new basis for the design of subtype-specific ligands targeting collagen-binding integrins.