Directed evolution is a useful technology for creating novel biomolecules that enhance or mimic protein function. Small polypeptides are desired as therapeutics and research tools as they are amenable to chemical synthesis and offer facile incorporation into biomaterials. Using molecular cloning, biologically active amino acid sequences derived from cell adhesion proteins were grafted into several stable, constrained peptide frameworks and were shown to bind to integrin receptors with modest affinity. Since three dimensional conformation is critical for a polypeptide to bind integrins with high affinity and specificity, prototype molecules were subjected to affinity maturation through yeast surface display. Combinatorial libraries of mutants displayed on the yeast cell surface were screened by flow cytometric sorting to isolate polypeptides with enhanced integrin binding affinity. These results demonstrate that naturally-occurring constrained peptide scaffolds 1) can be redirected to function as adhesion molecule mimics and 2) can be engineered for enhanced integrin binding affinity through directed evolution. These peptides are currently being tested for their ability to specifically modulate integrin-mediated cell adhesion, and for their use as in vivo tumor targeting agents for molecular imaging and therapeutic applications. Supported by NIH 5K01CA104706 and 1R25CA118681 (to RHK).

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