Identification of metal ion binding peptides containing unnatural amino acids by phage display

The bidentate metal binding amino acid bipyridylalanine (BpyAla) was incorporated into a disulfide linked cyclic peptide phage displayed library to identify metal ion binding peptides. Selection against Ni(2+)-nitrilotriacetic acid (NTA) enriched for sequences containing histidine and BpyAla. BpyAla predominated when selections were carried out at lower pH, consistent with the differential pKa's of histidine and BpyAla. Two peptides containing BpyAla were synthesized and found to bind Ni(2+) with low micromolar dissociation constants. Incorporation of BpyAla and other metal binding amino acids into peptide and protein libraries should enable the evolution of novel binding and catalytic activities.