Trinucleotide cassettes increase diversity of T7 phage-displayed peptide library

In this study, we evaluated and compared the diversity of a 12-mer T7 phage-displayed peptide library randomized using codon-corrected trinucleotide cassettes with a T7 and an M13 12-mer phage-displayed peptide library constructed using the degenerate codon randomization method.We herein demonstrate that the combination of trinucleotide cassette amino acid codon randomization and T7 phage display construction methods resulted in a significant enhancement to the functional diversity of a 12-mer peptide library. This novel library exhibited superior amino acid uniformity and order-of-magnitude increases in amino acid sequence diversity as compared to degenerate codon randomized peptide libraries. Comparative analyses of the biophysical characteristics of the 12-mer peptide libraries revealed the trinucleotide cassette-randomized library to be a unique resource.The combination of T7 phage display and trinucleotide cassette randomization resulted in a novel resource for the potential isolation of binding peptides for new and previously studied molecular targets.Bacteriophage (phage)-displayed random peptide libraries have become a widely-used screening resource for identifying ligands for molecular targets. Several applications of phage-displayed peptide library technology include epitope and protein-protein interaction mapping, and ligand and enzyme substrate specificity identification [1-5]. Successful application of the technology is dependent on screening methodology, on the biophysical characteristics of the target molecule and peptide library, and on the amino acid sequence diversity of the library. The library should be of adequate sequence diversity to contain binding ligands for the target, however, diversity can be limited both by biological censorship of phage-displayed amino acid sequences and by methods of amino acid sequence randomization.Biological censorship of libraries displayed on phage is a consequence of conducting combinatorial chemistry within a b