Screening Of BFGF-binding Peptide By Phage Display Technology

Objective: To obtain the bFGF-binding peptide via phage display technology, to predict the ligand binding sites on FGFR and the manner of bFGF-FGFR interaction, and to provide the base for developing peptide agonist or antagonist of bFGF.Methods: Using bFGF as the target, the phage display heptapeptide library was biopanned for 3 rounds to obtain the positive phage clones. The affinity and the specificity of the clones were assessed by ELISA. DNA sequencing and computation predicting were applied to analyze the positive clones to select the lead peptide mimicking FGFR2 consensus or homologous sequences. The competitive binding activity of the lead peptide phage were examined by ELISA. The biological activities of the lead peptide were also investigated by MTT and chick embryo chorioallantoic membrane angiogenesis (CAM) assay.Results: After 3 rounds screening, the rate of positive clone reached 30%. Twelve positive clones with high affinity and specificity were selected from the phages library. The amino acid sequences of two candidate peptides sharing a consensus sequence were homologous to that of the motif 255～261aa of FGFR2 D3 domain and also showed similar hydrophobic profile with that of the motif. The phages displaying this lead peptide specifically bind to bFGF and the binding was inhibited by free bFGF. The synthetic lead peptide could inhibit both bFGF-induced cell proliferation Balb/c 3T3 and the angiogenesis of chick embryo CAM.Conclusion: A bFGF-binding peptide was obtained by using the tool of phage display. It may mimic D3 domain of FGFR2 which specifically bind to bFGF. It can block the activity of bFGF. The study would offer the basis for further locating the bFGF binding sites on FGFR2, designing effective antiangiogenic peptides and developing tumor inhibitors.