Methodology

Cognate peptide-receptor ligand mapping by directed phage display

Thomas Stratmann^1,2 and Angray S Kang¹

¹  Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA

²  Universidad de Barcelona, Departamento de Fisiologia, Diagonal 645, 3°, 08028 Barcelona, Spain

author email corresponding author email

Proteome Science 2005, 3:7doi:10.1186/1477-5956-3-7

Published:	17 June 2005

Abstract

Background

A rapid phage display method for the elucidation of cognate peptide specific ligand for receptors is described. The approach may be readily integrated into the interface of genomic and proteomic studies to identify biologically relevant ligands.

Methods

A gene fragment library from influenza coat protein haemagglutinin (HA) gene was constructed by treating HA cDNA with DNAse I to create 50 – 100 bp fragments. These fragments were cloned into plasmid pORFES IV and in-frame inserts were selected. These in-frame fragment inserts were subsequently cloned into a filamentous phage display vector JC-M13-88 for surface display as fusions to a synthetic copy of gene VIII. Two well characterized antibodies, mAb 12CA5 and pAb 07431, directed against distinct known regions of HA were used to pan the library.

Results

Two linear epitopes, HA peptide 112 – 126 and 162–173, recognized by mAb 12CA5 and pAb 07431, respectively, were identified as the cognate epitopes.

Conclusion

This approach is a useful alternative to conventional methods such as screening of overlapping synthetic peptide libraries or gene fragment expression libraries when searching for precise peptide protein interactions, and may be applied to functional proteomics.