Automated detection of conformational epitopes using phage display Peptide sequences

Abstract

Background: Precise determination of conformational epitopes of neutralizing antibodies represents a key step in the rational design of novel vaccines. A powerful experimental method to gain insights on the physical chemical nature of conformational epitopes is the selection of linear peptides that bind with high affinities to a monoclonal antibody of interest by phage display technology. However, the structural characterization of conformational epitopes from these mimotopes is not straightforward, and in the past the interpretation of peptide sequences from phage display experiments focused on linear sequence analysis to find a consensus sequence or common sequence motifs.

Results: We present a fully automated search method, EpiSearch that predicts the possible location of conformational epitopes on the surface of an antigen. The algorithm uses peptide sequences from phage display experiments as input, and ranks all surface exposed patches according to the frequency distribution of similar residues in the peptides and in the patch. We have tested the performance of the EpiSearch algorithm for six experimental data sets of phage display experiments, the human epidermal growth factor receptor-2 (HER-2/neu), the antibody mAb Bo2C11 targeting the C(2) domain of FVIII, antibodies mAb 17b and mAb b12 of the HIV envelope protein gp120, mAb 13b5 targeting HIV-1 capsid protein and 80R of the SARS coronavirus spike protein. In all these examples the conformational epitopes as determined by the X-ray crystal structures of the antibody-antigen complexes, were found within the highest scoring patches of EpiSearch, covering in most cases more than 50% residues of experimental observed conformational epitopes. Input options of the program include mapping of a single peptide or a set of peptides on the antigen structure, and the results of the calculation can be visualized on our interactive web server.

Availability: Users can access the EpiSearch from our web server http://curie.utmb.edu/episearch.html.

Keywords: conformational epitopes; peptides; phage display.

Figure 1.

Flow chart of the EpiSearch method used to map peptide sequences obtained from phage display experiment onto 3D protein structure.

Figure 2.

A) Average score profile of five trastuzumab peptides CQMWAPQWGPDC, CKLYWADGELTC, CKLYWADGEFTC, CVDYHYEGTITC, and CVDYHYEGAITC in each patch centered on the surface exposed residues of HER2 protein surface is shown. B–C) shows the score profile for the five peptides at the two high scoring patches centered at residues 214 and 565, respectively.

Figure 3.

Comparison of the genuine epitope site on HER2 surface obtained from X-ray crystallography with the epitope site predicted by the EpiSearch method. A) The amino acids at the genuine epitope site on HER-2 surface (red). B) The amino acids predicted by the EpiSearch method present in the highest scoring patch centered on C565 and also present in the input peptide sequences (green), while the residues not present in the input peptide sequences are shown in orange. C) Amino acids correctly predicted and present in the genuine epitope binding site on HER2 surface (red), while the residues present in the epitope site and not predicted are shown in blue.

Figure 4.

A) Amino acids in C₂ domain of FVIII (red) complex with BO2C11 Fab. B) The residues predicted by the EpiSearch method in the highest scoring patch and present in the input peptide sequences are shown in green while the residues not present in the input peptide sequences are shown in orange. C) Amino acids correctly predicted and present in the BO2C11 binding site (red), while the residues present in the BO2C11 binding site and not predicted are shown in blue.