Screening and identification of antigenic mimics of the adhesion protein of Mycoplasma genitalium by phage display peptide library

Abstract

Objective To screen a 12-mer phage display peptide library by the polyclonal antibody (pAb) against the recombinant adhesion protein of Mycoplasma genitalium (rMgPa) in order to obtain the antigenic mimic epitopes of MgPa.Methods The purified pAb was used to screen the immunodominant mimic epitopes of MgPa by a random 12-peptide phage display library.Seventy-four recombinant phage clones were randomly selected,and then DNA sequence analysis and computer-based bioinformatics analysis were performed to define the consensus amino acid residues of the mimotopes by MIMOX.The binding specificities of the selected phage-displayed peptides to the purified pAb were confirmed by ELISA,competitive ELISA and Western blot analysis.Results After four rounds of biopanning,a significant enrichment of phages was achieved,the inserts from 74 phage clones distinguished 45 peptides based on the different amino acids sequences.Amongst 45 peptides,36 peptides were ELISA positive and 23 peptides that absorbance values were higher than 1.5 showed high reactivities with pAb and effectively inhibited the binding of pAb to rMgPa.Immunoscreening via phage display peptide library revealed three different mimptopes of adhesion protein of M.genitalium,P-S-A-A/V-X-R-F/W-E/S-L-S-P,A-K-I/L-T/Q-X-T-L-X-L and K-S-L-S-R-X-D-X-I.Results of bioinformatics analysis by MIMOX demonstrated that S,A,F for cluster 1,A,K,I,T and L for cluster 2,K,S,L,R,D and I for cluster 3,may be the key consensus amino acid residues in the aligned mimotopes,respectively.Conclusion Antigenic mimics on MgPa were successfully identified and the motif P-S-A-A/V-X-R-F/W-E/S-L-S-P,A-K-I/L-T/Q-X-T-L-X-L and K-S-L-S-R-X-D-X-I may represent the immunodominant mimic epitopes of MgPa.And S,A,F K,I,T,L,R and D may be the key amino acid residues for the epitopes of MgPa. Key words: Mycoplasma genitalium; Adhesin protein; Phage display random peptide library; Mimic epitope