Mapping of epitopes on porcine zona pellucida-3 α by monoclonal antibodies inhibiting oocyte-sperm interaction

Abstract

Zona pellucida glycoprotein 3 α(ZP3 α) has been designated as the primary sperm receptor ligand in porcine gamete interaction. In this study, epitopes were mapped on porcine ZP3 α (pZP3 α) by using monoclonal antibodies (mAbs) possessing in vitro contraceptive efficacy. Using Western blots, we tested recombinant pZP3 α fragments expressed as fusion proteins in Escherichia coli and corresponding to pZP3 α precursor protein amino acid residues 18-142 (F1), 140-243 (F2), 239-363 (F3), and 359-462 (F4), for reactivities with mAbs. MAb-403 reacted with F3, and mAbs-412 and -421 with F2. MAb-420 showed weak reactivity with F1. Synthesis of overlapping 12-mer peptides further resolved the epitope for mAb-420 to amino acid residues 133-144, mAb-421 to 157-168, mAb-412 to 205-216, and mAb-403 to 301-312. MAbs-412 and -420 inhibited the binding of boar sperm to zona-encased porcine oocytes. These results, the first to define peptide epitopes of pZP3 α, should assist in the design of a synthetic peptide-based immunocontraceptive vaccine.