MAPPING OF EPITOPES ON PORCINE ZONA PELLUCIDA-3-ALPHA BY MONOCLONAL-ANTIBODIES INHIBITING OOCYTE-SPERM INTERACTION

Zone pellucida glycoprotein 3 alpha (ZP3 alpha) has been designated as the primary sperm receptor ligand in porcine gamete interaction. In t his study, epitopes were mapped on porcine ZP3 alpha (pZP3 alpha) by u sing monoclonal antibodies (mAbs) possessing in vitro contraceptive ef ficacy. Using Western blots, we tested recombinant pZP3 alpha fragment s expressed as fusion proteins in Escherichia coli and corresponding t o pZP3 alpha precursor protein amino acid residues 18-142 (F1), 140-24 3 (F2), 239-363 (F3), and 359-462 (F4), for reactivities with mAbs. MA b-403 reacted with F3, and mAbs-412 and -421 with F2. MAb-420 showed w eak reactivity with F1. Synthesis of overlapping 12-mer peptides furth er resolved the epitope for mAb-420 to amino acid residues 133-144, mA b-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 a lpha, should assist in the design of a synthetic peptide-based immunoc ontraceptive vaccine.