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Analysis of an epitope sequence recognized by a monoclonal antibody MAb-5H4 against a porcine zona pellucida glycoprotein (pZP4) that blocks fertilization

The zona pellucida glycoprotein that surrounds the mammalian oocyte has several target antigens that have potential use in the development of a contraceptive vaccine. In the present study, an epitope sequence recognized by a monoclonal antibody to the porcine zona pellucida glycoprotein ZP4 was determined. Three candidate peptides were synthesized, based on an epitope mapping by cDNA and an analysis of chain flexibility of porcine ZP4. Only one synthetic peptide, corresponding to amino acid positions 50–67, reacted with the monoclonal antibody; the other synthetic peptides, corresponding to positions 60–79 and 70–100, did not react. The reactive epitope was identified as CTYVLDPENL, corresponding to positions 50–59 of porcine ZP4. The peptide inhibited the reaction of the monoclonal antibody binding to native ZP4 in a dose-dependent manner. When the synthetic peptide 50–67 was used to immunize mice, the resultant antisera reacted not only with the synthetic peptide but also with native pig zona pellucida. In addition, anti-peptide 50–67 antibody inhibited porcine fertilization in vitro. It is thus concluded that the peptide identified as an epitope for the monoclonal antibody would be a promising candidate for the development of a contraceptive vaccine.