Mapping of Minimal Motifs of B-Cell Epitopes on Human Zona Pellucida Glycoprotein-3

Xu, Wan-Xiang; He, Ya-Ping; Wang, Jian; Tang, Hai-Ping; Shi, Hui-Juan; Sun, Xiao-Xi; Ji, Chao-Neng; Gu, Shao-Hua; Xie, Yi

The human zona pellucida glycoprotein-3 (hZP3) by virtue of its critical role during fertilization has been proposed as a promising candidate antigen to develop a contraceptive vaccine. In this direction, it is imperative to map minimal motifs of the B cell epitopes (BCEs) so as to avoid ZP-specific oophoritogenic T cell epitopes (TCEs) in the ZP3-based immunogens. In this study, based on known results of mapping marmoset and bonnet monkey ZP3 (mstZP3 and bmZP3), two predictable epitopes(23–30 and 301–320) on hZP3 were first confirmed and five minimal motifs within four epitopes on hZP3 were defined using serum to recombinant hZP3a(22–176) or hZP3b(177–348) as well as a biosynthetic peptide strategy. These defined minimal motifs were QPLWLL(23–28) for hZP3(23–30), MQVTDD(103–108) for hZP3(93–110), EENW(178–181) for hZP3(172–190), as well as SNSWF(306–310) and EGP(313–315) for hZP3(301–320), respectively. Furthermore, the antigenicity of two peptides for hZP3(172–187) and hZP3(301–315) and specificity of the antibody response to these peptides were also evaluated, which produced high-titer antibodies in immunized animals that were capable of reacting to ZP on human oocytes, r-hZP3b(177–348) protein, as well as r-hZP3(172–190), r-hZP3(303–310), and r-hZP3(313–320) epitope peptides fused with truncated GST188 protein.