title: Contraceptive efficacy of recombinant fusion protein comprising zona pellucida glycoprotein-3 fragment and gonadotropin releasing hormone.
abstract: Contraceptive vaccines have been used for the management of wildlife population. In the present study, we have examined the contraceptive potential of Escherichia coli-expressed recombinant fusion protein comprising of 'promiscuous' T cell epitope of tetanus toxoid [TT; amino acid (aa) residues 830-844] followed by dilysine linker (KK), dog ZP3 fragment (aa residues 307-346), triglycine spacer (GGG), T cell epitope of bovine RNase (bRNase; aa residues 94-104), GnRH, T cell epitope of circumsporozoite protein of Plasmodium falciparum (CSP; aa residues 362-383), and GnRH. SDS-PAGE analysis of the purified refolded protein revealed a dominant ∼12 kDa band, which in Western blot reacted with mouse polyclonal antibodies against dog ZP3 fragment and mouse monoclonal antibodies against GnRH. Immunization of female FvB/J mice following two booster schedule with the above recombinant protein supplemented with alum led to high antibody titres against the immunogen as well as ZP3 and GnRH as determined by ELISA. The immune sera reacted with zona pellucida of mouse oocyte and also inhibited in-vitro fertilization. The qRT-PCR studies showed decrease in the ovarian GnRH receptor in mice immunized with the recombinant fusion protein. Mating studies revealed high contraceptive efficacy of the recombinant protein as in two independent experiments, 90% of the immunized female mice failed to conceive. Following one booster immunization schedule, 50% of the immunized female mice failed to conceive. However, in adjuvanted controls, all the female mice became pregnant. To conclude, the recombinant protein described herein has a good potential to be developed as candidate contraceptive vaccine.
authors: Reproductive Cell Biology Lab., National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110 067, India.
authors: Ananta Prasad
authors: AP
authors: Arukha
authors: Reproductive Cell Biology Lab., National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110 067, India.
authors: Vidisha
authors: V
authors: Minhas
authors: Reproductive Cell Biology Lab., National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110 067, India.
authors: Abhinav
authors: A
authors: Shrestha
authors: Reproductive Cell Biology Lab., National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110 067, India. Electronic address: skgupta@nii.ac.in.
authors: Satish Kumar
authors: SK
authors: Gupta
mesh_terms: D000818:Animals; D002417:Cattle; D005260:Female; D007987:Gonadotropin-Releasing Hormone / Q000235:genetics / Q000276:immunology* / Q000494:pharmacology; D051379:Mice; D011247:Pregnancy; D015800:Protozoan Proteins / Q000235:genetics / Q000276:immunology / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000276:immunology* / Q000494:pharmacology; D012260:Ribonucleases / Q000235:genetics / Q000276:immunology / Q000494:pharmacology; D022642:Vaccines, Contraceptive / Q000235:genetics / Q000276:immunology* / Q000494:pharmacology; D000072216:Zona Pellucida Glycoproteins / Q000235:genetics / Q000276:immunology* / Q000494:pharmacology
chemical_list: D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; D022642:Vaccines, Contraceptive; D000072216:Zona Pellucida Glycoproteins; C000606521:Zp3 protein, mouse; C041595:circumsporozoite protein, Protozoan; D007987:Gonadotropin-Releasing Hormone; D012260:Ribonucleases
keywords: Gonadotropin releasing hormone; Immunocontraception; Promiscuous T cell epitope; Tag-free recombinant fusion protein; Zona pellucida glycoprotein-3