Antibodies can cause interference in gonadotropinic activity disrupting reproductive processes and thereby forming the basis for the design of contraceptive vaccines. The isolation of new immunocontraceptives requires a precise understanding of the structural interactions between hormone and receptor. A definitive insight of the gonadotropins and their receptors will facilitate the design of molecules that will specifically inhibit or enhance hormone action. The objective of this proposal is to examine the structure-function relationship of gonadotropins. This will be accomplished collaboratively using two complementary approaches. First, is to determine the structure of novel analogs of the hormones exhibiting either agonistic or antagonistic properties. Earlier studies have shown that single chain of human chorionic gonadotropin (hCG) beta beta binds to the receptor without eliciting biological activity, as well as competitively inhibiting native hormone both in vitro and in vivo. The hyper-expression and purification of this molecule is currently underway in the Indian laboratory. The solution structures will be simplified by using the previously solved structures of heterodimeric hCG and follicle stimulating hormone (FSH). Structure solution analyses will use X-ray crystallography, as well as NMR spectroscopy. The second approach utilizes a panel of monoclonal antibodies already available plus new antibodies produced and characterized during this study to investigate the hormone-receptor interface. These studies will produce single chain fragment variables (ScFv) from hybridoma clones secreting monoclonal antibodies. We will explore the structure of ScFvs individually and in complex with the hormones and mutations of ScFvs, which may render higher affinity for the hormones. This proposal focuses on developing new biological tools to interfere in the actions of gonadotropins while also probing critical aspects of hormone structure. The laboratory of the Indian PI will produce and purify the hormones and antibodies while the laboratory of the U.S. PI will determine the structure of the proteins. The strength of the Indian laboratory is generating large quantities of purified and characterized protein while the strength of the U.S. laboratory is the ability to solve the structure of the proteins. Thus both laboratories uniquely complement each others activities. The Indian laboratory has already initiated efforts towards production of monoclonal antibodies (MAbs) and ScFvs. These proteins will be supplied to U.S. investigator who will attempt to solve the structure of the proteins produced. ? ? ?
