Gonadotropin Inhibitors:A Structural Biology Approach To Immunocontraception

说明(申请人提供)：抗体可以干扰促性腺激素的活动，扰乱生殖过程，从而形成避孕疫苗设计的基础。新型免疫避孕药的分离需要对激素和受体之间的结构相互作用有精确的了解。对促性腺激素及其受体的明确认识将有助于设计出专门抑制或增强激素作用的分子。这项建议的目的是研究促性腺激素的结构与功能的关系。这将通过两种相辅相成的方法协作完成。首先，确定具有激动性或拮抗性的激素的新类似物的结构。早期的研究表明，人绒毛膜促性腺激素(HCG)β的单链与受体结合，不会引发生物活性，也不会在体外和体内竞争性地抑制天然激素。该分子的超表达和纯化目前正在印度实验室进行。利用已有的异二聚体hCG和卵泡刺激素(FSH)的结构将简化溶液结构。结构溶液分析将使用X射线结晶学以及核磁共振光谱。第二种方法利用一组已有的单抗，加上在这项研究中产生和表征的新抗体来研究激素-受体的界面。这些研究将从分泌单抗的杂交瘤克隆中产生单链片段变量(ScFv)。我们将单独探索单链抗体的结构，以及与激素和单链抗体突变的复合体，这可能会使单链抗体对激素有更高的亲和力。这项建议的重点是开发新的生物工具来干预促性腺激素的活动，同时也探索激素结构的关键方面。印度PI的实验室将生产和纯化激素和抗体，而美国PI的实验室将确定蛋白质的结构。印度实验室的优势是产生大量纯化的和具有特征的蛋白质，而美国实验室的优势是解决蛋白质结构的能力。因此，两个实验室的活动具有独特的互补性。印度实验室已经开始努力生产单抗和单链抗体。这些蛋白质将被提供给美国的研究人员，他们将试图解决产生的蛋白质的结构。

DESCRIPTION (provided by applicant): 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.