HORMONE-BINDING DOMAINS OF LH/HCG RECEPTOR

促性腺激素受体(LHR)属于严重跨膜超家族 结构域G偶联受体。其长的胞外氨基末端 341-368个氨基酸残基的多肽具有较高的亲和力 激素结合序列。因为已经有可能准备展会 大肠杆菌胞外氨基末端多肽的量和 CHO细胞，现在已经可以进行其详细的 物理化学、免疫学和生物学特性。这个 这些研究的最终目标是尽可能准确地定义 受体的配基结合部位，目的是开发 人绒毛膜促性腺激素(HCG)拮抗剂。胞外结构域 的大鼠(LHR)(1-294和1-341残留物)将以毫克为单位制备 从大肠杆菌、CHO和SF-9昆虫细胞中提取的数量。在首字母之后 大鼠LHR重组氨基末端多肽的鉴定 氨基酸组成、肽图谱及氨基和羧基末端 测序，(1)配基结合部位测定的研究 将会被承担。几种方法，如使用删除和 利用蛋白水解物进行定点突变截断突变体 胞外区多肽和合成肽的片段 将对LHR的高亲和力激素结合位点进行定位。(2) LHR将通过研究一些 翻译后修饰，包括二硫键的位置 键和碳水化合物单位的详细结构 昆虫细胞CHO和SF-9的胞外区。(3)三个人 将对空间结构进行分析和尝试，实现晶化 氨基末端1-294和1-341残基多肽及其 只含硒的类似物。加入硒可以避免这个问题。 X射线衍射谱中重金属离子的类质同晶取代 学习。(4)抑制肿瘤生长的多克隆抗体和单抗 HCG与受体的结合。(5)体外和体内实验 大鼠LHR胞外区的生物学特性 人绒毛膜促性腺激素对大鼠促性腺激素升高的抑制作用 将进行子宫重量、排卵和孕酮合成。 这些研究将有助于评估 作为hCG拮抗剂的LHR胞外多肽对生育的影响 监管。对大鼠LHR的初步研究将扩展到人类LHR。

Lutropin receptor (LHR) belongs to the superfamily of sever transmembrane domain G-coupled receptors. Its long extracellular aminoterminal polypeptide of 341-368 amino acid residues contains the high affinity hormone binding sequences. Since it has been possible to prepare fair amounts of the extracellular aminoterminal polypeptide from E. coli and CHO cells, it has now become feasible to undertake its detailed physicochemical, immunological and biological characterization. The ultimate goal of these studies is to define as precisely as possible the ligand binding site of the receptor with the objective of developing antagonists of human choriogonadotropin (hCG). The extracellular domain of rat (LHR) (1-294 and 1-341 residues) will be prepared in milligram quantities from E. coli, CHO and SF-9 insect cells. After initial characterization of the recombinant aminoterminal peptides of rat LHR by amino acid composition, peptide mapping and amino and carboxyterminal sequencing, (1) studies on the determination of the ligand binding site will be undertaken. Several approaches such as the use of deletion and truncation mutants by site directed mutagenesis, use of proteolytic fragments of the extracellular domain polypeptides and synthetic peptides will be made to map the high affinity hormone binding site of LHR. (2) The LHR will be further characterized by studying some of the posttranslational modifications including the positions of the disulfide bonds and the detailed structures of the carbohydrate unites in the extracellular domains from CHO and SF-9 insect cells. (3) For three dimensional structural analysis and attempt will be made to crystallized the aminoterminal 1-294 and 1-341 residue polypeptides as well as its selenomethionly analogs. Insertion of selenium will obviate the problem of isomorphous replacement by heavy metal ion in x-ray diffraction studies. (4) Polyclonal and monoclonal antibodies which inhibit the binding of hCG to the receptor. (5) Finally, the in vitro and in vivo biological characterization of the extracellular domain of rat LHR by studying the inhibition of hCG action in hCG induced increase in rat uterine weight, ovulation and progesterone synthesis will be carried out. These studies will help in the evaluation of the potential of the extracellular polypeptides of LHR as antagonists of hCG for fertility regulation. Initial studies on rat LHR will be extended to human LHR.