ESTROGEN SYNTHETASE: CHARACTERIZATION AND CONTROL

我们建议从人胎盘中纯化细胞色素P-450 催化雄激素芳构化的酶 雌激素。该酶将根据以下方面进行表征 亚态专一性和动力学、物理化学性质和 部分氨基酸序列。我们还提议为这两项工作做准备 纯化蛋白的多克隆抗体和单抗。 此外，我们建议研究几个 作为结构探针的活性位点定向抑制剂的类型 和酶的功能。两种可逆的抑制剂-- 模拟雄激素底物的非甾体化合物和 一种新型的双功能类固醇抑制剂将用于 映射活动场地几何图形。几种类型的化学成分 活性类固醇--直接烷基化、自杀底物和 光亲和试剂-将被用来共价标记 蛋白质，以便识别蛋白质序列中 包括活动站点。这些抑制剂将具有启发式的用途。 用于研究芳构化的分子事件的细节 反应，并将在药物设计中有应用。 雌激素依赖型癌症的治疗。

We proposed to purify from human placenta the cytochrome P-450 enzyme which catalyzes the aromatization of androgens to estrogens. The enzyme will be characterized with respect to substate specificity and kinetics, physicochemical properties and partial amino acid sequence. We also proposed to prepare both polyclonal and monoclonal antibodies to the purified protein. In addition, we propose to investigate the interaction of several types of active site-directed inhibitors as probes of the structure and function of the enzyme. Two types of reversible inhibitors -- non-steroidal compounds which mimic the androgen substrates and a new type of bifunctional steroid inhibitor - will be used for mapping the active site geometry. Several types of chemically reactive steroids -- direct alkylators, suicide substrates and photo-affinity reagents - will be used to covalently label the protein in order to identify regions of the protein sequence which comprise the active site. The inhibitors will be of heuristic use for studying details of the molecular events of the aromatization reaction and will have an application in the design of drugs for the treatment of estrogen-dependent cancers.