DESIGN OF CHEMICAL PROBES TO STUDY PHOSPHOLIPASE C

研究磷脂酶 C 的化学探针的设计

• 批准号: 3301616
• 负责人: STEPHEN MARTIN
• 金额: $ 11.48万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-07-01 至 1994-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3301616
• 关键词: X ray crystallography; chemical synthesis; enzyme mechanism; enzyme substrate analog; isozymes; phosphatidylinositols; phospholipase C; phospholipid inhibitor; phospholipids; protein structure function; radiotracer

The overall goal of this research program is to design and develop chemical probes that may be exploited to enhance our understanding of the mechanism of action of bacterial and mammalian isoenzymes of the phospholipase C (PLC) class. Substrate analogues that may serve as potential mechanistic probes and inhibitors will be the synthetic targets of these efforts. Inasmuch as it is known that the mammalian phosphoinositide-specific PLC plays a key role signal transduction by releasing the second messengers 1,2-diacylglycerol (DAG) and 1,4,5-inositol trisphosphate (IP3), these studies should enable elucidation of certain mechanistic features of this important process. Compounds prepared during these studies could have beneficial impact in a variety of disease areas, including anticancer, cardiovascular, and anti-inflammatory. The principal foci of these investigations will be to: (1) develop efficient, general methods for the syntheses of all classes of phospholipids and derivatives thereof that contain modified head groups and/or modified phosphatidic acid subunits; (2) design and prepare rationally selected phospholipids to test hypotheses regarding the mechanism of enzymatic hydrolysis of the phosphodiester bond in different classes of phospholipids; (3) design and synthesize phospholipid substrate analogues for biological screening as potential inhibitors of bacterial and mammalian PLC isoenzymes; and (4) collaborate in single crystal X-ray studies of inhibitors complexed with bacterial PLC to examine phospholipid- enzyme interactions thereby gaining insights to design second generation mechanistic probes. Biological assays and screening experiments to survey structure-activity relationships with bacterial PLC (B. cereus) and PI-PLC (B. thuringiensis) will be executed in our laboratories according to standard protocols. The X-ray crystallographic studies of inhibitors of bacterial PLC will be carried out in collaboration with Professor Edward Hough (University of Tromso, Norway). The in vitro screening for structure-activity relationships with mammalian PI-PLC and PIP2-PLC will be performed in collaboration with scientists at DuPont (Dr. Pat N. Confalone's group), with Professor Philip J. Majerus (Washington University School of Medicine) and Dr. Sue Goo Rhee (National Institutes of Health, Heart, Lung and Blood Institute).

这项研究计划的总体目标是设计和开发化学品 可能被利用来增强我们对机制的理解的探测 细菌和哺乳动物磷脂酶C同工酶的作用 (PLC)类。可作为潜在机制的底物类似物 探针和抑制剂将是这些努力的合成目标。 由于已知哺乳动物磷脂酰肌醇特异的PLC 通过释放第二信使发挥关键的信号转导作用 1，2-二酰基甘油(DAG)和1，4，5-三磷酸肌醇(IP3)，这些 研究应该能够阐明这一现象的某些机制特征 重要的过程。在这些研究中制备的化合物可能具有 对各种疾病领域的有益影响，包括抗癌， 心血管和抗炎作用。 这些调查的主要焦点将是：(1)开发 有效的、通用的方法来合成所有类别的 含有修饰头基的磷脂及其衍生物 和/或修饰的磷脂酸亚基；(2)设计和制备 合理选择磷脂以检验关于 不同底物中磷酸二酯键的酶促降解机理 磷脂的分类；(3)磷脂底物的设计与合成 作为细菌和细菌的潜在抑制物的生物筛选类似物 哺乳动物的PLC同工酶；以及(4)单晶X射线的协作 细菌PLC与缓蚀剂复配检测磷脂的研究 酶的相互作用从而获得设计第二代的洞察力 机械探头。 用于结构活性研究的生物测定和筛选实验 细菌PLC(蜡样芽孢杆菌)和PI-PLC(苏云金芽孢杆菌)的关系 将根据标准协议在我们的实验室执行。这个 细菌PLC抑制剂的X射线结晶学研究将是 与Edward Hough教授(华盛顿大学)合作进行 挪威特罗姆索)。构效关系的体外筛选 与哺乳动物PI-PLC和PIP2-PLC的关系将在 与杜邦公司(Pat N.Confone博士的团队)的科学家合作， 与菲利普·J·马杰鲁斯教授(华盛顿大学医学院) 和Sue Goo Rhee博士(国立卫生、心脏、肺和血液研究所 研究所)。