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).

该研究计划的总体目标是设计和开发化学 可用于增强我们对该机制的理解的探针 磷脂酶 C 的细菌和哺乳动物同工酶的作用 （PLC）类。 可能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。 因为众所周知，哺乳动物磷酸肌醇特异性 PLC 通过释放第二信使在信号转导中发挥关键作用 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种现象的某些机制特征 重要过程。 在这些研究期间制备的化合物可能具有 对多种疾病领域产生有益影响，包括抗癌、 心血管、抗炎。 这些调查的主要重点是：（1） 合成所有类别的有效、通用方法 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)设计与准备 合理选择磷脂来检验有关的假设 不同酶解磷酸二酯键的机理 磷脂类别； (3)磷脂底物的设计与合成 作为细菌和潜在抑制剂的生物筛选类似物 哺乳动物PLC同工酶； (4) 在单晶 X 射线方面合作 研究抑制剂与细菌 PLC 复合以检查磷脂 酶相互作用从而获得设计第二代的见解 机械探针。 用于调查结构活性的生物测定和筛选实验 与细菌 PLC（蜡状芽孢杆菌）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中执行。 这 细菌 PLC 抑制剂的 X 射线晶体学研究将 与 Edward Hough 教授（英国大学）合作进行 挪威特罗姆瑟）。 结构活性的体外筛选 与哺乳动物 PI-PLC 和 PIP2-PLC 的关系将在 与杜邦公司的科学家（Pat N. Confalone 博士的小组）合作， 与 Philip J. Majerus 教授（华盛顿大学医学院） 和 Sue Goo Rhee 博士（美国国立卫生研究院、心脏、肺和血液研究所） 研究所）。