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 可能被利用以增强我们对机制的理解的探针 磷脂酶C的细菌和哺乳动物同工酶的作用 （PLC）类。 可以用作潜在机械的底物类似物 探针和抑制剂将是这些努力的合成目标。 众所周知，哺乳动物磷酸肌酸特异性PLC 通过释放第二使者扮演关键的作用信号转导 1,2-二酰基甘油（DAG）和1,4,5-肌醇三磷酸盐（IP3），这些 研究应能够阐明此的某些机械特征 重要过程。 在这些研究中制备的化合物可能具有 在包括抗癌在内的各种疾病地区的有益影响， 心血管和抗炎。 这些调查的主要重点将是：（1）发展 所有类别的合成的高效，一般方法 磷脂及其包含改良头基的衍生物 和/或改良的磷脂酸亚基； （2）设计和准备 合理选择的磷脂来检验有关的假设 磷酸二酯键的酶水解机制在不同 磷脂类别； （3）设计和合成磷脂底物 用于生物学筛查的类似物，作为细菌和细菌抑制剂的类似物 哺乳动物PLC同工酶； （4）单晶X射线合作 研究与细菌PLC复合的抑制剂检查磷脂 - 酶相互作用，从而获得了设计第二代的见解 机械探针。 生物测定和筛查实验调查结构活性 与细菌PLC（B. cereus）和Pi-Plc（苏云金氏芽孢杆菌）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射线晶体学研究将是 与爱德华·霍夫教授合作（大学 挪威Tromso）。 体外筛查结构活性 将与哺乳动物PI-PLC和PIP2-PLC的关系进行 与杜邦（Pat N. Confalone博士小组）的科学家合作， 与Philip J. Majerus教授（华盛顿大学医学院） 和苏·古鲁（Sue Goo Rhee）博士（国立卫生研究院，心脏，肺和血液 研究所）。