STRUCTURE AND PHARMACOLOGY OF CHOLINERGIC PROTEINS

胆碱能蛋白的结构和药理学

• 批准号: 3478402
• 负责人: STEWART N ABRAMSON
• 金额: $ 4.25万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-04-01 至 1997-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3478402
• 关键词: Baculoviridae; Torpedo; acetylcholinesterase; active sites; affinity labeling; alternatives to animals in research; animal poison; clone cells; conformation; crosslink; fish electric organ; high performance liquid chromatography; neuropharmacology; neurotoxins; nicotinic receptors; protein structure function; radiotracer; site directed mutagenesis; transfection

Acetylcholinesterase is the enzyme responsible for the hydrolysis of acetylcholine, while nicotinic acetylcholine receptors contain cation-selective channels that open in response to binding acetylcholine. These two proteins play central roles in cholinergic neurotransmission, and they are therefore essential for normal neuronal function. Drugs that affect their activity are widely used as insecticides, as adjuvants in surgical anesthesia, and as therapeutic agents for the treatment of glaucoma, paralytic ileus, atony of the urinary bladder, and myasthenia gravis. All of the clinically useful drugs that affect the activity of these proteins do so by interacting with their acetylcholine-recognition sites. Therefore, structural and pharmacological characterization of these sites is essential for understanding the function of these proteins and for developing new and more useful drugs. The research proposed herein is directed towards the structural and functional characterization of the acetylcholine-recognition sites of acetylcholinesterase and the nicotinic acetylcholine receptor. In the first stage of the project an emphasis will be placed on using small, covalent, active-site directed affinity reagents to directly identify amino acids within the acetylcholine-binding sites. Examples include synthetic active-site directed crosslinking reagents, and the naturally occurring toxins onchidal and lophotoxin. (Onchidal and lophotoxin are active-site directed covalent inhibitors of acetylcholinesterase and nicotinic acetylcholine receptors, respectively). In the second stage of the project the amino acids identified with the active-site directed affinity reagents will be altered by site-directed mutagenesis and their function will be assessed by heterologous expression. Integration of experimental pharmacology, medicinal chemistry, protein chemistry, and molecular biology will increase our understanding of the molecular mechanisms by which these important proteins recognize and bind acetylcholine. As a result, such studies will ultimately aid in the design of new and more useful drugs and insecticides.

乙酰胆碱酯酶是一种负责水解酶 乙酰胆碱，而烟碱型乙酰胆碱受体含有 对结合的乙酰胆碱作出反应而开放的阳离子选择性通道。 这两种蛋白质在胆碱能神经传递中起着核心作用， 因此，它们对正常的神经元功能是必不可少的。毒品 影响其活性的物质被广泛用作杀虫剂、佐剂 在手术麻醉中，并作为治疗药物治疗 青光眼、麻痹性肠梗阻、膀胱无力和肌无力 格雷维斯。所有临床上有用的影响血管活性的药物 这些蛋白质通过与它们的乙酰胆碱识别相互作用来做到这一点。 网站。因此，其结构和药理特征 这些位点对于了解这些蛋白质的功能是必不可少的。 以及开发新的更有用的药物。 本文提出的研究是针对结构和 乙酰胆碱识别位点的功能表征 乙酰胆碱酯酶和烟碱型乙酰胆碱受体。在 该项目的第一阶段将重点放在使用小型、 共价、活性部位导向的亲和试剂直接鉴定 乙酰胆碱结合部位内的氨基酸。示例包括 合成的活性中心定向交联剂和天然的 发生的毒素有杀线虫和钩端螺旋藻毒素。(Onchidal和Lophotoxx是 活性部位定向的乙酰胆碱酯酶共价抑制剂和 烟碱型乙酰胆碱受体)。在第二阶段 该项目确定了与活性部位定向的氨基酸 亲和试剂将通过定点突变和它们的 功能将通过异源表达来评估。集成 实验药理学、药物化学、蛋白质化学和 分子生物学将增加我们对分子的理解。 这些重要蛋白质识别和结合的机制 乙酰胆碱。因此，这样的研究最终将有助于 设计新的更有用的药物和杀虫剂。