STRUCTURE AND PHARMACOLOGY OF CHOLINERGIC PROTEINS

胆碱能蛋白的结构和药理学

• 批准号: 2268030
• 负责人: STEWART N ABRAMSON
• 金额: $ 10.11万
• 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/2268030
• 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.

乙酰胆碱酯酶是负责水解的酶 乙酰胆碱，而烟碱乙酰胆碱受体含有 响应结合乙酰胆碱的阳离子选择通道。 这两种蛋白质在胆碱能神经传递中起着核心作用， 因此，它们对于正常的神经元功能至关重要。 毒品 影响其活性被广泛用作杀虫剂，作为佐剂 在手术麻醉中，作为治疗的治疗剂 青光眼，瘫痪的肠胃，膀胱和肌无力 重力。 影响影响活性的所有临床上有用的药物 这些蛋白质通过与乙酰胆碱识别相互作用来做到这一点 站点。 因此，结构和药理表征 这些位点对于理解这些蛋白质的功能至关重要 并用于开发新的和更有用的药物。 本文提出的研究针对结构和 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，主动位置的定向亲和力试剂直接识别 乙酰胆碱结合位点内的氨基酸。 示例包括 合成主动站点的定向交联试剂，自然 发生毒素的毒素和糖毒素。 （Onchidal和Lophotoxin是 乙酰胆碱酯酶的主动位置定向的共价抑制剂和 烟碱乙酰胆碱受体分别）。 在第二阶段 该项目的氨基酸用主动位置定向识别 亲和力试剂将通过定位的诱变及其 功能将通过异源表达评估。 整合 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。 结果，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。