STRUCTURE AND PHARMACOLOGY OF CHOLINERGIC PROTEINS

乙酰胆碱酯酶是负责水解 乙酰胆碱，而烟碱乙酰胆碱受体含有 响应结合乙酰胆碱而打开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核心作用， 因此它们对于正常的神经元功能是必需的。 药物 影响其活性物质被广泛用作杀虫剂、佐剂 在外科麻醉中，以及作为治疗剂用于治疗 青光眼、麻痹性肠梗阻、膀胱弛缓和肌无力 gravis. 所有临床上有用的药物，影响活动的 这些蛋白质是通过与它们的乙酰胆碱识别 网站. 因此，结构和药理学表征的 这些位点对于理解这些蛋白质的功能至关重要 以及开发新的和更有用的药物。 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乙酰胆碱受体。 在 该项目的第一阶段将侧重于使用小型， 共价的、活性位点定向的亲和试剂， 乙酰胆碱结合位点内的氨基酸。 实例包括 合成的活性位点定向交联试剂，和天然的 出现毒素onchidal和lophotoxin。（Onchidal和lophotoxin是 乙酰胆碱酯酶的活性位点定向共价抑制剂， 烟碱乙酰胆碱受体）。 第二阶段 该项目的氨基酸确定与活性位点定向 亲和试剂将通过定点诱变改变， 通过异源表达来评估功能。 一体化 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 这些重要的蛋白质识别和结合的机制 乙酰胆碱 因此，这些研究最终将有助于 设计新的和更有用的药物和杀虫剂。

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.