Studies of P2X ATP Receptors

P2X ATP 受体的研究

• 批准号: 7034026
• 负责人: RICHARD IRWIN HUME
• 金额: $ 33.64万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-03-01 至 2009-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7034026
• 关键词: Xenopus; Xenopus oocyte; adenosine triphosphate; binding sites; biological signal transduction; calcium flux; cell line; glutamate receptor; ligands; molecular genetics; molecular site; neurophysiology; protein structure function; purinergic receptor; receptor binding; receptor expression; receptor sensitivity; site directed mutagenesis; voltage /patch clamp

DESCRIPTION (provided by applicant): The P2X proteins are ATP gated channels that depolarize cells and also allow calcium to enter. P2X receptors are expressed in virtually every tissue, including neurons and glia of the central and peripheral nervous system, smooth, skeletal and cardiac muscle, cochlear hair cells, platelets, most classes of white blood cells, hepatocytes, and endothelial cells in the lung and gastrointestinal tract. The importance of members of this gene family for normal physiology is apparent from the range of phenotypes that are seen in their absence, Mice in which specific P2X receptors are knocked out show dysfunction in pain perception, ability to void the bladder, gut motility, neuronal control of ejaculation, the ability of the nervous system to monitor the oxygen level in the blood, the ability to fight bacterial infection, and blood clotting. A major problem in the purinergic receptor field is the limited specificity of agonists and antagonists that can be used to alter ATP signaling in vivo. The goal of the experiments described here is to better characterize the molecular mechanisms that allow ATP and allosteric modulators to open P2X receptor channels. The results of these studies should facilitate the development of agents that act more specifically on particular receptors. We will use electrophysiological, biochemical, and molecular approaches to study receptors bearing complementary mutations in adjacent or non-adjacent subunits. The specific aims are: Goal 1 - To test whether the zinc binding sites that modulate channel activity in P2X2, P2X3, and P2X4 receptors are within or between subunits, and to define residues that participate in these binding sites. We will also define our understanding about the mechanisms by which zinc promotes channel opening in P2X2 receptors. Goal 2 - To test whether the ATP binding site of P2X receptors is within or between subunits and to define additional residues that are exposed in the ATP binding pocket. These experiments will also test the number of molecules of ATP that must be bound in order to open a channel. Goal 3 - To define the molecular movements that are a consequence of zinc or ATP binding to P2X2 receptors. These experiments are of particular relevance to making progress in understanding and treating pain associated with tissue injury, as P2X2 and P2X3 receptors have been implicated as playing essential roles as sensing the damage and signaling the central nervous system. Having a better understanding of the structure of these receptors should allow the development of new treatments for this type of pain, and so greatly ease the suffering of individuals with burns and other injuries that produce persistent pain.

描述（由申请人提供）：P2X蛋白是ATP封闭通道，它去极化细胞并允许钙进入。 P2X受体几乎在每个组织中表达，包括中枢和周围神经系统的神经元和神经胶质，光滑，骨骼和心肌，耳蜗毛细胞，血小板，大多数白细胞，肝细胞，肝细胞以及肺和内皮细胞胃肠道。该基因家族成员对正常生理的重要性从缺乏的表型范围很明显，在其缺席的情况下，将特定的P2X受体敲除出的小鼠表现出疼痛感的功能障碍，能力使膀胱，肠运动，神经元，神经元，神经元，神经元的能力控制射精，神经系统监测血液中氧气水平的能力，对抗细菌感染的能力和血液凝结的能力。嘌呤能受体场中的一个主要问题是激动剂和拮抗剂的特异性有限，可用于在体内改变ATP信号传导。这里描述的实验的目的是更好地表征允许ATP和变构调节剂打开P2X受体通道的分子机制。这些研究的结果应促进更具体地对特定受体起作用的药物的发展。我们将使用电生理，生化和分子方法来研究在相邻或非粘附亚基中带有互补突变的受体。具体目的是：目标1-测试调节P2X2，P2X3和P2X4受体中通道活性的锌结合位点是否在亚基内或之间，并定义参与这些结合位点的残基。我们还将定义我们对锌在P2X2受体中促进通道开口的机制的理解。目标2-测试P2X受体的ATP结合位点是否在亚基内或之间，并定义在ATP结合袋中暴露的其他残基。这些实验还将测试必须绑定的ATP分子数量才能打开通道。目标3-定义分子运动是锌或ATP与P2X2受体结合的结果。这些实验与在理解和治疗与组织损伤相关的疼痛方面取得进展特别相关，因为P2X2和P2X3受体被暗示是在感觉到中枢神经系统的损害和发出信号时起着至关重要的作用。对这些受体的结构有了更好的了解，应允许为这种疼痛开发新的治疗方法，并极大地减轻了烧伤和其他造成持续疼痛的人的苦难。