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）：P2 X蛋白是ATP门控通道，可使细胞去极化并允许钙进入。P2 X受体在几乎所有组织中表达，包括中枢和外周神经系统的神经元和神经胶质、平滑肌、骨骼肌和心肌、耳蜗毛细胞、血小板、大多数类别的白色血细胞、肝细胞以及肺和胃肠道中的内皮细胞。该基因家族的成员对于正常生理学的重要性从其缺失时所见的表型范围是显而易见的。敲除特定P2 X受体的小鼠显示疼痛感知、排空膀胱的能力、肠道运动、射精的神经元控制、神经系统监测血液中氧水平的能力、对抗细菌感染的能力、和血液凝固。嘌呤能受体领域的一个主要问题是可用于改变体内ATP信号传导的激动剂和拮抗剂的特异性有限。本文所述实验的目的是更好地表征允许ATP和变构调节剂打开P2 X受体通道的分子机制。这些研究的结果应该有助于开发对特定受体更特异性地起作用的药物。我们将使用电生理学，生物化学和分子方法来研究在相邻或不相邻亚基中具有互补突变的受体。具体目标是：目的1 -测试调节P2 X2、P2 X3和P2 X4受体通道活性的锌结合位点是否在亚基内或亚基之间，并确定参与这些结合位点的残基。我们还将定义我们对锌促进P2 X2受体通道开放的机制的理解。目标2 -测试P2 X受体的ATP结合位点是否在亚基内或亚基之间，并确定暴露在ATP结合口袋中的其他残基。这些实验还将测试为了打开通道而必须结合的ATP分子的数量。目标3 -确定锌或ATP与P2 X2受体结合的分子运动。这些实验对于理解和治疗与组织损伤相关的疼痛具有特别的相关性，因为P2 X2和P2 X3受体被认为在感知损伤和向中枢神经系统发出信号方面发挥重要作用。更好地了解这些受体的结构应该允许开发这种类型疼痛的新治疗方法，从而大大减轻烧伤和其他产生持续疼痛的伤害的个体的痛苦。