Structure and Finction of AMPA subtype ionotropic glutamate receptors

AMPA 亚型离子型谷氨酸受体的结构和功能

• 批准号: 8650439
• 负责人: Alexander Sobolevsky
• 金额: $ 34.72万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-30 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8650439
• 关键词: AMPA Receptors; Acids; Adamantane; Agonist; Alzheimer' s Disease; Amyotrophic Lateral Sclerosis; Binding; Binding Sites; Biochemical; Calcium; Cations; Chemicals; Clinical; Complex; Computer Simulation; Crystallization; Cysteine; Data; Detergents; Down-Regulation; Drug Design; Electrophysiology (science); Engineering; Entropy; Epilepsy; Fluorescence; Glutamate Agonist; Glutamate Receptor; Glutamates; Goals; Integral Membrane Protein; Ion Channel; Ischemia; Kinetics; Knowledge; Length; Ligands; Lipids; Location; Mediating; Methods; Modeling; Modification; Molecular; Molecular Conformation; Molecular Sieve Chromatography; Mutation; Neuraxis; Neurons; Neurosciences; Patients; Pharmaceutical Preparations; Polyamines; Primary Lateral Sclerosis; Process; Proteins; Rattus; Regulation; Resolution; Site-Directed Mutagenesis; Structural Models; Structure; Surface; Techniques; Temperature; Testing; Therapeutic; Toxin; Work; base; chemical synthesis; crosslink; desensitization; design; electron density; excitotoxicity; improved; ion channel blocker; nervous system disorder; neuron loss; neurotransmission; public health relevance; receptor; receptor function; research study; response

PROJECT SUMMARY AMPA (¿-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) subtype ionotropic glutamate receptors mediate the majority of excitatory neurotransmission in the central nervous system. Alteration in AMPA receptor function is associated with numerous devastating neurological diseases including Alzheimer's disease, amyotrophic lateral sclerosis, epilepsy and ischemia. Regulation of AMPA receptor activity is therefore an important clinical goal. AMPA receptors function by opening their ion channel for the flow of permeant cations in response to binding of agonist glutamate to the receptor. This process of ion channel opening or activation gating is accompanied by typically slower process of desensitization. Desensitization leads to the closure of the ion channel pore and appears as a 10-500 fold reduction of AMPA receptor- mediated currents in the continuous presence of glutamate. Desensitization represents a natural way of suppressing AMPA receptor activity that protects neurons from the excitotoxic calcium overload. Enhancing or weakening desensitization is therefore an effective way of regulating AMPA receptor activity in pathological conditions. An alternative way to suppress AMPA receptor activity that has therapeutic potential is to use ion channel blockers, molecules that plug the channel pore. At the molecular level, desensitization and ion channel block are expected to have common features that allow them to lock the ion channel in the closed conformation. We plan to study AMPA receptor desensitization and block using a combination of structural and functional approaches and to reveal similarities and differences in these processes. Accordingly, our specific aims are: 1) Obtaining a high-resolution structure of an AMPA receptor in the desensitized state and 2) Building a structural model of ion channel block. AMPA receptor is a challenging target for structure-functional studies because it represents a multimeric integral membrane protein of a large size with typically low expression level. To achieve our goals, we will use a combination of structural and functional approaches including modern crystallographic techniques, Fluorescence-based Size Exclusion Chromatography (FSEC) and electrophysiology. We will use different crystallization methods and temperatures, screen detergents, lipids and ligands to obtain full length AMPA receptor structures in the desensitized and blocker-bound states. We will then combine nascent structural information with electrophysiological recordings and kinetic modeling to figure out mechanisms of AMPA receptor desensitization and block. Achieving our aims will have a significant impact on molecular neuroscience and will result in a new structural/functional model of AMPA receptor that can serve as a dynamic template for theoretical prediction, in silico fitting and chemical synthesis of new compounds that can be tested in different models of neurological diseases and eventually become safe and effective medications.

项目摘要 AMPA（<$-氨基-3-羟基-5-甲基-4-异恶唑丙酸）亚型离子型谷氨酸受体 介导中枢神经系统中的大部分兴奋性神经传递。AMPA的变更 受体功能与包括阿尔茨海默氏症在内的许多破坏性神经系统疾病有关 疾病、肌萎缩性侧索硬化症、癫痫和缺血。AMPA受体活性的调节是 因此是一个重要的临床目标。AMPA受体通过打开其离子通道来发挥作用， 在一些实施方案中，受体可响应于激动剂谷氨酸与受体的结合而释放渗透阳离子。这个过程中的离子通道 打开或激活门控通常伴随着较慢的脱敏过程。脱敏 导致离子通道孔关闭，并表现为AMPA受体减少10-500倍， 介导的电流在连续存在的谷氨酸。脱敏是一种自然的方式， 抑制AMPA受体活性，保护神经元免受兴奋性毒性钙超载。增强或 因此，减弱脱敏作用是调节病理性AMPA受体活性的有效途径。 条件具有治疗潜力的抑制AMPA受体活性的另一种方法是使用离子通道。 通道阻滞剂，堵塞通道孔的分子。在分子水平上，脱敏和离子通道 块预期具有共同的特征，允许它们将离子通道锁定在封闭的 构象我们计划研究AMPA受体脱敏和阻断，使用结构和功能的组合。 功能的方法，并揭示在这些过程中的相似性和差异。因此，我们的具体 目的是：1）获得脱敏状态下AMPA受体的高分辨率结构，和2） 离子通道阻滞结构模型的建立。AMPA受体是一个具有挑战性的结构-功能靶点， 因为它代表了一种大尺寸的多聚体整合膜蛋白， 表达水平。为了实现我们的目标，我们将使用结构和功能方法相结合的方法 包括现代晶体学技术、基于双折射的尺寸排阻色谱法（FSEC） 和电生理学。我们将使用不同的结晶方法和温度，屏幕清洁剂，脂质 和配体以获得脱敏和阻断剂结合状态的全长AMPA受体结构。我们 然后将联合收割机将新生结构信息与电生理记录和动力学建模相结合， 阐明AMPA受体脱敏和阻断的机制。实现我们的目标将产生重大影响。 对分子神经科学的影响，并将导致AMPA受体的新结构/功能模型， 可以作为理论预测、计算机拟合和化学合成新化合物的动态模板。 这些化合物可以在不同的神经系统疾病模型中进行测试，并最终变得安全， 有效的药物。