3D Structure and Function of Ligand-Gated Ion Channels

配体门控离子通道的 3D 结构和功能

• 批准号: 6858588
• 负责人: James E Gouaux
• 金额: $ 25.55万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-03-19 至 2005-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6858588
• 关键词: AMPA receptors; NMDA receptors; X ray crystallography; Xenopus oocyte; affinity chromatography; analytical ultracentrifugation; cell line; conformation; glutamate receptor; immunoaffinity chromatography; ion exchange chromatography; ligands; membrane channels; protein structure function; receptor binding; site directed mutagenesis; structural biology; voltage /patch clamp; western blottings

DESCRIPTION (provided by applicant): Ionotropic glutamate receptors mediate the majority of excitatory signal transmission in the mammalian nervous system and are essential to its normal development and function. There are three distinct and well defined receptor subtypes that are the AMPA, kainate and NMDA receptors. Ionotropic glutamate receptors 'work' by coupling the binding of glutamate to gating, or opening, of a transmembrane ion channel. Because ionotropic glutamate receptors are linchpins of the human nervous system, they have been the target of a large number of pharmacological investigations and, therefore, a vast number of agonists, antagonists and allosteric effectors are known. Because in many cases these molecules elicit specific functional responses, they serve as excellent probes of the relationships between receptor structure and function. A substantial portion of the research proposed in this application is aimed at determining atomic resolution structures of the extracellular domains of AMPA and NMDA receptors, and understanding how agonists, antagonists and allosteric modulators exert their functional effects on the receptor. In addition, we have recently discovered bacterial homologs of the eukaryotic receptors and these molecules are promising species for over expression, crystallization and structure determination. Thus, a second element of the research proposed in this application is concentrated on determining the high resolution, three-dimensional structure of a bacterial receptor, in order to understand how ligand-binding is coupled to channel gating. A final element of the research proposed in this application is related to testing structure-based mechanistic hypotheses using site-directed mutagenesis and electrophysiology. By carrying out two electrode voltage clamp experiments in my laboratory, we will be able to evaluate the validity of specific mechanistic hypotheses. Taken together, the overall aims of the proposed research are to provide new and deep insights into the relationships between structure and function in ionotropic glutamate receptors. The resulting atomic structures and the mechanistic schemes will likely prove useful in the design of novel molecules that may have significant therapeutic value.

描述（由申请人提供）：离子型谷氨酸受体介导哺乳动物神经系统中的大多数兴奋性信号传播，对于其正常发育和功能至关重要。 AMPA，海谷和NMDA受体有三种不同且定义明确的受体亚型。通过耦合谷氨酸与跨膜离子通道的门控或开放的结合，离子型谷氨酸受体“工作”。由于离子型谷氨酸受体是人类神经系统的关键，因此它们是大量药理研究的靶标，因此已知大量激动剂，拮抗剂和变构效应子。因为在许多情况下，这些分子会引起特定的功能响应，因此它们是对受体结构与功能之间关系的极好探针。本应用中提出的大部分研究旨在确定AMPA和NMDA受体细胞外结构域的原子分辨率结构，并了解激动剂，拮抗剂和变构调节剂如何对受体发挥其功能作用。此外，我们最近发现了真核受体的细菌同源物，这些分子是过度表达，结晶和结构测定的有前途的物种。因此，本应用中提出的研究的第二个要素集中于确定细菌受体的高分辨率，三维结构，以了解如何将配体结合耦合到通道门控。本应用中提出的研究的最终元素与使用位置定向的诱变和电生理学测试基于结构的机械假设有关。通过在我的实验室进行两个电极电压夹具实验，我们将能够评估特定机械假设的有效性。 综上所述，拟议的研究的总体目的是为离子型谷氨酸受体的结构和功能之间的关系提供新的深刻见解。所得的原子结构和机械方案可能会被证明在可能具有显着治疗值的新分子的设计中有用。