Molecular Mechanisms of Desensitization and Drug Modulation in Ligand-Gated Ion C

配体门控离子 C 脱敏和药物调节的分子机制

• 批准号: 8916155
• 负责人: Sudha Chakrapani
• 金额: $ 30.12万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8916155
• 关键词: Affinity; Alcohols; Area; Automobile Driving; Barbiturates; Behavior; Benzodiazepines; Binding; Calorimetry; Cholesterol; Communication; Complex; Consensus; Coupled; Coupling; Drug Design; Drug Modulation; Drug Targeting; Environment; Equilibrium; Event; Fluorescence; Functional disorder; Gated Ion Channel; General anesthetic drugs; Goals; Health; Homologous Gene; Hydration status; Ion Channel Gating; Ions; Lead; Ligand Binding; Ligands; Lipids; Measurement; Measures; Mediating; Membrane; Membrane Lipids; Molecular; Molecular Conformation; Motion; Movement; Nature; Neuromuscular Junction; Neurons; Neurotransmitters; Pharmaceutical Preparations; Pharmacologic Substance; Physiological; Play; Prevention; Property; Proteins; Research; Role; Structure; Synaptic Transmission; Techniques; Testing; Therapeutic; Therapeutic Agents; Thermodynamics; Titrations; Vestibule; Water; Work; base; biophysical properties; conformational conversion; desensitization; design; drug sensitivity; improved; insight; interdisciplinary approach; nervous system disorder; novel strategies; patch clamp; proteoliposomes; reconstitution; research study

DESCRIPTION (provided by applicant): The long-term goal of this proposal is to determine the structural basis for allosteric mechanisms in pentameric ligand gated ion channels (LGIC). Upon neurotransmitter binding, LGIC rapidly switch from closed to open to desensitized channel conformations. Despite a substantial understanding of the physiological and pathophysiological role of desensitization, there is very little structural information available on this state. The fcus of this study is to use bacterial LGIC homologues, GLIC and ELIC, to identify the mechanism of desensitization and to determine how drugs modulate transitions between the closed, open and desensitized states. We propose to apply spectroscopic techniques to functionally defined states of GLIC and ELIC in a membrane environment. Our studies will involve a multidisciplinary approach that includes spectroscopic techniques (EPR and fluorescence), patch-clamp measurements in reconstituted proteoliposomes, and thermodynamic measurements using isothermal titration calorimetry. We will 1) Measure the conformational transitions underlying the desensitized conformation. 2) Determine the role of lipid-water-protein interface in gating transitions. 3) Identify the structural basis for allosteric modulation by general anesthetics and alcohols. These experiments will test the central hypothesis that membrane lipids play a key role in maintaining the conformational integrity, in driving gating transitions, and in governing drug modulation. The proposed studies are expected to be significant in that they will provide a dynamic view of LGIC gating in a physiologically- relevant membrane environment. These findings will establish a structural framework to enhance our understanding of LGIC function at the molecular level and thereby improve therapeutic strategies and drug design.

描述（由申请人提供）：该提案的长期目标是确定五聚体配体离子通道（LGIC）中变构机制的结构基础。在神经递质结合后，LGIC从闭合到脱敏的通道构象。尽管对脱敏的生理和病理生理作用有深刻的了解，但在该状态下，结构信息很少。这项研究的FCU是使用细菌LGIC同源物，胶质和精致，以确定脱敏的机理，并确定药物如何调节封闭，开放和脱敏状态之间的过渡。我们建议将光谱技术应用于膜环境中胶质和ELIC的功能定义状态。我们的研究将涉及一种多学科方法，其中包括光谱技术（EPR和荧光），重建蛋白脂质体的贴片钳测量以及使用等温滴定热量法的热力学测量。我们将1）测量脱敏构象基础的构象转变。 2）确定脂质 - 水 - 蛋白质界面在门控跃迁中的作用。 3）确定一般麻醉和醇的变构调节的结构基础。这些实验将测试膜脂质在保持构象完整性，驱动门控过渡和管理药物调节方面起关键作用的中心假设。预计拟议的研究将具有重要意义，因为它们将在生理相关的膜环境中提供LGIC门控的动态视图。这些发现将建立一个结构框架，以增强我们在分子水平上对LGIC功能的理解，从而改善治疗策略和药物设计。