Structural Basis for Antiarrhythmic Drug Action

抗心律失常药物作用的结构基础

• 批准号: 10364048
• 负责人: WILLIAM A CATTERALL
• 金额: $ 74.39万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-06 至 2026-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10364048
• 关键词: Action Potentials; Affinity; Anti-Arrhythmia Agents; Arrhythmia; Binding; Brugada syndrome; Calcium; Calcium Channel; Cardiac; Cardiac Myocytes; Cells; Clinical; Complex; Coupling; Cryoelectron Microscopy; Crystallization; Cytosol; Dependence; Dihydropyridines; Dilated Cardiomyopathy; Disulfides; Drug Antagonism; Drug Receptors; Drug usage; Elderly; Electron Microscopy; Functional disorder; Image; Inherited; Ion Channel; Ions; Lead; Learning; Length; Life; Lipid Bilayers; Lipids; Long QT Syndrome; Maps; Membrane Potentials; Methods; Modeling; Molecular; Molecular Conformation; Molecular Target; Mutation; Pathogenicity; Pharmaceutical Preparations; Prevention; Resolution; Rest; Role; Safety; Side; Site; Sodium; Sodium Channel; Specificity; Structure; Surface; Timothy syndrome; Work; X-Ray Crystallography; base; crosslink; cryogenics; design; drug action; drug mechanism; heart rhythm; improved; insight; next generation; phenylalkylamine; prevent; receptor; reconstitution; sensor; voltage

Voltage-gated sodium (Nav1.5) channels initiate action potentials and voltage-gated calcium (Cav1.2) channels initiate excitation/contraction coupling in cardiac myocytes. They are molecular targets for mutations that cause arrhythmias and for antiarrhythmic drugs (AADs) used in control and prevention of life-threatening arrhythmias. We have determined the structures of the bacterial Nav channel NavAb and the model calcium channel CavAb by X-ray crystallography, and we have determined the structure of the primary cardiac Nav channel Nav1.5 at high resolution by cryogenic electron microscopy (cryo-EM). This work gave new insight into the structure of the ion selectivity filter and mechanism of Na and Ca selectivity, the structure of the voltage sensors and mechanisms of voltage-dependent gating, the activation and inactivation gates and their functional interaction, the receptor sites for AADs, and the mechanism of access of AADs to their receptor site through the open activation gate and fenestrations in the sides of the pore. Mutations in Nav1.5 that cause inherited cardiac arrhythmias, including Dilated Cardiomyopathy and Long QT Syndrome, map onto the pore module, voltage sensor, activation gate, and fast inactivation gate of Nav and Cav channels, opening the way to probing the pathophysiology of these mutations at the structural level. Here we will investigate the structural basis for Nav and Cav channel function, the complex pore-blocking mechanisms of AADs, and the mechanisms underlying the pathophysiological effects of arrhythmia mutations. Aim 1. We will determine the structure of Nav1.5 channels in resting, open, and inactivated states and analyze the molecular mechanisms for ion selectivity and conductance in the open state of Nav1.5. Aim 2. We will prepare complexes of Nav1.5 in the closed, open, and inactivated states with AADs bound, and we will resolve their structures at high resolution. We will probe structural differences in the drug- receptor complexes formed by Class IA, IB, and IC AADs in order to understand the structural basis for the differences in drug action that lead to their different clinical uses. We will examine the role of the fenestrations in resting-state block by Class IA, IB, and IC AADs. Aim 3. We will insert mutations that cause Dilated Cardiomyopathy and Long QT Syndrome Type-3 into NavAb and Nav1.5, characterize their pathophysiological effects on Na currents and gating pore currents, and resolve their structures at high resolution by X-ray crystallography and/or cryo-EM. Aim 4. We will insert mutations that cause Timothy Syndrome into CavAb and Cav1.2, determine their pathophysiological effects on Ca and Ba currents, and resolve their structures at high resolution by X-ray crystallography and cryo-EM. Overall, these studies open the exciting possibility of understanding cardiac Nav and Cav channels in atomic detail in native and pathogenic conformations and learning how to manipulate the structures of AADs to make them more specific, more effective, and safer.

电压门控钠（NAV1.5）通道启动动作电位和电压门控钙（CAV1.2）通道 在心肌细胞中启动激发/收缩耦合。它们是导致突变的分子靶标 心律失常和用于控制和预防威胁生命心律失常的抗心律失常药物（AAD）。 我们已经确定了细菌NAV通道Navab和模型钙通道Cavab的结构 通过X射线晶体学，我们已经确定了主要心脏NAV通道NAV1.5的结构 高温电子显微镜（Cryo-EM）高分辨率。这项工作给了新的见解 离子选择性滤波器和Na和Ca选择性的机理，电压传感器的结构和机制的结构 电压依赖性门，激活和失活门及其功能相互作用，受体 AAD的站点，以及通过开放激活门进入其受体位点的机制， 孔的侧面。导致遗传心律失常的NAV1.5突变，包括 扩张的心肌病和长QT综合征，映射到孔模块，电压传感器，激活门， 和NAV和CAV通道的快速灭活门，为探测这些病理生理的道路开辟了道路 结构层的突变。在这里，我们将研究NAV和CAV通道功能的结构基础， AAD的复杂孔隙阻滞机制以及病理生理作用的基础机制 心律不齐突变。目标1。我们将确定静止，打开和 灭活状态并分析开放状态下离子选择性和电导的分子机制 NAV1.5。目标2。我们将在封闭，开放和灭活状态中准备NAV1.5的复合物 绑定，我们将以高分辨率解决它们的结构。我们将探测药物的结构差异 由IA类，IB和IC AAD形成的受体复合物，以了解该结构基础 导致其不同临床用途的药物作用差异。我们将研究Fenestrations在 按IA，IB和IC AADS划分的静止状态块。目标3。我们将插入引起扩张的突变 心肌病和长QT综合征3型Navab和Nav1.5的病理生理特征 对Na电流和门控孔电流的影响，并通过X射线高分辨率解决其结构 晶体学和/或冷冻EM。目标4。我们将插入引起蒂莫西综合症的突变中的cavab和 Cav1.2，确定其对CA和BA电流的病理生理影响，并在高处解决其结构 X射线晶体学和冷冻EM的分辨率。总体而言，这些研究开辟了令人兴奋的可能性 了解本地和致病构象原子细节中的心脏NAV和CAV通道以及 学习如何操纵AAD的结构，以使其更具体，更有效和更安全。