Structural Studies on Prokaryotic Potassium Channels

描述(由申请人提供)：离子通道在膜电位、受体电位和动作电位的产生中起重要作用。它们是神经科学中许多细胞的一个基本特征：兴奋性的分子构建块。这些蛋白与所有可兴奋组织的生理学和病理生理学有关，是包括癫痫和心律失常在内的许多疾病过程的基础，也是临床药物使用的主要靶点。钾通道是一个结构相关的离子通道超家族的系统发育创始人，包括核苷酸门控通道、钠通道和钙通道。钾通道通常由四个相同的亚基以四重对称的方式组装而成。这种相当简单的结构蓝图和只有一种亚基类型的实质性实用优势使钾通道成为一个被广泛研究的模型系统。这项建议探讨了钾通道的两个方面：药物结合和电压依赖。在目标1中，我们将确定药物-通道相互作用的结构基础。在目标2中，我们将尝试以其本机形式描述通道的电压感应域。我们将结合半胱氨酸诱变、生物化学、电生理学、定点自旋标记和X射线结晶学来研究两种原核钾通道：KCSA和KvAP。这一提议的长期目标是在结构水平上了解离子通道的功能属性。 与公共健康相关：离子通道是一种膜蛋白，允许离子从细胞膜的一侧传递到另一侧。它们是产生人体内所有电信号的分子硬件。这些信号被用来协调心脏的跳动，并构成中枢和外周神经系统复杂功能的基础。当离子通道发生故障时，严重的疾病接踵而至，如心律失常、癫痫，甚至死亡。离子通道也是重要的药物靶点。作用于离子通道的药物在临床上每天都在使用，往往会导致严重的药物副作用。确定这些渠道的确切结构和作用机制将有助于开发更安全和更有效的药物。

DESCRIPTION (provided by applicant): Ion channels are instrumental in the generation of membrane potential, receptor potential, and action potential. They are the molecular building blocks for what is an essential characteristic of many cells in neuroscience: excitability. These proteins are implicated in the physiology and pathophysiology of all excitable tissues, underlie many disease processes including epilepsies and arrhythmias, and are major targets of essential drugs used in clinical medicine. Potassium channels are the phylogenetic founders of a large superfamily of structurally related ion channels that includes nucleotide gated channels, sodium channels, and calcium channels. Potassium channels are typically assembled from four identical subunits in a four-fold symmetrical fashion. This rather simple structural blueprint and the substantial practical advantage of only one subunit type have made potassium channels a much studied model system. This proposal explores two aspects of potassium channels: drug binding and voltage- dependence. In aim 1 we will determine the structural basis of the drug-channel interaction. In aim 2 we will attempt to describe the voltage-sensing domain of the channel in its native form. We will use a combination of cysteine mutagenesis, biochemistry, electrophysiology, site-directed spin labeling, and X-ray crystallography to study two prokaryotic potassium channels: KcsA and KvAP. The long-term goal of this proposal is to understand functional properties of ion channels at a structural level. PUBLIC HEALTH RELEVANCE: Ion channels are membrane proteins that allow ions to pass from one side of the cell membrane to the other. They are the molecular hardware that generates all electrical signals in the human body. These signals are used to coordinate the beating of the heart and underlie the complex function of the central and peripheral nervous system. When ion channels malfunction, serious maladies ensue, such as heart arrhythmias, epilepsy, and even death. Ion channels are also important drug targets. Drugs effecting ion channels are used every day in clinical medicine and are often implicated in serious drug side effects. Determining the precise structure and mechanism of action of these channels will allow for the development of safer and more effective drugs.