Structural Studies on Prokaryotic Potassium Channels

描述（由申请方提供）：离子通道有助于产生膜电位、受体电位和动作电位。它们是神经科学中许多细胞的基本特征：兴奋性的分子构建块。这些蛋白质涉及所有可兴奋组织的生理学和病理生理学，是许多疾病过程（包括癫痫和心律失常）的基础，并且是临床医学中使用的基本药物的主要靶标。钾通道是结构上相关的离子通道的大超家族的系统发育创始者，所述超家族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研究较多的模型系统。这个提议探讨了钾通道的两个方面：药物结合和电压依赖性.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，我们将试图描述其天然形式的通道的电压敏感域。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射线晶体学来研究两种原核钾通道：KcsA和KvAP。该提案的长期目标是在结构水平上了解离子通道的功能特性。

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.