Molecular Mechanism of Cation Channel Selectivity

描述（由申请人提供）：离子穿过生物膜的转移对于神经兴奋、肌肉细胞收缩、信号转导和激素分泌等生理过程至关重要。离子通道通过在膜内提供通道（离子传导孔）以允许特定离子沿其电化学梯度向下移动而发挥重要作用。在人类中，离子通道存在于几乎所有的组织中，提供各种生理学上的基本功能。由于它们在人体中的普遍性和重要性，通道的功能障碍通常是导致广泛的人类病理学的原因。选择特定离子种类的能力被称为离子选择性，并且是定义离子通道功能的基本性质。在四聚体阳离子通道中，其包括单个最大的离子通道家族，包括K+、Ca 2+、Na+和环核苷酸门控（CNG）通道，选择性通常是离子通道孔（选择性过滤器）的一部分内的独特结构和化学环境的直接结果，所述选择性过滤器在不同通道之间是不同的。我们的理解，在这组通道的离子选择性的分子细节，已经走过了漫长的道路与进步的遗传，生物化学和电生理分析的离子通道，最近，结构表征的几个成员开始的开创性工作的KcsA K+通道结构和随后的工作对其他K+选择性通道。虽然这些结构研究提供了一个直接的可视化的选择性过滤器的K+通道，决定因素contributingto K+选择性仍然是在激烈的争论。此外，有很少的结构信息可用于其他四聚体阳离子通道和它们的离子选择性的分子基础仍然不清楚。我们研究的总体目标是了解四聚体阳离子通道中离子选择性的结构基础。利用极高分辨率的晶体结构的几个模型蛋白代表的离子传导孔的选择性和非选择性通道，我们的目标是阐明离子选择性的基本原则，在两组生理上必需的阳离子通道。一种是非选择性的、Ca 2+可渗透的环核苷酸门控（CNG）通道，使用来自蜡状芽孢杆菌的NaK及其CNG模拟嵌合体作为模型系统;另一种是K+选择性通道，使用K+选择性NaK突变体（NaK 2K）和来自热自养甲烷杆菌的MthK K+通道作为模型系统。本研究将结合蛋白质晶体学、电生理学和蛋白质化学合成的方法进行

DESCRIPTION (provided by applicant): Transfer of ions across biological membranes is central to physiological processes like nerve excitation, muscle cell contraction, signal transduction and hormone secretion. Ion channels play a vital role by providing a passageway, the ion conduction pore, within membranes to allow specific ions to traverse down their electrochemical gradient. In humans, ion channels are found in nearly all tissues serving a variety of physiologically essential functions. Because of their prevalence and importance in the human body, dysfunction of a channel is often to blame for a wide range of human pathologies. The ability to select for specific ionic species is known as ion selectivity and is a fundamental property defining ion channel function. In tetrameric cation channels, which comprise the single largest family of ion channels including the K+, Ca2+, Na+, and cyclic nucleotide-gated (CNG) channels, selectivity is usually a direct consequence of the unique structural and chemical environment within part of the ion channel pore, the selectivity filter, which is distinct among different channels. Our understanding of the molecular details governing ion selectivity in this group of channels has come a long way with the advancement of genetic, biochemical, and electrophysiological analysis of ion channels and, more recently, the structural characterization of several members beginning with the ground breaking work of the KcsA K+ channel structure and subsequent work on other K+ selective channels. Although these structural studies offer a direct visualization of the selectivity filter of K+ channels, the determinant factors contributingto K+ selectivity are still under heated debate. Moreover, there is little structural information available for other tetrameric cation channels and the molecular basis for their ion selectivity remains unclear. The overall goal of our research is to understand the structural basis of ion selectivity in tetrameric cation channels. Taking advantage of the extremely high resolution crystal structures of several model proteins representing the ion conduction pores of both selective and non-selective channels, we aim to elucidate basic principles of ion selectivity in two groups of physiologically essential cation channels. One is the non-selective, Ca2+ permeable cyclic nucleotide-gated (CNG) channels, using NaK from Bacillus cereus and its CNG- mimicking chimeras as model systems; the other is K+ selective channels, using a K+ selective NaK mutant (NaK2K) and the MthK K+ channel from Methanobacterium thermoautotrophicum as model systems. A combined approach of protein crystallography, electrophysiology and protein chemical synthesis will be employed in the proposed studies