Structural mechanism of K channel modulation by cellular redox state

细胞氧化还原态调节 K 通道的结构机制

• 批准号: 7369847
• 负责人: Ming Zhou
• 金额: $ 39.08万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-03-15 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7369847
• 关键词: Architecture; Binding; Catalysis; Cell membrane; Cells; Communication; Complex; Condition; Coupled; Coupling; Development; Diffuse; Family; Goals; Heart; Hormones; Hypoxia; Integral Membrane Protein; Membrane; Membrane Potentials; Molecular; Molecular Conformation; NADP; Neurons; Oxidation-Reduction; Oxidative Stress; Physiological Processes; Physiology; Potassium Channel; Proteins; Reagent; Research Personnel; Resolution; Rest; Side; Structure; Testing; Therapeutic; X-Ray Crystallography; base; carbonyl reductase (NADPH); cofactor; macromolecule; member; potassium ion; programs; response; voltage

Voltage-dependent potassium channels (Kv) are integral membrane proteins that, in response to membrane voltage changes, catalyze potassium ions to diffuse across the cell membrane. Kv channels regulate membrane excitability and are essential to many physiological processes such as the rhythmic beating of heart, the communication between neurons, and the secretion of hormones. The beta subunit (Kvp) of the Shaker type Kv channels (Kv1) permanently attaches to the intracellular side of a channel and is implicated in channel modulation during oxidative stresses and hypoxic conditions. Sequence conservation suggests that Kvp resembles an aldo-keto reductase (AKR), and the crystal structure of a Kvp showed that it has a canonical AKR fold, a tightly bound cofactor nicotinamide adenine dinucleotide phosphate (NADPH), and highly conserved catalytic residues in the right geometry for catalysis to happen. However, the enzymatic activity of Kvp has never been demonstrated. The overall objectives of this proposal are to examine how Kvp as a functional AKR modulates channel function, to investigate how the enzymatic activity is coupled to channel activities, and to develop an atomic level understanding of the coupling mechanism. The long-term goals of the project are to understand the physiology of Kvp, and the principles governing Kv channel modulations. We have recently identified several Kvp substrates, and demonstrated that Kvp is a functional aldo-keto reductase. We also found that the substrates modulates channel function only when a Kvp is co- expressed. These exciting new results led us to hypothesize that: 1) the AKR function of Kvp is coupled to Kv channel functions; 2) the coupling is achieved through interactions between intracellular domains and Kvp 3) different redox states of Kvp have different conformations that induce a conformational change of a channel domain. To test these hypotheses, we propose the following three specific aims: Aim 1: To examine the functional coupling between channel activities and the AKR activity of Kvp. Aim 2: To investigate the molecular bases of the coupling mechanism. Aim 3: To investigate the structural bases of the coupling. Results from this project will help us understand modulations of the various Kv channel families, and will help develop therapeutic reagents that target the macromolecule complex.

电压依赖性钾通道 (Kv) 是完整的膜蛋白，响应膜 电压变化，催化钾离子扩散穿过细胞膜。 Kv通道调节 膜的兴奋性对于许多生理过程至关重要，例如有节奏的跳动 心脏、神经元之间的通讯以及激素的分泌。 β 亚基 (Kvp) 摇床型 Kv 通道 (Kv1) 永久附着在通道的细胞内侧并受到牵连 氧化应激和缺氧条件下的通道调节。序列守恒表明 Kvp 类似于醛酮还原酶 (AKR)，并且 Kvp 的晶体结构表明它具有 典型的 AKR 折叠，紧密结合的辅因子烟酰胺腺嘌呤二核苷酸磷酸 (NADPH)，以及 高度保守的催化残基以正确的几何形状进行催化。然而，酶促 Kvp 的活动从未被证明。该提案的总体目标是研究 Kvp 如何 作为功​​能性 AKR 调节通道功能，以研究酶活性如何与 通道活动，并形成对耦合机制的原子级理解。长期来看 该项目的目标是了解 Kvp 的生理学以及控制 Kv 通道的原理 调制。我们最近鉴定了几种 Kvp 底物，并证明 Kvp 是一种功能性的 醛酮还原酶。我们还发现，只有当 Kvp 共同作用时，基底才会调节通道功能。 表示。这些令人兴奋的新结果使我们做出假设：1）Kvp 的 AKR 函数与 Kv通道功能； 2）耦合是通过细胞内结构域和Kvp之间的相互作用实现的 3）Kvp的不同氧化还原态具有不同的构象，从而引起构象变化 通道域。为了检验这些假设，我们提出以下三个具体目标： 目标 1：检查通道活动与 Kvp AKR 活动之间的功能耦合。 目标2：研究偶联机制的分子基础。 目标 3：研究联轴器的结构基础。 该项目的结果将帮助我们了解各种 Kv 通道系列的调制，并将有助于 开发针对大分子复合物的治疗试剂。