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: 7570624
• 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/7570624
• 关键词: Architecture; Binding; Catalysis; Cell membrane; Cells; Communication; Complex; Coupled; Coupling; Diffuse; Family; Goals; Heart; Hormones; Hypoxia; Integral Membrane Protein; Membrane; Membrane Potentials; Molecular; Molecular Conformation; NADP; Neurons; Oxidation-Reduction; Oxidative Stress; Oxidoreductase; Physiological Processes; Physiology; Potassium Channel; Proteins; Reagent; Research Personnel; Resolution; Rest; Side; Structure; Testing; Therapeutic; X-Ray Crystallography; base; cofactor; macromolecule; member; potassium ion; programs; response; therapeutic development; 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）是整体膜蛋白，响应于膜 电压变化，催化钾离子以扩散在细胞膜上。 KV通道调节 膜兴奋性，对于许多生理过程，例如节奏的节奏至关重要 心脏，神经元之间的交流和激素的分泌。 beta亚基（KVP） 振荡器类型的KV通道（KV1）永久连接到通道的细胞内侧，牵涉 在氧化应激和低氧条件下的通道调节中。序列保存建议 KVP类似于Aldo-Keto还原酶（AKR），并且KVP的晶体结构表明它具有 规范AKR折叠，紧密结合的辅助烟酰胺腺苷二核苷酸磷酸盐（NADPH）和 催化的右几何形状中高度保守的催化残基发生。但是，酶促 KVP的活动从未得到证明。该提案的总体目标是研究KVP 作为功​​能性AKR，调节通道函数，以研究酶活性如何耦合到 渠道活动，并发展原子水平对耦合机制的理解。长期 该项目的目标是了解KVP的生理学以及管理KV渠道的原则 调制。我们最近已经确定了几个KVP底物，并证明KVP是一种功能 Aldo-Keto还原酶。我们还发现，仅当KVP是共同的时，底物才能调节通道函数 表达。这些令人兴奋的新结果导致我们假设：1）KVP的AKR函数与 KV通道功能； 2）通过细胞内结构域与KVP之间的相互作用来实现耦合 3）KVP的不同氧化还原状态具有不同的构象，可引起构象变化 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：检查信道活动与KVP的AKR活性之间的功能耦合。 目标2：研究耦合机制的分子碱基。 目标3：研究耦合的结构底座。 该项目的结果将有助于我们了解各种KV渠道家庭的调制，并将有助于 开发靶向大分子复合物的治疗试剂。