High-resolution crystallographic and functional studies of K+ channel gating

K 通道门控的高分辨率晶体学和功能研究

• 批准号: 8290873
• 负责人: Luis Gonzalo Cuello
• 金额: $ 28.12万
• 依托单位: TEXAS TECH UNIVERSITY HEALTH SCIS CENTER
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-01 至 2017-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8290873
• 关键词: Address; Affect; Alkali Metals; Amino Acids; Aromatic Amino Acids; Biological Process; Cell membrane; Cells; Cellular Membrane; Cessation of life; Characteristics; Chemicals; Communication; Complex; Coupled; Coupling; Cysteine; Data; Disease; Drug Design; Elements; Engineering; Functional disorder; Goals; Human body; Immune system; Ions; Kinetics; Knowledge; Lead; Life; Mechanics; Mediating; Membrane; Metabolic; Methods; Molecular; Molecular Conformation; NK Cell Activation; Nature; Pancreas; Pharmaceutical Preparations; Phenylalanine; Physiological Processes; Potassium Channel; Property; Proteins; Resolution; Role; Series; Side; Structure; Therapeutic; X-Ray Crystallography; base; blood glucose regulation; design; disulfide bond; driving force; electrical potential; extracellular; improved; information model; mutant; solute; voltage

DESCRIPTION (provided by applicant): The long-term goal of this project is to determine the structural changes that underlie K+ channel function. These membrane-spanning proteins are critical in controlling the electrical potential difference across the membrane, which, in turn, forms the basis for solute exchange and cellular excitability. It follows that a full understanding of the relation between structure and function in K+ channels is a high priority. Achieving this goal will require knowledge of the conformational changes that mediate two key properties, ion permeation and gating. In this proposal, we will characterize at the atomic level these two properties by trapping the channel complex in different kinetic states. Subsequent structural analysis will address the following fundamental but unanswered questions: 1) what is the high-resolution structure of KcsA trapped in the open and C-type inactivated state? 2) what is the amino-acid network underlying allosteric communication between the activation gate and the selectivity filter of KcsA? and 3) which are the structural changes of the KcsA selectivity filter that lead to C-type inactivation? The answers to these questions will provide us with the structural background needed to understand ion selectivity, permeation, and gating of K+ channels in a dynamic context and eventually will assist in the rational design of drugs for the treatment of K+ channel- related diseases. We will focus our study on an archetypal prokaryotic channel, KcsA, that contains all the structural elements characteristic of K+ channel function (i.e., ion permeation, activation, and C-type inactivation gating) but possesses a structure simple enough to facilitate analysis. A dual approach combining crystallographic and electrophysiological methods will provide high-resolution functional and structural information for this model channel in various kinetic states. PUBLIC HEALTH RELEVANCE: K+ channels are molecules conveniently located in the plasma membrane of all living cells, from which they effectively control the flow of K+ ions coming out of the cell, which is crucial for a large number of physiological processes i.e. activation of natural killer cells in the immune system or the regulation of the blood glucose content by the pancreatic cells. Given the critical role of K+ channel in the normal functioning of the human body, it is not surprising that their dysfunction has catastrophic metabolic consequences that very often lead to death. For this reason, it is of surmount importance to determine what are the structural changes at the atomic level that a K+ channel has to endure when it is doing its biological function, which in turn it will assist us in the design of safer therapeutic drugs to treat many K+ channel related diseases.

描述（由申请人提供）：该项目的长期目标是确定K+通道功能的结构变化。这些跨膜蛋白对于控制整个膜的电势差至关重要，这反过来又构成了溶质交换和细胞兴奋性的基础。因此，充分理解 在K+通道中结构和功能之间的关系是一个高优先级。实现这一目标将需要了解介导两个关键特性的构象变化，即离子渗透和门控。在此提案中，我们将在原子水平上表征这两种特性，通过捕获不同动力学状态的通道复合物。随后的结构分析将解决以下基本但未解决的问题：1）被困在开放和C型灭活状态中的KCSA的高分辨率结构是什么？ 2）激活门和KCSA选择性滤波器之间的变构通信的基础氨基酸网络是什么？ 3）KCSA选择性过滤器的结构变化是导致C型失活的？这些问题的答案将为我们提供理解离子选择性，渗透性和K+通道门控所需的结构背景，并最终将有助于对K+通道相关疾病的治疗药物的合理设计。我们将研究将研究重点放在原型的原核通道KCSA上，其中包含K+通道功能的所有结构元素特征（即离子渗透，激活和C型灭活门），但具有简单的结构 足以促进分析。结合晶体学和电生理方法的双重方法将为高分辨率的功能和结构信息提供 该模型通道在各种动力学状态下。 公共卫生相关性：K+通道是所有活细胞的质膜中方便的分子，它们从中有效地控制了从细胞中流出的K+离子的流动，这对于大量生理过程至关重要。考虑到K+通道在正常功能中的关键作用 人体，他们的功能障碍具有经常导致死亡的灾难性新陈代谢后果并不奇怪。由于这个原因，确定在原子水平上的结构变化是什么，当它在执行其生物学功能时必须忍受的结构变化是什么，这反过来又将有助于我们设计更安全的治疗药物来治疗许多K+通道相关疾病。