Mass spectrometric studies on potassium channels

钾通道的质谱研究

• 批准号: 6620406
• 负责人: Adrian Gross
• 金额: $ 14.7万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-12-10 至 2004-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6620406
• 关键词: Xenopus oocyte; alkylation; binding sites; cysteine; electrospray ionization mass spectrometry; high performance liquid chromatography; intermolecular interaction; mass spectrometry; membrane structure; potassium channel; protein structure; protein structure function; site directed mutagenesis; structural biology; technology /technique development

DESCRIPTION (provided by applicant): Voltage-dependent 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. Despite the obvious biological and clinical importance of these proteins, we still know relatively little about their structure. Although important progress has been made in understanding ion permeation in potassium channels, large parts of these essential proteins remain structurally unresolved. This proposal explores a novel technique for addressing structural questions involving ion channels and other membrane proteins. We will use a combination of cysteine scanning mutagenesis, alkylation, and mass spectrometry to study structural aspects of two potassium channels, KcsA and Shaker. We refer to this novel technique as site-directed mass tagging. The long-term goal of this proposal is to provide low-resolution structural information about functional ion channels. The proposal is high risk in the sense that we do not yet know whether the technique will fulfill its potential and prove to be a useful structural tool. If successful, however, site-directed mass tagging may be able to provide important insight into the gross structure of many rare membrane proteins. It is anticipated that results from this study will provide new insight into the general structural layout of Shaker.

描述（由申请人提供）：电压依赖性离子通道是 在产生膜电位，受体电位和 动作潜力。它们是一个分子构建块 神经科学中许多细胞的基本特征：兴奋性。这些 蛋白质与所有可激发的生理学和病理生理有关 组织，包括癫痫和心律不齐的许多疾病过程的基础， 并且是临床医学中使用的必需药物的主要靶标。尽管有 这些蛋白质的显而易见的生物学和临床重要性，我们仍然知道 关于它们的结构相对较少。虽然重要的进步已经 在了解钾通道中的离子渗透中，大部分 这些必需蛋白在结构上仍未解决。 该建议探讨了一种解决结构性问题的新技术 涉及离子通道和其他膜蛋白。我们将使用一个组合 半胱氨酸扫描诱变，烷基化和质谱法的研究 两个钾通道的结构方面，KCSA和振动筛。我们指的是 新技术作为定向的质量标记。这个长期目标 建议是提供有关功能的低分辨率结构信息 离子通道。从我们尚不知道的意义上说，该提议是高风险 该技术是否会发挥其潜力并证明是有用的 结构工具。但是，如果成功的话，则以网站为导向的质量标签可能能够 为许多稀有膜的总结构提供重要的见解 蛋白质。预计这项研究的结果将提供新的 深入了解振动筛的一般结构布局。