Refining a structural model of ENaC

完善 ENaC 的结构模型

• 批准号: 8564819
• 负责人: OSSAMA B KASHLAN
• 金额: $ 2.51万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-01 至 2014-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8564819
• 关键词: ASIC channel; Aldosterone; Bile Acids; Binding; Blood Pressure; Blood Vessels; Cations; Cell surface; Characteristics; Chickens; Cleaved cell; Color; Cues; Cysteine; Data; Distal; Disulfides; Environment; Epithelial; Epithelial Cells; Epithelium; Extracellular Domain; Extracellular Fluid; Extracellular Structure; Family; Family member; Fingers; Gene Family; Goals; Health; Homologous Protein; Human; Ion Channel; Kidney; Location; Lung; Mechanics; Modeling; Mucociliary Clearance; Mutation; Nephrons; Peptide Hydrolases; Peptides; Play; Process; Proteins; Protons; Regulation; Relative (related person); Reporting; Research Design; Site; Smooth Muscle Myocytes; Sodium; Sodium Channel; Sodium Chloride; Stimulus; Stretching; Structural Models; Structure; Taste Perception; Testing; Thumb structure; Tongue; Transmembrane Domain; airway surface liquid; apical membrane; base; blood pressure regulation; comparative; crosslink; disulfide bond; epithelial Na+ channel; extracellular; improved; member; mouse ASIC1 protein; novel; public health relevance; research study; response; shear stress

DESCRIPTION (provided by applicant): The epithelial Na+ channel (ENaC) has a key role in the regulation of extracellular fluid volume and blood pressure. ENaC belongs to a family of ion channels that sense the external environment. ENaC responds to several environmental cues including Na+, Cl-, protons, proteases and shear stress. The ENaC extracellular domains interact with the extracellular environment to sense these cues. Having an accurate model of ENaC structure is therefore critical to understanding ENaC regulation by extracellular factors. Acid sensing ion channel 1 (ASIC1) is the only protein homologous to ENaC whose structure has been reported. The ENaC subunit extracellular domains are largely homologous the corresponding ASIC1 domains, except for the so called finger domain where ENaC subunits have a large insertion. The ENaC finger domain has been implicated in the channel's response to Na+, proteases, and shear stress. The goal of this proposal is to improve upon a structural model of the extracellular domains of the ENaC ? subunit. The proposed studies will investigate the structural interactions that involve a stretch of residues that includes two cysteines in the finger domain insertion. We will then use this information to refine a structural model of the ENaC ? subunit. We previously hypothesized that these two finger domain cysteines formed a disulfide bridged pair, but our preliminary data contradicts this notion. Experiments are proposed to define specific interactions between these cysteines and other specific sites in the channel, and between these cysteines and an inhibitory peptide. We will derive distance constraints from our data and combine them with constraints based on homology to ASIC1 and on other available data to refine a model of the ENaC ? subunit extracellular domains. We will then assess our model by developing novel hypotheses of function to test.

描述（申请人提供）：上皮Na+通道（ENaC）在细胞外液容量和血压的调节中发挥关键作用。 ENaC 属于感知外部环境的离子通道家族。 ENaC 对多种环境信号做出反应，包括 Na+、Cl-、质子、蛋白酶和剪切应力。 ENaC 细胞外结构域与细胞外环境相互作用以感知这些线索。因此，拥有准确的 ENaC 结构模型对于了解细胞外因子对 ENaC 的调节至关重要。酸敏感离子通道 1 (ASIC1) 是唯一与 ENaC 结构已报道的同源蛋白。 ENaC 亚基胞外结构域在很大程度上与相应的 ASIC1 结构域同源，除了所谓的指结构域（其中 ENaC 亚基具有大量插入）。 ENaC 指状结构域与通道对 Na+、蛋白酶和剪切应力的响应有关。该提案的目标是改进 ENaC 细胞外域的结构模型？亚基。拟议的研究将调查涉及指状结构域插入中包含两个半胱氨酸的一段残基的结构相互作用。然后我们将使用这些信息来完善 ENaC 的结构模型？亚基。我们之前假设这两个指状结构域半胱氨酸形成二硫桥对，但我们的初步数据与这一观点相矛盾。建议进行实验来定义这些半胱氨酸与通道中其他特定位点之间以及这些半胱氨酸与抑制肽之间的特定相互作用。我们将从我们的数据中导出距离约束，并将它们与基于 ASIC1 同源性和其他可用数据的约束相结合，以完善 ENaC 的模型？亚基胞外结构域。然后，我们将通过开发新的功能假设来测试来评估我们的模型。