Allosteric ENaC Regulation

上皮性钠离子通道(ENaC)在调节细胞外液容量和血液中起着关键作用 压力。ENAC属于感知外部环境的离子通道家族，它对 包括Na+、Cl-、质子、蛋白水解酶和切应力在内的几种环境信号。这些渠道是如何 对这些细胞外信号的反应还知之甚少。这项提案的目标是确定 外源Na+和蛋白酶调节ENaC的分子机制。它们结合在一起以创建特定的 结构性活跃的通道池，以及仅在细胞外Na+时才被招募的其他通道池 浓度很低。拟议中的基于结构模型的研究将确定分子 这两个因素的调节机制。Na+和抑制肽的结合部位将定义为 以及Na+或抑制肽结合时引起的动态变化。实验将测试是否 酸性口袋含有Na+抑制结合部位，而与酸性口袋相邻的区域含有结合 抑制肽的位置。无论哪种效应器都要引起通道活动的改变，它们必须引起局部 当结合传播到通道孔时构象变化。有人提议用实验来诱捕 使用化学交联和静电效应的通道的功能构象。机械师 了解细胞外因子如何调节ENaC将有助于我们理解ENaC的功能 监管以及相关渠道如何感知其环境。

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 and it responds to several environmental cues including Na+, Cl-, protons, proteases and shear stress. How these channels respond to these extracellular cues is poorly understood. The goal of this proposal is to determine the molecular mechanisms of ENaC regulation by external Na+ and proteases. They combine to create specific pools of channels that are constitutively active, and others that are recruited only when extracellular Na+ concentrations are low. The proposed studies based on structural models will determine molecular mechanisms of regulation by both factors. Binding sites for Na+ and an inhibitory peptide will be defined, as well as dynamic changes induced upon binding of Na+ or an inhibitory peptide. Experiments will test whether an acidic pocket hosts a Na+ inhibitory binding site, and a region adjacent to the acidic pocket hosts the binding site for an inhibitory peptide. For either effector to induce changes in channel activity, they must induce local conformational changes upon binding that propagate to the channel pore. Experiments are proposed to trap functional conformations of the channel using chemical crosslinks and electrostatic effects. A mechanistic understanding of how extracellular factors regulate ENaC will inform our understanding of ENaC functional regulation and how related channels may sense their environments.