Ion-selectivity and mechanisms of the Na/K pump

细胞将细胞与其环境分开的细胞膜上的离子（带电原子或分子）浓度存在巨大差异。这种浓度差用于驱动许多重要的细胞过程，并由许多膜泵建立和维持，这些膜泵是负责跨膜运输这些离子的蛋白质。该项目将让本科生（包括在科学领域代表性不足的少数群体成员）和研究生参与对特定膜泵选择和跨膜运输钠离子和钾离子的基本特性的研究。除了生物物理技术培训外，学生还将获得支持参加全国科学会议，在会上展示他们的研究成果。Na/K 泵是电动的，因为它在每个催化循环中将 3 Na 输送出细胞，以换取 2 K 。三分之一的位点可以专门结合 Na（Na/K 泵与其他 P 型 ATP 酶的区别特征）。所有 P 型 ATP 酶在两种主要构象之间交替，E1 具有向内的离子结合位点，E2 具有向外的离子结合位点。 Na/K 泵离子选择性的分子机制仍不清楚，但了解这些对于了解这些蛋白质如何将 ATP 水解的化学能与离子易位所需的机械功耦合起来至关重要。该项目将采用集成高时间分辨率电生理学、计算化学和生物化学的创新方法来研究 Na/K 泵中离子选择的机制。我们实验室现有的晶体结构和结果将用于生成以下方面的假设： (a) 阐明两种主要泵构象中离子选择性的机制； (b) 确定细胞内 Na 结合和外部 Na 释放的顺序； (c) 确定通过 Na/K 泵非耦合 H 流入的途径和机制。

Cells maintain large differences in the concentration of ions (electrically charged atoms or molecules) across the membrane that separates the cell from its environment. This concentration difference is used to drive many essential cellular processes and it is established and maintained by numerous membrane pumps, which are proteins responsible for transporting these ions across the membrane. This project will engage undergraduate students (including members of minorities underrepresented in science) and graduate students in investigations of the fundamental properties by which a particular membrane pump selects and transports sodium and potassium ions across membranes. In addition to training in biophysical techniques, the students will be supported to attend national scientific meetings at which they will be able to present the results of their research.The Na/K pump is electrogenic because it transports 3 Na+ out of the cell in exchange for 2 K+, in each catalytic cycle. One out of three sites can exclusively bind Na+ (a feature that distinguishes the Na/K pump from other P-type ATPases). All P-type ATPases alternate between two major conformations, E1 with inward facing ion binding sites and E2 with outward facing ion binding sites. The molecular mechanisms of ion selectivity by the Na/K pump remain unclear, yet knowledge of these is critical to an understanding of how these proteins couple the chemical energy of ATP hydrolysis to the mechanical work required for ion translocation. This project will study the mechanisms of ion selection in the Na/K pump with an innovative approach that integrates high-temporal-resolution electrophysiology, computational chemistry and biochemistry. The available crystal structures and results from our laboratory will be used to generate hypotheses with respect to (a) Elucidating the mechanisms of ion selectivity in the two major pump conformations; (b) Determining the order of intracellular Na+ binding and external Na+ release; and (c) Identifying the pathway and mechanisms of uncoupled H+ influx through Na/K pumps.