Ion-selectivity and mechanisms of the Na/K pump

细胞在隔膜上的离子(带电的原子或分子)的浓度保持着很大的差异，该膜将细胞与其环境分开。这种浓度差被用来驱动许多基本的细胞过程，它是由许多膜泵建立和维持的，膜泵是负责跨膜运输这些离子的蛋白质。该项目将邀请本科生(包括在科学界代表性不足的少数族裔)和研究生研究特定膜泵选择和跨膜传输钠离子和钾离子的基本性质。除了生物物理技术培训外，学生们还将被支持参加国家科学会议，在会上展示他们的研究成果。钠/钾泵是电生的，因为在每个催化循环中，它将3个钠输送到细胞外，以换取2个K。每三个位点中就有一个能与Na结合(这是Na/K泵区别于其他P型ATPase的一个特征)。所有的P型ATPase都在两个主要构象之间交替，E1具有内向的离子结合位点，而E2具有面向外向的离子结合位点。Na/K泵选择性离子的分子机制尚不清楚，但了解这些机制对于了解这些蛋白质如何将ATP水解的化学能与离子转移所需的机械功相结合是至关重要的。本项目将采用一种集高时间分辨率电生理学、计算化学和生物化学于一体的创新方法来研究Na/K泵的离子选择机制。我们实验室现有的晶体结构和结果将被用来产生如下假说：(A)阐明两个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；以及(C)确定通过Na/K泵非偶联H内流的途径和机制。

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.