The structural and mechanistic basis of K(+) translocation by the KtrAB system

K（+）转运蛋白超家族（SKT）单位蛋白，来自除动物以外的所有生物王国，在质膜上转运K（+）和/或Na（+）。这些蛋白质是渗透调节、pH稳态和抵抗高盐度和干燥的关键成分。SKT蛋白与KcsA等K（+）通道密切相关，但也显示出一些与功能改变有关的显著差异。在SKT蛋白中，真核成员仅由易位亚基组成，而原核成员由至少一个额外的细胞质亚基调节。目前的假设是，原核生物成员的易位亚基单独表现出通道样活性，而调节亚基可能根据需要介导转运蛋白活性到系统中以实现其生理作用。在这里，我们将研究细菌的Na（+）依赖性K（+）易位KtrAB系统，其中KtrB是K（+）易位亚基，KtrA介导Na（+）依赖性，K（+）特异性和摄取速度的增加。KtrB包含一个独特的门控序列基序，但系统的结构-功能关系尚未解决。通过使用x射线晶体学、EPR测量、输运和电生理测量，我们将阐明以下问题：(i) KtrA对KtrB的结构和功能影响，（ii）在分子水平上控制系统内K（+）通量的机制，以及（iii） Na（+）是作为配体激活通道还是作为耦合离子共同运输。

The superfamily of K(+) transporters (SKT) units proteins from all kingdoms of life but animals that translocate K(+) and/or Na(+) over the plasma membrane. These proteins are key components of osmotic regulation, pH homeostasis and resistance to high salinity and dryness. SKT proteins are closely related to K(+) channels like KcsA but also show several striking differences which are assigned to an altered function. Within the SKT proteins eukaryotic members consist of only the translocating subunit while prokaryotic members are regulated by at least one additional cytoplasmic subunit. The current hypothesis is that the translocating subunits of prokaryotic members alone show channel-like activities while the regulatory subunits may mediate transporter activity to the system as needed to fulfill its physiological role. Here, we will study the Na(+)-dependent K(+)-translocating KtrAB system of bacteria, in which KtrB is the K(+)-translocating subunit and KtrA mediates Na(+) dependency, K(+) specificity and an increase in uptake velocity. KtrB was shown to contain a unique sequence motif for gating but the structure - function relationship of the system remained unsolved. By use of X-ray crystallography, EPR measurements, transport and electrophysiological measurements we will elucidate the questions (i) which structural and by that also functional impact KtrA has on KtrB, (ii) which mechanisms control K(+) flux within the system on a molecular level and (iii) whether Na(+) acts as ligand to activate channeling or is co-transported as coupling ion.