Expression, purification and biochemical characterization of the cytoplasmic loop of PomA, a stator component of the Na(+) driven flagellar motor.
Expression, purification and biochemical characterization of the cytoplasmic loop of PomA, a stator component of the Na(+) driven flagellar motor.
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DOI：
10.2142/biophysics.9.21
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发表时间：
2013
期刊：
影响因子：
--
通讯作者：
Homma M
中科院分区：
文献类型：
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作者：
Abe-Yoshizumi R;Kobayashi S;Gohara M;Hayashi K;Kojima C;Kojima S;Sudo Y;Asami Y;Homma M
嵌在细菌膜中的鞭毛马达是由特定离子流驱动的分子机器。每个电机都由定子和转子组成，这些部件的相互作用被认为会产生扭矩。Na+内流通过溶藻弧菌的PomA/PomB定子复合物耦合到扭矩产生，并推测引发PomA的胞质结构域的结构变化，其与C环中的转子蛋白FliG相互作用以驱动旋转。在这项研究中，我们试图过量生产PomA的胞质环（PomA-Loop），但它是不溶性的。因此，我们制备了具有小的可溶性标签（GB 1）的融合蛋白，其允许我们表达和表征重组蛋白。PomA环的结构似乎非常细长或具有松散的三级结构。当PomA-Loop蛋白在E.大肠杆菌，轻微的显性效应，观察运动。我们的结论是，细胞质环单独保留了一定的功能。
Flagellar motors embedded in bacterial membranes are molecular machines powered by specific ion flows. Each motor is composed of a stator and a rotor and the interactions of those components are believed to generate the torque. Na+ influx through the PomA/PomB stator complex of Vibrio alginolyticus is coupled to torque generation and is speculated to trigger structural changes in the cytoplasmic domain of PomA that interacts with a rotor protein in the C-ring, FliG, to drive the rotation. In this study, we tried to overproduce the cytoplasmic loop of PomA (PomA-Loop), but it was insoluble. Thus, we made a fusion protein with a small soluble tag (GB1) which allowed us to express and characterize the recombinant protein. The structure of the PomA-Loop seems to be very elongated or has a loose tertiary structure. When the PomA-Loop protein was produced in E. coli, a slight dominant effect was observed on motility. We conclude that the cytoplasmic loop alone retains a certain function.