Distinct Roles of Highly Conserved Charged Residues at the MotA-FliG Interface in Bacterial Flagellar Motor Rotation
Distinct Roles of Highly Conserved Charged Residues at the MotA-FliG Interface in Bacterial Flagellar Motor Rotation
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DOI：
10.1128/jb.01971-12
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发表时间：
2013-02-01
影响因子：
3.2
通讯作者：
Minamino, Tohru
中科院分区：
文献类型：
--
作者：
Morimoto, Yusuke V.;Nakamura, Shuichi;Minamino, Tohru
定子蛋白MoTA和转子蛋白FliG之间的静电相互作用对于细菌鞭毛马达的旋转是重要的。Mota的Arg90和Glu98不仅是产生扭矩所必需的，也是转子周围的定子组装所必需的，但它们的实际作用尚不清楚。在这里，我们分析了Mota-FliG界面上重要的带电残基在电机性能中的作用。约75%的MOTA(R90E)细胞和45%的MOTA(E98K)细胞没有显示绿色荧光蛋白(GFP)-MOTB的荧光点，这表明定子组装在转子周围的效率降低。FliG(D289K)和FliG(R281V)突变分别恢复了mota(R90E)和mota(E98K)突变体的运动能力，也显示出GFP-MOTB斑点的减少和强度。FliG(D289K)突变显著恢复了MOTA(R90E)突变体中GFP-MOTB对运动的定位，而FliG(R281V)突变不能恢复MOTA(E98K)突变体中GFP-MOTB的定位。这些结果表明，Mota-Arg90-FliG-Asp289相互作用对于定子在转子周围的正确定位至关重要，而Mota-Glu98-FliG-Arg281相互作用对扭矩产生更重要。
Electrostatic interactions between the stator protein MotA and the rotor protein FliG are important for bacterial flagellar motor rotation. Arg90 and Glu98 of MotA are required not only for torque generation but also for stator assembly around the rotor, but their actual roles remain unknown. Here we analyzed the roles of functionally important charged residues at the MotA-FliG interface in motor performance. About 75% of the motA(R90E) cells and 45% of the motA(E98K) cells showed no fluorescent spots of green fluorescent protein (GFP)-MotB, indicating reduced efficiency of stator assembly around the rotor. The FliG(D289K) and FliG(R281V) mutations, which restore the motility of the motA(R90E) and motA(E98K) mutants, respectively, showed reduced numbers and intensity of GFP-MotB spots as well. The FliG(D289K) mutation significantly recovered the localization of GFP-MotB to the motor in the motA(R90E) mutant, whereas the FliG(R281V) mutation did not recover the GFP-MotB localization in the motA(E98K) mutant. These results suggest that the MotA-Arg90-FliG-Asp289 interaction is critical for the proper positioning of the stators around the rotor, whereas the MotA-Glu98-FliG-Arg281 interaction is more important for torque generation.