FliH and FliI ensure efficient energy coupling of flagellar type III protein export in Salmonella.
FliH and FliI ensure efficient energy coupling of flagellar type III protein export in Salmonella.
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DOI：
10.1002/mbo3.340
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发表时间：
2016-06
期刊：
影响因子：
3.4
通讯作者：
Namba K
中科院分区：
文献类型：
--
作者：
Minamino T;Kinoshita M;Inoue Y;Morimoto YV;Ihara K;Koya S;Hara N;Nishioka N;Kojima S;Homma M;Namba K
为了构建细菌鞭毛，鞭毛蛋白通过其特定的输出装置从细胞质输出到生长的鞭毛结构的远端。鞭毛输出装置由跨膜（TM）输出门复合物和由FliH、FliI和FliJ组成的细胞质ATP酶复合物组成。FlhA是一种TM输出门蛋白，在蛋白质转运的能量偶联中起重要作用。然而，能量耦合机制仍然未知。在这里，我们进行了交叉互补测定，以测量输出装置的能量转导系统对遗传扰动的鲁棒性。弧菌FlhA恢复了沙门氏菌ΔflhA突变体的运动性，但不能恢复ΔfliH‐fliI flhB（P28 T）ΔflhA突变体的运动性。flgM突变显著增加鞭毛基因表达水平，使弧菌FlhA在ΔfliH‐fliI flhB（P28 T）ΔflhA突变体中发挥其输出活性。拉下试验显示，弧菌FlhA对FliJ和FlgN-FlgK伴侣-底物复合物的结合亲和力远低于沙门氏菌FlhA的结合亲和力。这些表明，弧菌FlhA需要FliH和FliI的支持，以有效和适当地与FliJ和FlgN-FlgK复合物相互作用。我们建议，FliH和FliI确保强大的和有效的能量耦合的蛋白质出口鞭毛组装过程中。
For construction of the bacterial flagellum, flagellar proteins are exported via its specific export apparatus from the cytoplasm to the distal end of the growing flagellar structure. The flagellar export apparatus consists of a transmembrane (TM) export gate complex and a cytoplasmic ATPase complex consisting of FliH, FliI, and FliJ. FlhA is a TM export gate protein and plays important roles in energy coupling of protein translocation. However, the energy coupling mechanism remains unknown. Here, we performed a cross‐complementation assay to measure robustness of the energy transduction system of the export apparatus against genetic perturbations. Vibrio FlhA restored motility of a Salmonella ΔflhA mutant but not that of a ΔfliH‐fliI flhB(P28T) ΔflhA mutant. The flgM mutations significantly increased flagellar gene expression levels, allowing Vibrio FlhA to exert its export activity in the ΔfliH‐fliI flhB(P28T) ΔflhA mutant. Pull‐down assays revealed that the binding affinities of Vibrio FlhA for FliJ and the FlgN–FlgK chaperone–substrate complex were much lower than those of Salmonella FlhA. These suggest that Vibrio FlhA requires the support of FliH and FliI to efficiently and properly interact with FliJ and the FlgN–FlgK complex. We propose that FliH and FliI ensure robust and efficient energy coupling of protein export during flagellar assembly.