Weak Interactions between Salmonella enterica FlhB and Other Flagellar Export Apparatus Proteins Govern Type III Secretion Dynamics.
Weak Interactions between Salmonella enterica FlhB and Other Flagellar Export Apparatus Proteins Govern Type III Secretion Dynamics.
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DOI：
10.1371/journal.pone.0134884
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Namba K
中科院分区：
文献类型：
--
作者：
McMurry JL;Minamino T;Furukawa Y;Francis JW;Hill SA;Helms KA;Namba K
细菌鞭毛含有其自己的 III 型分泌装置，可协调蛋白质输出与远端的组装。虽然输出装置蛋白之间的许多相互作用已被报道，但很少有人对控制输出机制的差异亲和力和动态关系进行检查。 FlhB 是一种完整的膜蛋白，在钩子完成时发生的输出和底物特异性转换中发挥着关键作用。本文报道的是 FlhB (FlhBC) 的胞质结构域与其他输出装置蛋白（包括 FliK、FlhAC 和 FliI）之间相互作用的定量表征。 FliK 和 FlhAC 以微摩尔亲和力结合。在没有 ATP 的情况下，FliI 结合的 KD 为 84 nM。 ATP 诱导的 FliI 寡聚化引起动力学变化，刺激快速结合、快速关闭结合并降低亲和力。在增溶、非变性条件下纯化的全长 FlhB 通过其跨膜结构域和稳定结合的 FliH 形成稳定的二聚体。总之，目前的结果支持了先前假设的 FlhB 的核心作用，并阐明了 III 型分泌中蛋白质-蛋白质相互作用的动态。
The bacterial flagellum contains its own type III secretion apparatus that coordinates protein export with assembly at the distal end. While many interactions among export apparatus proteins have been reported, few have been examined with respect to the differential affinities and dynamic relationships that must govern the mechanism of export. FlhB, an integral membrane protein, plays critical roles in both export and the substrate specificity switching that occurs upon hook completion. Reported herein is the quantitative characterization of interactions between the cytoplasmic domain of FlhB (FlhBC) and other export apparatus proteins including FliK, FlhAC and FliI. FliK and FlhAC bound with micromolar affinity. KD for FliI binding in the absence of ATP was 84 nM. ATP-induced oligomerization of FliI induced kinetic changes, stimulating fast-on, fast-off binding and lowering affinity. Full length FlhB purified under solubilizing, nondenaturing conditions formed a stable dimer via its transmembrane domain and stably bound FliH. Together, the present results support the previously hypothesized central role of FlhB and elucidate the dynamics of protein-protein interactions in type III secretion.