Division of Biological Science, Graduate School of Science, Nagoya University, Furo-cho, Chikusa-Ku, Nagoya 464-8602, Japan.
Department of Macromolecular Science, Graduate School of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka 560-0043, Japan.
Biomolecules. 2020 Jan 11;10(1):126. doi: 10.3390/biom10010126.
The bacterial flagellum is a filamentous organelle extending from the cell surface. The axial structure of the flagellum consists of the rod, hook, junction, filament, and cap. The axial structure is formed by axial component proteins exported via a specific protein export apparatus in a well-regulated manner. Although previous studies have revealed the outline of the flagellar construction process, the mechanism of axial structure formation, including axial protein export, is still obscure due to difficulties in direct observation of protein export and assembly in vivo. We recently developed an in vitro flagellar protein transport assay system using inverted membrane vesicles (IMVs) and succeeded in reproducing the early stage of flagellar assembly. However, the late stage of the flagellar formation process remained to be examined in the IMVs. In this study, we showed that the filament-type proteins are transported into the IMVs to produce the filament on the hook inside the IMVs. Furthermore, we provide direct evidence that coordinated flagellar protein export and assembly can occur at the post-translational level. These results indicate that the ordered construction of the entire flagellar structure can be regulated by only the interactions between the protein export apparatus, the export substrate proteins, and their cognate chaperones.
细菌鞭毛是一种从细胞表面伸出的丝状细胞器。鞭毛的轴向结构由杆、钩、连接、丝和帽组成。轴向结构由通过特定的蛋白输出装置以一种精确调控的方式输出的轴向组成蛋白形成。尽管先前的研究已经揭示了鞭毛结构形成的大致过程，但由于在体内直接观察蛋白输出和组装的困难，轴向结构形成的机制，包括轴向蛋白输出，仍然不清楚。我们最近开发了一种使用反转膜囊泡（IMVs）的体外鞭毛蛋白转运测定系统，并成功地复制了鞭毛组装的早期阶段。然而，鞭毛形成过程的后期阶段仍需在 IMVs 中进行检查。在这项研究中，我们表明，丝型蛋白被转运到 IMVs 中，在 IMVs 内的钩上产生丝。此外，我们提供了直接的证据表明，协调的鞭毛蛋白输出和组装可以在翻译后水平发生。这些结果表明，整个鞭毛结构的有序构建可以仅通过蛋白输出装置、输出底物蛋白及其同源伴侣之间的相互作用来调节。
