Novel insights into the mechanism of well-ordered assembly of bacterial flagellar proteins in Salmonella

Yumi Inoue,Tohru Minamino,Yusuke V Morimoto,Keiichi Namba

doi:10.1038/s41598-018-20209-3

Yumi Inoue, Tohru Minamino + Show 2 more

Open Access

https://doi.org/10.1038/s41598-018-20209-3

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Abstract

The FliI ATPase of the flagellar type III protein export apparatus forms the FliH2FliI complex along with its regulator FliH. The FliH2FliI complex is postulated to bring export substrates from the cytoplasm to the docking platform made of FlhA and FlhB although not essential for flagellar protein export. Here, to clarify the role of the FliH2FliI complex in flagellar assembly, we analysed the effect of FliH and FliI deletion on flagellar protein export and assembly. The hook length was not controlled properly in the ∆fliH-fliI flhB(P28T) mutant compared to wild-type cells, whose hook length is controlled to about 55 nm within 10% error. The FlhA(F459A) mutation increased the export level of the hook protein FlgE and the ruler protein FliK by about 10-fold and 3-fold, respectively, and improved the hook length control in the absence of FliH and FliI. However, the ∆fliH-fliI flhB(P28T) flhA(F459A) mutant did not produce flagellar filaments efficiently, and a large amount of flagellin monomers were leaked out into the culture media. Neither the hook length control nor flagellin leakage was affected by the FlhB(P28T) and FlhA(F459A) mutations. We will discuss a hierarchical protein export mechanism of the bacterial flagellum.

Highlights

The ruler protein FliK by about 10-fold and 3-fold, respectively, and improved the hook length control in the absence of FliH and FliI
The flagellar type III protein export apparatus switches its substrate specificity upon completion of hook assembly, thereby terminating hook assembly and initiating filament assembly (Fig. 1)
These suggest that a deletion of FliH and FliI considerably reduces the targeting efficiency of FlgE and FliK to the docking platform of the export gate complex made of FlhAC and FlhBC, and that the FlhA(F459A) mutation recovers their targeting efficiency, thereby restoring the hook length control

Summary

Introduction

The ruler protein FliK by about 10-fold and 3-fold, respectively, and improved the hook length control in the absence of FliH and FliI. The flagellar type III protein export apparatus is composed of a transmembrane export gate complex made of FlhA, FlhB, FliP, FliQ and FliR and a cytoplasmic ATPase complex consisting of FliH, FliI and FliJ4–6. The cytoplasmic ATPase ring complex of the export apparatus is formed by six copies of the FliH homo-dimer, six copies of the FliI ATPase and one copy of FliJ22–24 and is located at the flagellar base through interactions of the extreme N-terminal region of FliH with FlhA and a C ring protein FliN (Fig. 1)[25,26,27,28]. Since FliI-YFP exchanges between the flagellar basal body and the cytoplasmic pool, the FliH2FliI complex is postulated to bring export substrates and type III export chaperone–substrate complexes from the cytoplasm to the FlhAC-FlhBC platform[28]

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Conclusion