The FlhA linker mediates flagellar protein export switching during flagellar assembly

Yumi Inoue,Tohru Minamino,Katsumi Imada,Mamoru Kida,Keiichi Namba,Norihiro Takekawa,Miki Kinoshita

doi:10.1038/s42003-021-02177-z

Abstract

The flagellar protein export apparatus switches substrate specificity from hook-type to filament-type upon hook assembly completion, thereby initiating filament assembly at the hook tip. The C-terminal cytoplasmic domain of FlhA (FlhAC) serves as a docking platform for flagellar chaperones in complex with their cognate filament-type substrates. Interactions of the flexible linker of FlhA (FlhAL) with its nearest FlhAC subunit in the FlhAC ring is required for the substrate specificity switching. To address how FlhAL brings the order to flagellar assembly, we analyzed the flhA(E351A/W354A/D356A) ΔflgM mutant and found that this triple mutation in FlhAL increased the secretion level of hook protein by 5-fold, thereby increasing hook length. The crystal structure of FlhAC(E351A/D356A) showed that FlhAL bound to the chaperone-binding site of its neighboring subunit. We propose that the interaction of FlhAL with the chaperon-binding site of FlhAC suppresses filament-type protein export and facilitates hook-type protein export during hook assembly.

Highlights

The flagellar protein export apparatus switches substrate specificity from hook-type to filament-type upon hook assembly completion, thereby initiating filament assembly at the hook tip
The flagellum of Salmonella enterica is a supramolecular motility machine consisting of the basal body, which acts as a bi-directional rotary motor, the hook, which functions as a universal joint, and the filament, which works as a helical propeller[1]
The flhA(E351A/D356A) and flhA(W354A) mutants produce the hook–basal body (HBB) without the filament attached[10]

Summary

Introduction

The flagellar protein export apparatus switches substrate specificity from hook-type to filament-type upon hook assembly completion, thereby initiating filament assembly at the hook tip. The C-terminal cytoplasmic domain of FlhA (FlhAC) serves as a docking platform for flagellar chaperones in complex with their cognate filament-type substrates. The FlhAC–FlhBC-docking platform provides binding sites for the cytoplasmic ATPase complex, flagellar export chaperones (FlgN, FliS, FliT), and export substrates to mediate hierarchical protein targeting and secretion[4]. The FlhAC–FlhBCdocking platform serves as an export switch to induce substrate specificity switching from rod-/hook-type proteins to filamenttype ones when the hook reaches its mature length of about 55 nm in Salmonella, thereby terminating hook assembly and initiating filament formation (Fig. 1)[8,9,10,11]

Methods

Results

Conclusion