FliK-Driven Conformational Rearrangements of FlhA and FlhB Are Required for Export Switching of the Flagellar Protein Export Apparatus.

FlhA and FlhB are transmembrane proteins of the flagellar type III protein export apparatus, and their C-terminal cytoplasmic domains (FlhA and FlhB) coordinate flagellar protein export with assembly. FlhB undergoes autocleavage between Asn-269 and Pro-270 in a well-conserved NPTH loop located between FlhB and FlhB polypeptides and interacts with the C-terminal domain of the FliK ruler when the length of the hook has reached about 55 nm in As a result, the flagellar protein export apparatus switches its substrate specificity, thereby terminating hook assembly and initiating filament assembly. The mechanism of export switching remains unclear. Here, we report the role of FlhB cleavage in the switching mechanism. Photo-cross-linking experiments revealed that the () and () mutations did not affect the binding affinity of FlhB for FliK. Genetic analysis of the () mutant revealed that the P270A mutation affects a FliK-dependent conformational change of FlhB, thereby inhibiting the substrate specificity switching. The () mutation in FlhA suppressed the () mutation, suggesting that an interaction between FlhB and FlhA is critical for the export switching. We propose that the interaction between FliK and a cleaved form of FlhB promotes a conformational change in FlhB responsible for the termination of hook-type protein export and a structural remodeling of the FlhA ring responsible for the initiation of filament-type protein export. The flagellar type III protein export apparatus coordinates protein export with assembly, which allows the flagellum to be efficiently built at the cell surface. Hook completion is an important morphological checkpoint for the sequential flagellar assembly process. The protein export apparatus switches its substrate specificity from the hook protein to the filament protein upon hook completion. FliK, FlhB, and FlhA are involved in the export-switching process, but the mechanism remains a mystery. By analyzing a slow-cleaving () mutant, we provide evidence that an interaction between FliK and FlhB induces conformational rearrangements in FlhB, followed by a structural remodeling of the FlhA ring structure that terminates hook assembly and initiates filament formation.