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Mol Microbiol. 2003 May;48(4):1043-57.

Substrate specificity of type III flagellar protein export in Salmonella is controlled by subdomain interactions in FlhB.

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Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520-8114, USA.


FlhB, an integral membrane protein, gates the type III flagellar export pathway of Salmonella. It permits export of rod/hook-type proteins before hook completion, whereupon it switches specificity to recognize filament-type proteins. The cytoplasmic C-terminal domain of FlhB (FlhBC) is cleaved between Asn-269 and Pro-270, defining two subdomains: FlhBCN and FlhBCC. Here, we show that subdomain interactions and cleavage within FlhB are central to substrate-specificity switching. We found that deletions between residues 216 and 240 of FlhBCN permitted FlhB cleavage but abolished function, whereas a deletion spanning Asn-269 and Pro-270 abolished both. The mutation N269A prevented cleavage at the FlhBCN-FlhBCC boundary. Cells producing FlhB(N269A) exported the same amounts of hook-capping protein as cells producing wild-type FlhB. However, they exported no flagellin, even when the fliC gene was being expressed from a foreign promoter to circumvent regulation of expression by FlgM, which is itself a filament-type substrate. Electron microscopy revealed that these cells assembled polyhook structures lacking filaments. Thus, FlhB(N269A) is locked in a conformation specific for rod/hook-type substrates. With FlhB(P270A), cleavage was reduced but not abolished, and cells producing this protein were weakly motile, exported reduced amounts of flagellin and assembled polyhook filaments.

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