Premium
Substrate specificity of type III flagellar protein export in Salmonella is controlled by subdomain interactions in FlhB
Author(s) -
Fraser Gillian M.,
Hirano Takanori,
Ferris Hedda U.,
Devgan Lara L.,
Kihara May,
Macnab Robert M.
Publication year - 2003
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1046/j.1365-2958.2003.03487.x
Subject(s) - flagellin , flagellum , biology , cleavage (geology) , cytoplasm , microbiology and biotechnology , protein filament , mutant , gene , genetics , paleontology , fracture (geology)
Summary 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 (FlhB C ) is cleaved between Asn‐269 and Pro‐270, defining two subdomains: FlhB CN and FlhB CC . 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 FlhB CN permitted FlhB cleavage but abolished function, whereas a deletion spanning Asn‐269 and Pro‐270 abolished both. The mutation N269A prevented cleavage at the FlhB CN –FlhB CC 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.