title: Interaction of a bacterial flagellar chaperone FlgN with FlhA is required for efficient export of its cognate substrates.
abstract: FlgN chaperone acts as a bodyguard to protect its cognate substrates, FlgK and FlgL, from proteolysis in the cytoplasm. Docking of the FlgN-FlgK complex with the FliI ATPase of the flagellar type III export apparatus is key to the protein export process. However, a ΔfliH-fliI flhB(P28T) mutant forms some flagella even in the absence of FliH and FliI, raising the question of how FlgN promotes the export of its cognate substrates. Here, we report that the interaction of FlgN with an integral membrane export protein, FlhA, is directly involved in efficient protein export. A ΔfliH-fliI flhB(P28T) ΔflgN mutant caused extragenic suppressor mutations in the C-terminal domain of FlhA (FlhA(C) ). Pull-down assays using GST affinity chromatography showed an interaction between FlgN and FlhA(C) . The FlgN-FlgK complex bound to FlhA(C)  and FliJ to form the FlgN-FlgK-FliJ-FlhA(C)  complex. The FlgN-FlhA(C)  interaction was enhanced by FlgK but not by FliJ. FlgN120 missing the last 20 residues still bound to FlgK and FliJ but not to FlhA(C) . A highly conserved Tyr-122 residue was required for the interaction with FlhA(C) . These results suggest that FlgN efficiently transfers FlgK/L subunits to FlhA(C)  to promote their export.
authors.affiliation: Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan. tohru@fbs.osaka-u.ac.jp
authors.forename: Tohru
authors.initials: T
authors.lastname: Minamino
authors.forename: Miki
authors.initials: M
authors.lastname: Kinoshita
authors.forename: Noritaka
authors.initials: N
authors.lastname: Hara
authors.forename: Shiori
authors.initials: S
authors.lastname: Takeuchi
authors.forename: Akira
authors.initials: A
authors.lastname: Hida
authors.forename: Satomi
authors.initials: S
authors.lastname: Koya
authors.forename: Helen
authors.initials: H
authors.lastname: Glenwright
authors.forename: Katsumi
authors.initials: K
authors.lastname: Imada
authors.forename: Phillip D
authors.initials: PD
authors.lastname: Aldridge
authors.forename: Keiichi
authors.initials: K
authors.lastname: Namba
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D008954:Models, Biological; D008958:Models, Molecular; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D025941:Protein Interaction Mapping; D021381:Protein Transport; D013489:Suppression, Genetic
chemical_list: D001426:Bacterial Proteins; C087498:FlgN protein, Bacteria; C080688:FlhA protein, Bacteria; D008565:Membrane Proteins; D050505:Mutant Proteins