Structure of the cytoplasmic domain of FlhA and implication for flagellar type III protein export.
FlhA is the largest integral membrane component of the flagellar type III protein export apparatus of Salmonella and is composed of an N-terminal transmembrane domain (FlhA(TM)) and a C-terminal cytoplasmic domain (FlhA(C)). FlhA(C) is thought to form a platform of the export gate for the soluble components to bind to for efficient delivery of export substrates to the gate. Here, we report a structure of FlhA(C) at 2.8 A resolution. FlhA(C) consists of four subdomains (A(C)D1, A(C)D2, A(C)D3 and A(C)D4) and a linker connecting FlhA(C) to FlhA(TM). The sites of temperature-sensitive (ts) mutations that impair protein export are distributed to all four domains, with half of them at subdomain interfaces. Analyses of the ts mutations and four suppressor mutations to the G368C ts mutation suggested that FlhA(C) changes its conformation for its function. Molecular dynamics simulation demonstrated an open-close motion with a 5-10 ns oscillation in the distance between A(C)D2 and A(C)D4. These results along with further mutation analyses suggest that a dynamic domain motion of FlhA(C) is essential for protein export.
Saijo-Hamano
Yumiko
Y
Dynamic NanoMachine Project, International Cooperative Research Project, Japan Science and Technology Agency, Suita, Osaka, Japan.
Imada
Katsumi
K
Minamino
Tohru
T
Kihara
May
M
Shimada
Masafumi
M
Kitao
Akio
A
Namba
Keiichi
K
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D018360:Crystallography, X-Ray; D005408:Flagellin / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism*; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012486:Salmonella typhimurium / Q000737:chemistry* / Q000378:metabolism*; D013489:Suppression, Genetic; D013696:Temperature
D001426:Bacterial Proteins; C080688:FlhA protein, Bacteria; D008565:Membrane Proteins; D005408:Flagellin