Assembly and stoichiometry of FliF and FlhA in Salmonella flagellar basal body.
The bacterial flagellar export apparatus is required for the construction of the bacterial flagella beyond the cytoplasmic membrane. The membrane-embedded part of the export apparatus, which consists of FlhA, FlhB, FliO, FliP, FliQ and FliR, is located in the central pore of the MS ring formed by 26 copies of FliF. The C-terminal cytoplasmic domain of FlhA is located in the centre of the cavity within the C ring made of FliG, FliM and FliN. FlhA interacts with FliF, but its assembly mechanism remains unclear. Here, we fused yellow fluorescent protein (YFP) and cyan fluorescent protein (CFP) to the C-termini of FliF and FlhA and investigated their subcellular localization by fluorescence microscopy. The punctate pattern of FliF-YFP localization required FliG but neither FliM, FliN, FlhA, FlhB, FliO, FliP, FliQ nor FliR. In contrast, FlhA-CFP localization required FliF, FliG, FliO, FliP, FliQ and FliR. The number of FlhA-YFP molecules associated with the MS ring was estimated to be about nine. We suggest that FlhA assembles into the export gate along with other membrane components during the MS ring complex formation in a co-ordinated manner.
Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita, Osaka, 565-0871, Japan; Quantitative Biology Center, RIKEN, 6-2-3 Furuedai, Suita, Osaka, 565-0874, Japan.
Yusuke V
YV
Morimoto
Mariko
M
Ito
Koichi D
KD
Hiraoka
Yong-Suk
YS
Che
Fan
F
Bai
Nobunori
N
Kami-Ike
Keiichi
K
Namba
Tohru
T
Minamino
D001426:Bacterial Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D064652:Basal Bodies / Q000737:chemistry* / Q000378:metabolism*; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D008164:Luminescent Proteins / Q000032:analysis / Q000235:genetics; D008565:Membrane Proteins / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000235:genetics; D012475:Salmonella / Q000737:chemistry* / Q000378:metabolism*
D001426:Bacterial Proteins; C485182:Cyan Fluorescent Protein; C080688:FlhA protein, Bacteria; C075661:Flif protein, Bacteria; D008164:Luminescent Proteins; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; C054562:yellow fluorescent protein, Bacteria; D049452:Green Fluorescent Proteins