The conserved charged residues of the C-terminal region of FliG, a rotor component of the Na+-driven flagellar motor.
FliG is an essential component of the flagellar motor and functions in flagellar assembly, torque generation and regulation of the direction of flagellar rotation. The five charged residues important for the rotation of the flagellar motor were identified in Escherichiacoli FliG (FliG(E)). These residues are clustered in the C terminus and are all conserved in FliG(V) of the Na(+)-driven motor of Vibrioalginolyticus (Lys284, Arg301, Asp308, Asp309 and Arg317). To investigate the roles of these charged residues in the Na(+)-driven motor, we cloned the VibriofliG gene and introduced single or multiple substitutions into the corresponding positions in FliG(V). FliG(V) with double Ala replacements in all possible combinations at these five conserved positions still retained significant motile ability, although some of the mutations completely eliminated the function of FliG(E). All of the triple mutants constructed in this study also remained motile. These results suggest that the important charged residues may be located in different places and the conserved charged residues are not so important for the Na(+)-driven flagellar motor of Vibrio. The chimeric FliG protein (FliG(VE)), composed of the N-terminal domain from V.alginolyticus and the C-terminal domain from E.coli, functions in Vibrio cells. The mutations of the charge residues of the C-terminal region in FliG(VE) affected swarming ability as in E.coli. Both the FliG(V) and the FliG(VE) proteins with the triple mutation were more susceptible to proteolysis than proteins without the mutation, suggesting that their conformations were altered.
Yorimitsu
Tomohiro
T
Division of Biological Science, Graduate School of Science, Nagoya University, Chikusa-ku, 464-8602, Nagoya, Japan.
Mimaki
Atsushi
A
Yakushi
Toshiharu
T
Homma
Michio
M
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D017124:Conserved Sequence; D010450:Endopeptidases; D004926:Escherichia coli / Q000737:chemistry* / Q000378:metabolism; D005407:Flagella / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics*; D011485:Protein Binding; D017386:Sequence Homology, Amino Acid; D013347:Subcellular Fractions; D013550:Swimming; D014733:Vibrio / Q000378:metabolism
D001426:Bacterial Proteins; C075661:Flif protein, Bacteria; C075660:Flig protein, Bacteria; D008164:Luminescent Proteins; D008565:Membrane Proteins; D049452:Green Fluorescent Proteins; D010450:Endopeptidases