+,)+,)*-mno  pp

3////2IYaKBP3///////
.('Vf^Hkkkkkkq
SeCLbkkkkk
ZBFekkkkkk#IYaKBP .'
VYK_EDhkkk .'_COkkkkkkk 
BXkkkkkkkk
Sj\jNlkkkk!
]Vkkkkkkkk".
DjWCkkkkkk!
VCciJMkkkk
dgkkkkkkkJaFkkkkkkk _T[fjMR# :IYaKBP

 5:4>:

09="<@6`Gi:clQl1



!$&%7?;A8:#;AMOUaSmoothimmingsw         
Tumbling

Smoothimmingsw+

"# "# !$ &'+%,376;0-/2<'+&81:

Flagellin
HAP1HAP2(FliD)

Flagellin (FliC)

Region I
3

FlgBCFG

3flaF

2flgBCDEGHIJ

flgKL flaC flaD

flaE

flaB

3

flaLM

flaAGHIJK

2



fliEFGHIKFLMNOPQR flhB2

flhAFG fliA cheYZAB orf1 orf2 cheW orf3FliA,FliHI
FlgI

2Region II

L-ring(FlgH)
P-ring(FlgI)
Rod(FlgBCFG)
MS-ring(FliF)
C-ring(FliGMN)
FlgD

cheVR



HAP3(FlgK)
HAP1(FlgL)
FlgE

FlgH

33

flgNM flgA

FlgE

            

HAP2

HAP3

Region III

FliFRegion IVRegion V

32,32

pomAB

motX

motY

FliGMN   

FliAHI

459.

'+

(multi-Pof)&
WT

WT

'+&%*()



KK148
KK148NMB155

30℃, 5hr

NMB1551μm#

()&()&'+

FlhF0FlhG1684: 4:209 .80%# 
C/2# ,flhF0flhG1

flhF

flhA

flhGfliA
1.0 kbp

FlhF2 FlhF

FlhF"

FlhG"

FlhG

cheY

FlhG (FleN)2 

Putative GTPase
Similar to E. coli FtsY (SRP recepter)
Involved in flagellar placement(in Pseudomonas putida)
Overproduction causes hyperflagellation(in Pseudomonas putida)

Putative ATPase
Similar to E. coli MinD
(cell division inhibitor)
Overproduction prevents flagellation(in Pseudomonas aeruginosa)
Depletion causes hyperflagellation(in Pseudomonas aeruginosa)

Vibrio alginolyticus-7%flhF 0%flhG # 9;@BA%

11

1

4:ΔflhF

VIO5(wt, Pof+Laf-) 1

KK148(flhG mutant)

ΔflhFG
Bar; 1 μm

FlhF34:  15$!?

FlhF 0%FlhG1684:202
Vibrio alginolyticus Cell>=?

FlhF localization

GTPase activity?

FlhF>B<B2 
>B<B2!

FlhF

directly?

FlhG

Flagellation
Overexpression of FlhG

Depletion of FlhG



Ｎａ+駆動モーター特異的なMotXとMotYはモーター基部にリング構造を作る

MotX
MotY

PomA, PomB (MotA, MotB)
No crystals( V. alginolyticus )( E. coli )FliG-C2.2Å structure
Lloyd et al, 1999

FliG-middleFliN

3.4Å diffraction
Molecular replacement
Brown et al, 2005

FliM-middle

FliG-N

2.8Å structure
Brown et al, 2002

LP ring
LP ringT-ring2.0Å structure
4.0Å diffraction
Park et al, 2006

proteins

MS ring

MS ring(Terashima et al, Mol. Microbiol. 2006)

Bar; 50nm

V. alginolyticus basal body contained an additional structure, T-ring,and T-ring is composed of MotX and MotY.

* 

MotX と MotYの存在部位 ! " 
KK148(motX+motY+)

TH2(ΔmotY)

TH1(ΔmotX)

Previous model

TH3(ΔmotX ΔmotY)

OM MotY

New model from 2006

Hook

MotX

L ringP ring

PG

MotY & MotX (T-ring)
Pom B

Pom BIMPom A

MS ring

Pom AC ring    !$013'"   #$26+6"%'5   013'"#$!#$"*%-,

H+-driven
Lateral flagellaHigh viscosityH+#$26+6"

 $#""#! !&.4(+5/)

H+ -driven
E.coliNa + -driven
V.alginolyticus

unction

MotA
MotB

PomA
PomB
MotX
MotY

ChannelChannel/Stator
T-ringT-ringLow viscosity

Na+Na+-drivenPolar flagellum,-*,-*+/"& ,'-(% Intermoleculardimerization
PomA (27.2 kD)5=:VKFP_ 3<P`U]" "

PomB (35.5 kD)
PBD

P172

C

D24
N
D148
R88
C
R89
E99
E96 E97

BE_JZO]<24?(BE_$N

Critical site
of sodium binding(PomA)4(PomB)2 
heterohexamer complex

P16

Phenamil resistance
(D148Y or P16S)

Rotor-Stator interaction
(Sato & Homma, J. Biol. Chem. 2000)

T^L_;NL\CW)

=QBS\KMFX[Y`I`=

KcsA....prokaryotic K+ channel
similar to Shaker type(Science, Doyle et al., 1998)

BE_$RA]I`,'=\_H<%69& :0
69K+ BE_;608= =DO]G`@ 7?1


QBS\KMFX[Y`I`<>?(a)
H+ typeMS ring

Na+ type

PGB

MotA

MotB

MS ring

FliGE

in E. coli

FliM/N

0 mM NaCl

100 mM NaCl

M254 (motB mutant) expressing MotA/MotB (WT)



malfunction

PGB

MotA

PomB

MS ring

PomA
PomA

PGB

MotB

FliG Ein E. coli!/NaCl0$0 mM NaClPo B

in E. coli

FliM/N

FliGE

/#*+0.)!/0$PomA(d)malfunction

FliM/NPGB

FliGE(c)MS ring!=#(b)FliM/Nin E. coli#*+/E. coli0.) 
100 mM NaCl

M254 (motB mutant) expressing PomA/PotB7.

  E. coli 

@3BG<>7DFEI;I+ K
?=G4C36H(!     0IJ
#5H9H'J

PomA/PotB7

PomA/PotB7

WT-MotA/MotB

WT-MotA/MotB     
#5H9H1#&(

36HAF<8:($.,2EI;I1-.")( .%J //*EI;IEI;I+
EI;I+.1)1 #

モーター回転計測システム





Power/generator = 1.75 x 105 pN nm s-1( x 70 of kinesin, x 400 of myosin, x 450 of F1-ATPase)計測系

遅い領域の 角度 vs 時間

Quadrant Photodiode200 nm fluorescent bead
500 nmbead1064 nm laser

High speed CCD camera(Ryu, Berry and Berg (2000) Nature)

ビーズの重心位置はnm, msの精度で計測することができる。

26 (+*-/$ "!%#)0,&FliG#"!%ステップサイズの分布/0'360º / 13.7 º≈ 26 steps/rev

1,400 revolution from 9 cellsFliG
~26(Suzuki et al. 2004)

FliM & FliN ~34  (Thomas et al. 1999) The main steps size is ~13.7 º.
It corresponds to ~26 steps / revolution.

45245236

 o8]_dkXkC:;FGlrm FoWjYdk[h9       
1  1   8   
8 'HL.ekXkFGlrm]_dk[h9               
8&'FHLekXkG /E *>JFr           
8HLekXkF *FN^hQkGr ShTkV        0  MOj8#, FN^hQkGr ShTkV        0l MOj
8 'HLekXkGBA:JFr 8XjaR)      1  %)
8HLKI$=?JE(D-Gr9 on  pq  qn  onn1  pnn  qnn
8 'FHL"!CekXkK@D<+F Gr
 iZ\  UgMj[8  `Zc8  PfZc8  bZR

7