motormax forcemax speedmax power rotary rotary flagellar motor (8 units, e. coli) 2400 pn nm 95 pn...

Motor Max Force Max Speed Max Power

ROTARY

Flagellar motor (8 units, E. coli)

2400 pN nm

95 pN

300 Hz

35 m/s

2000 pN nm @ 150 Hz

1.9 x 106 pN nm /s

(Vibrio) 1700 Hz

220 m/s

(single unit) 300 pN nm

12 pN

300 Hz

35 m/s

250 pN nm @ 150 Hz

2.4 x 105 pN nm /s

F1 -ATPase 40 pN nm

40 pN

150 Hz

0.9 m/s

20 pN nm @ 75 Hz

9 x 103 pN nm /s

LINEAR

Myosin (single molecule in muscle or in vitro)

6 pN 10 m/s(speed of array, each

molecule mostly detached)

2 pN @ 10 /s x 20 nm

400 pN nm /s

Kinesin 5 pN 1 m/s 2.5 pN @ 0.5 m /s

1.25 x 103 pN nm /s

RNA polymerase

20 pN 0.01 m/s ~200 pN nm /s

Overview of lectures

• BIOLOGY was introduced by Judy Armitage’s

• BIOPHYSICS - Experimental Techniques to measure rotary molecular motors

Flagellar Motor

F1-ATPase

Single-molecule experiments on bacterial flagellar motors

Rotor

Stator

Continuous switch model

Switching

F Bai, RW. Branch, D Nicolau, TPilizota, BC Steel, PK Maini, RM Berry (2010)Conformational spread as a mechanism for cooperativity in the bacterial flagellar switchScience 327:685-689

Bacterial Chemotaxis

1-D Ising model of flagellar switch

movie

Tethered cells

Cell body rotates at ~ 10 Hz

1 m

Flagellum tethered to coverslip

Cell body

Coverslip in microscope

Frequency (Hz)

fast beads, small.mov

Work = torque x angle

Torque = d(work) / d(angle)

Low Reynolds number:Torque = viscous drag coefficient x angular velocity

Beads attached to the motor

Finite, variable switch times

Switch times distribution predicted by model

… further detailed tests of model

C / Co

Resurrection

resurrection

steady-state expression

One motor can contain at least 11 stators

1 m bead

0.3 m bead

Torque-versus speed

Stepping rotation

Speed control for step detection using sodium-driven chimaera

pmf or smf = Vm + kT/e ln (Cin/Cout)

Low numbers of stators:

Low-level induction of stator proteins

De-energization also affects stator number

Slow rotation with (probably) one stator unit

Back-focal plane detection Fluorescence detection

Real speed 30x slower

26 steps per revolution

Kinesin, myosin II, Myosin V, F1-ATPase:

Step size is set by the track

by energy conservation based on full energization and high loads, one proton gives a max step size of ~10 degrees. Maybe there are 2 ions per step?

34-fold model refines C-ring : 25-fold model refines M ring (& C-inner)Thomas et al 2006

One ATP per step

ATP

ADP+ Pi

10nm

F1

FO

H+ or Na+

Single-molecule experiments on ATP-synthase

F1

Biotin-avidin link to rotating handle(actin filament, beads)

His-tag link to surface

0.5 micron beadsRotated by F1

Fluorescent actin filamentRotated by F1

(movie: Wolfgang Junge)

High [ATP] : no wait before 90° step

Medium [ATP] : ~ms before 90° step

Low [ATP] : long wait before 90° step

(ms)

All [ATP] : ~ms before 30° step

Low [ATP] : - 90° step rate-limiting- exponential distribution- single step

High [ATP] : - 30° step rate-limiting- peaked distribution- double (or more) step