F1 - ATPase: A Molecular Transducer ofChemical and Mechanical Energies

Kazuhiko Kinosita, Jr.

Department of Physics, School of Science and EngineeringWaseda University

http://www.k2.phys.waseda.ac.jp

Why Do We Eat, and Why Do We Breathe?

400 µM ATP

A Two-Foot Linear Molecular Motor That Transports Nutrients in the Brain

White: closer to youBlack: away from you 4x fast replay

M. Y. Ali, S. Uemura, S. Ishiwata

Myosin V

Actin filament

Literature quotation

Major Players

Watching How Myosin May Walk

K. Shiroguchi

36 nm

ATP (Adenosine Triphosphate):A Universal Currency of Energy in the Biological World

ATP Inorganic phosphate (Pi) ADP

+Free energy drop: DG = DG0 + kBT ln[ADP][Pi]/[ATP] DG0 = -50 pN·nm (@pH 7) DG = -(80~100) pN·nm in cells = -(20~25) kBT

Why do We Eat, and Why do We Breathe?To obtain energy (= to synthesize ATP)

by slowly burning the food we ingest with oxygen we inhale.

A mitochondrionP. D. Mitchell

ATP SynthaseProton-driven and ATP- driven motors connected by a common shaft (g).

ATP synthesis/hydrolysis on three bs.

Isolated F1 only hydrolyzes ATP; hence it is called F1 -ATPase

g

g

P. D. Boyer F. Oosawa & S. Hayashi J. E. Walker & colleagues

A Protein Molecule Is Madeby Lock-and-Key Mechanisms

+

+

–

–

N-H •••• O=C

N •••• H-O

+

+

–

–

N-H •••• O=C

N •••• H-O

F1-ATPase:A Rotary Motor Made of a Single Molecule

H. Noji, R. Yasuda & M. Yoshida

Streptavidin

Actin filament

10 nm

γ

α β

Physiological Function of F1 Is to Synthesize ATPthrough (Forced) Clockwise Rotation

Isolated F1 : CCW rotation powered by ATP hydrolysis in the three b subunits

F1 in vivo: CW rotation driven by proton flow through the Fo motor　　　　　　　↓　ATP synthesis in the three b subunits

γ

α β

FO

Attempt at Mechanical Synthesis of ATP

H. Itoh

Luciferin/Luciferase

A. Einstein

Manipulating Many Motors Simultaneously

H. Itoh

Magnet rotated at 10 Hz

Hiro Itoh @Hamamatsu Photonics Co., Ltd.

Reverse Rotation Led to ATP Synthesis!!

hyd

10 6 3

no rotation

Ph

oto

ns

Det

ecte

d

5 min at indicated rates (Hz)

First chemical synthesis driven by force

2 ATP / turn

Chemical Synthesis by Force (Torque)!

Free energy of ATP hydrolysis:

DG = DG0 + kBT ln[ADP][Pi]/[ATP] DG0 = _58 pN·nm (pH7.6) DG = _ (80~100) pN·nm (in cells) = _ (20~25) kBT

DG ~ _ 30 pN·nm （this experiment） ~ _ 7 kBT [ADP]=200mM,[Pi]=10mM,[ATP]~1nM

Hiro Itoh @Hamamatsu Photonics Co., Ltd.

A Force (Torque) Applied to One PointCan Reverse the Operation of F1 !!

g - Dictator Mechanism: The g angle dictates how chemical reactions proceed in b’s. In hydrolysis, three b’s cooperate through the g angle.

γ

β

How Can a Molecular Machine Convert ChemicalEnergy of ATP Hydrolysis into Mechanical Work?

ATP Inorganic phosphate (Pi) ADP

+Free energy drop: DG = DG0 + kBT ln[ADP][Pi]/[ATP] DG0 = -50 pN·nm (@pH 7) DG = -(80~100) pN·nm in cells = -(20~25) kBT

Liberation of free energy == Reaction goes toward right

Stepwise Rotation of F1

[ATP] = 20 nM

R. Yasuda & H. Noji

Watch!!

ab (catalytic)

g (rotor)