Conformational Changes

Conformational Changes in Proteins - VIII

ATPase is a protein complex composed of two parts usually labelled F₀ and F₁

F₀ is membrane-bound and acts as a proton pump

The pH difference across the membrane creates a chemical potential, m
m = RT ln (DpH)

F₁ is a soluble, multi-unit protein, a₃b₃gde

When connected to F₀ it uses the energy of the proton pump to synthesize ATP; when disconnected, it hydrolyzes ATP

The crystal structure of F₁ ATPase from cow heart mitochondria shows a mushroom shaped molecule:

ATPase F₁, Side View (1bmf)	ATPase F₁, Top View

The stem of the mushroom, or the axle of the wheel, is the g-subunit
In the complete F₀+F₁ protein, this stem inserts into the F₀ unit
The a and b subunits alternate to form the cap of the mushroom
The binding site is between pairs of a and b subunits, with most of the catalysis involving the b

ATPase F₁, Side View, ADP Bound (1h8e)	ATPase F₁, Top View, ADP Bound

Each of the three catalytic sites has three states:

L (oose), which binds ADP and P_i weakly and is not catalytic
T (ight), which binds ADP and P_i tightly, and converts them to ATP
O (pen), which releases ATP

The energy of the proton pump in F₀ is used to rotate the g subunit. (The interaction of this subunit with the others is not symmetrical.)

ATPase F₁, Side View, 50% Slab

The rotation induces conformational changes in a and b that rotate the sites among the three states:

The whole mechanism looks like this:

The term "mechanism" is exactly correct; the intact enzyme is a mechanical system that converts a chemical potential (pH difference) into a molecule (ATP) that releases energy upon hydrolysis, and can be used to power a large variety of other chemical processes.

Here are some movies of the process, from the Gogarten lab at UConn,showing the conformational changes in F₁:

Side View	Top View

Cutaway Side View

What's driving this? Details of the proton pump are not known; there is no structure. Here is a cartoon best guess from Oster's lab at Berkeley:

Mechanism of the Proton Pump

The negatively charged residues are aspartates, the lone positive residue is an arginine. Direction of rotation is outward from the left, inward to the right, or vice versa, depending on the pH gradient.

The motor must generate sufficient torque to produce about 20 kT per ATP molecule.

Given the current intense interest in nanotechnology and nanomachines, doing something cute with the F₁ system was inevitable.

Noji, et al [Nature, 1997, 386, 299] attached the F₁ unit to a solid support, and connected a fluorescent actin filament to the g subunit.

Adding ATP runs the reaction backward, hydrolyzing the ATP, and rotating the g shaft.
Rotation of the fluorescent tag was easily detected.

This page last modified 11:18 AM on Friday March 2nd, 2012.
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