A13 oxidative phosphorylation

What does the vectorial equation for electron transport represent?

It summarizes electron transport either with respect to one molecule of oxygen or one NADH.

What causes the mitochondrial matrix to become more negative?

Proton pumping from the matrix into the intermembrane space creates a charge difference—matrix (negative), intermembrane space (positive).

Why must the proton gradient be dissipated?

To prevent “pressure buildup” that stops proton pumping and halts electron transport and respiration.

What does the chemiosmotic theory state?

ATP synthesis is coupled to electron transfer through a proton gradient across the inner mitochondrial membrane, which stores energy used to make ATP.

What was the purpose of the first experiment (succinate + cyanide)?

To determine if electron transport is required for ATP synthesis.

What did the results of the first experiment show?

When cyanide inhibits electron transport (by blocking Complex IV), both oxygen consumption and ATP synthesis stop—showing coupling between the two processes.

What was tested in the second experiment with oligomycin and DNP?

Whether blocking ATP synthesis affects electron transport.

What does oligomycin inhibit?

ATP synthase, blocking ATP synthesis.

What is 2,4-DNP (dinitrophenol) and what does it do?

A weak acid uncoupler that dissipates the proton gradient by carrying protons across the membrane, allowing electron transport to continue without ATP synthesis.

Why does adding DNP restore electron transport after oligomycin?

DNP releases the built-up proton gradient, relieving back pressure and allowing electron flow to resume.

What drives ATP synthesis in mitochondria?

The proton gradient created by electron transport.

What happens when the proton gradient is experimentally increased by lowering pH outside the membrane?

ATP synthesis occurs even without electron transport, confirming that the gradient alone drives ATP production.

What are the two major domains of ATP synthase?

FO (membrane-embedded) and F₁ (matrix-facing catalytic head).

Which domain performs ATP synthesis?

The F₁ domain.

How many α and β subunits does the F₁ domain have?

Three α and three β subunits arranged alternately in dimers.

Where does catalysis occur in ATP synthase?

At the β subunits of the αβ dimers.

What role does the γ shaft play?

It rotates, inducing conformational changes in the αβ dimers to drive ATP formation and release.

What are the three conformational states of the αβ subunits?

Open (binds ADP + Pi), Tight (forms ATP), and Loose (releases ATP).

What drives rotation of the FO subunit?

The energy of protons moving down their gradient through the FO channel.

What does the C-ring of the FO complex do?

It rotates as protons bind and unbind, transmitting torque to the γ shaft of F₁.

How is the Ferris wheel analogy related to FO rotation?

Protons “load” onto one side of the C-ring (like people boarding) and “unload” on the other, causing rotation and energy transfer.

What residues are involved in proton loading/unloading?

Glutamate residues on the C-ring and an arginine residue on the A subunit.

How are ADP and ATP exchanged across the mitochondrial membrane?

Via adenine nucleotide translocase (imports ADP, exports ATP).

How is phosphate transported into the matrix?

Through phosphate translocase (a symporter that also brings in protons).

does not drive ATP synthesis

Why can’t NADH cross the mitochondrial membrane?

There’s no NADH/NAD⁺ transporter; reducing equivalents must be shuttled indirectly.

What is the malate-aspartate shuttle used for?

To transfer cytosolic NADH electrons into the mitochondrial matrix via redox reactions.

What is the glycerol 3-phosphate shuttle and where is it used?

An alternative NADH shuttle in skeletal muscle and brain; faster but less efficient because it transfers electrons to FADH₂ (Complex II).

How are electron transport and ATP synthesis coupled?

Electron transport creates the proton gradient; ATP synthesis dissipates it. Each depends on the other.

What happens if ATP synthesis stops?

The proton gradient builds up, halting electron transport and respiration.

What does DNP demonstrate about coupling?

It uncouples ATP synthesis from respiration, showing that the proton gradient links the two.