Oxidative phosphorylation pt2 (final exam)

ATP synthase (F1 F0 ATP synthase)

-catalyzes the formation of ATP from ADP + Pi

-uses the proton motive force as its energy force

-converts energy stored in a proton gradient into chemical energy (ATP)

F1 Portion of ATP Synthase

-Peripheral portion facing the mitochondrial matrix

-Catalyzes ATP synthase

-Contains 9 subunits (3a, 3b, y, d, e)

β Subunits of ATP Synthase

• Each β subunit contains a catalytic site for ATP synthesis.

αβ Arrangement of ATP Synthase

-α and β subunits assemble like "sections of an orange."

γ (Gamma) Subunit of ATP Synthase

-Forms a stalk through the center of the αβ complex

-is off-center

-as it rotates it contacts different αβ pairs

-contact causes conformational changes

Conformations of αβ Pairs in ATP Synthase

Each αβ pair cycles through 3 stages

1. Empty

-No ATP or ADP bound

2. ADP + Pi state

-Ready for ATP synthesis

3. ATP state

-ATP has been formed

Explain: Gamma Rotation → Conformational Change → ATP Formation

1. Gamma rotates.

2. Gamma contacts a different αβ pair.

3. Contact forces a conformational change.

4. ADP + Pi are joined to form ATP.

5. ATP is released from another αβ pair.

6. Cycle repeats.

ATP Hydrolysis

-Exergonic

-releases energy

ATP Synthesis

-Endergonic

-Requires energy input

F0 portion of ATP Synthase

-Composed of three subunits: a, b2, c8-12

-ATP yield calculations depend on the number of C subunits: Humans 8c, Yeast 10c, E. coli 12c

F0 C subunit

-contains two transmembrane helices

-looks like “nunchuncks”

-arrange in a circular ring

-small hydrophobic peptide

Explain how the rotation in the C subunit comes from the protonation of Asp61

1. Aspartate begins as unprotonated

2. A proton binds to Asp61

3. Protonation causes conformational change

4. Conformational change causes rotation

5. Asp passes the proton to the next Asp

6. The next Asp picks up a proton

7. Rotation continues around the ring

8. Rotation of the C ring rotates ε

9. ε rotates γ

10. γ drives ATP synthesis

How many ATP molecules are produced from one complete rotation of gamma?

3 ATP molecules per gamma rotation

ATP yield in E. coli

-12 C subunits

-12 protons = 1 complete rotation

-12 protons = 3 ATP

-4 protons required per ATP

Why NADH Produces 2.5 ATP?

Per 1 NADH entering Complex 1

-10 protons pumped

-Need: 12 protons —> 3 ATP

-Math: 10 × 3/12 = 2.5 ATP

Why FADH2 produces 1.5 ATP?

Per 1 FADH2 entering complex 2

-6 protons pumped

-Need: 12 protons —> 3 ATP

-Math: 6 × 3/12 = 1.5 ATP

ATP/ADP Antiporter

1. ATP leaves matrix

2. ADP enters matrix

Proton/Pi Symporter

1. proton moves inward

2. inorganic phosphate travels with proton inside the matrix

Malate-Aspartate Shuttle

-Transfers reducing equivalents (e-) from cytosolic NADH into the mitochondrial matrix

-Produces mitochondrial NADH

-2.5 ATP per NADH

Explain the mechanism of Malate-Aspartate Shuttle

Cytosol

1. NADH donates reducing equivalents

2. OAA is converted to malate by malate dehydrogenase

3. electricity now stored in malate

Transport

4. Malate enters matrix

5. α-Ketoglutarate exits

6. Uses malate/α-ketoglutarate antiporter.

Matrix

7. Malate converted back to OAA

8. NADH produced in matrix

9. NADH enters complex 1

10. ETC makes 10 protons

11. 2.5 ATP produced

Aspartate Cycle

12. OAA + glutamate → aspartate + α-ketoglutarate

13. Aspartate exported

14. Glutamate imported

15. shuttle repetes

Glycerol-3-Phosphate Shuttle

-Transfers the reducing equivalents of NADH to ubiquinone

-1.5 ATP per NADH

-6 protons pumped

Explain the mechanism of the glycerol-3-phosphate shuttle

Cytosol

1. NADH reduces DHAP

2. DHAP becomes G3P

Mitochondrial Side

3. Glycerol-3-phosphate dehydrogenase oxidizes glycerol-3-phosphate

4. FAD becomes FADH2

5. FADH2 reduces ubiquinone (Q)

6. Q becomes QH2

7. Electrons enter ETC at complex 3

8. 1.5 ATP produced

Regulation of Oxidative Phosphorylation

Main Regulators

-ADP + Pi

High ADP + Pi

-cell need ATP

-Ox phos. increase

High ATP

-cell has sufficient energy

-Ox Phos. slowed

Reactive Oxygen Species

Superoxide Dismutase

-Superoxide Dismutase converts superoxide into hydrogen peroxide

Catalase

-catalase converts hydrogen peroxide into water

Hydroxyl Radicals

-cause immediate damage to nearby molecules

-super reactive