Biochem Lec 27- Oxidative Phosphorylation I

What does oxidative phosphorylation do? How much ATP is needed everyday?

Oxidative phosphorylation→Production of ATP from the re-oxidation of FADH2 and NADH

• Electrons are transferred from NADH and FADH2 to O2 through a series of electron carriers.

• We need about 83 kg (!!!!) of ATP everyday to satisfy energy needs. OxPhos efficiently recycles ADP back to ATP.

What is the general flow of oxidative phosphorylation?

Redox power→ Electrochemical gradient→ Chemical energy

1. Electron transport: Harvest reducing power from NADH and FADH₂

2. Proton gradient: “proton motive force”

3. ATP synthesis

Describe the parts of a redox reaction using the following reaction: AH₂ + B→ A + BH₂

• AH₂ is more reduced than A (more electrons/hydrogens)

• B is more oxidized than BH₂

• In this reaction 2 H and 2 e^- are transferred from AH₂ to B

• AH₂ is the reductant/reducing agent

• B is the oxidant/oxidizing agent

• The product A is oxidized (lost electrons)

• BH₂ is reduced (gained electrons)

Why is a redox reaction separated into ½ reaction pairs? On what side of the arrow is the reductant/oxidant drawn?

• ½ reactions→ to judge the power of each oxidant

• Oxidant drawn on left side and reductant drawn on right side

What does E_º‘ measure?

It is the biological standard reduction potential→ is a measure of the tendency of a compound to give up or accept electrons under standard biological conditions

• Compounds that are strong reductants/high tendency to give up electrons→ Negative E_º‘

• Compounds that are strong oxidants/high tendency to accept electrons→ Positive E_º‘

What is the reaction catalyzed by lactate dehydrogenase? What are the half reactions? Is pyruvate or NAD⁺ a stronger reductant?

NADH + H⁺ + pyruvate→ NAD⁺ + lactate

1. NAD⁺ + 2e^- + 2H⁺→ NADH + H⁺: E_º‘= -0.32 volts

2. Pyruvate + 2e^- +2H⁺→ Lactate: E_º‘= -0.19 volts

NAD⁺ is a stronger reductant

What is ΔE_o’ and how is it calculated? What is ΔE_o’ for the lactate dehydrogenase reaction (ΔE_o’_pyruvate= -0.19V and ΔE_o’_NAD= -0.32 V)?

ΔE_o’ is the change in standard reduction potential for an oxidation-reduction reaction and is a measure of “reducing power”

• ΔE_o’= ΔE_o’_oxidant - ΔE_o’_reductant

ΔE_o’= ΔE_o’_pyruvate - ΔE_o’_NAD= -0.19 V - (-0.32V)= +0.13 V

How can ΔG^o’ be calculated from ΔE_o’? What is this value for the lactate dehydrogenase reaction? (ΔE_o’= +0.13 V, F= 96.5 kJ/mol, n= number of electrons transferred)

ΔG^o’ can be calculated from ΔE_o’ using the Nernst equation:

• ΔG^o’= -nFΔE_o’

1. n= number of e^- transferred

2. F= Faraday’s constant→ 96.5 kJ/mol V

ΔG^o’= -nFΔE_o’= -(2)(96.5 kJ/mol V)(+0.13 V)= -25.1 kJ/mol

• Each reductant and oxidant pair has its own ΔE_o’ value→ from these the free energy available from any redox reaction can be calculated

Calculate ΔE_o’ for both the NADH and FADH₂ reactions.

NADH + H + ½ O₂→ NAD⁺ + H₂O

1. NAD⁺ + 2e^- + 2H⁺→ NADH + H⁺: E_o’_NAD= -0.32 volts

2. ½ O₂ + 2e^- + 2H⁺→ H₂O: E_o’_oxygen= +0.82 volts

• ΔE_o’= E_o’_oxygen- E_o’_NAD= +0.82 V - (-0.32 V)= +1.14 V

FADH₂ + ½ O₂→ FAD + H₂O

1. FAD + 2e^- + 2H⁺→ FADH₂: E_o’_FAD= -0.22 V

2. ½ O₂ + 2e^- + 2H⁺→ H₂O: E_o’_oxygen= +0.82 V

• ΔE_o’= E_o’_oxygen- E_o’_FAD= +0.82 V - (-0.22 V)= +1.04 V

Calculate ΔG^o’ for NADH (ΔE_o’= +1.14 V) and FADH₂ (ΔE_o’= +1.04 V).

NADH:

• ΔG^o’= -(2)(96.5 kJ/mol V)(1.14 V)= -220.02 kJ/mol

FADH₂:

• ΔG^o’= -(2)(96.5 kJ/mol V)(1.04 V)= -200.72 kJ/mol

What is the cost of making one ATP? How does this relate to the previously calculated free energies of the NADH and FADH₂ reactions?

• The cost of making one ATP is +30.5 kJ/mol

• Theoretically, many ATP should be able to be produced from this reducing power

How many large protein complexes make up the electron transport chain?

4:

Complex I→ NADH-Q oxidoreductase

Complex II→ succinate Q-reductase

Complex III→ Q-cytochrome c oxidoreductase

Complex IV→ Cytochrome c oxidase

How do the complexes use NADH and what specific complexes are involved?

• Electrons flow from NADH to O2 through four large protein complexes embedded in the inner mitochondrial membrane

• Three complexes move protons out of the mitochondrial matrix, generating a proton gradient:

1. Complex I→ NADH-Q oxidoreductase

2. Complex III→ Q-cytochrome c oxidoreductase

3. Complex IV→ Cytochrome c oxidase

What is the role of complex II in the ETC?

Complex II (succinate Q-reductase)→ delivers electrons from FADH₂ to complex III

• succinate Q-reductase is NOT a proton pump

What are the mobile carriers in the transport chain?

1. Ubiquinone (coenzyme Q)

2. Cytochrome C

They connect the transport chains.

What are the electrons donated by NADH and FADH₂ passed to? Where are they located?

They are passed to electron carriers:

• Coenzyme Q, which is derived from isoprene, binds protons (QH₂) as well as electrons and can exist in several oxidation states

• Oxidized and reduced Q are present in the inner mitochondrial membrane in what is called the Q pool

• Cytochrome C is an electron carrier that employs an iron incorporated into a heme. Cytochrome C carries electrons from Complex III to Complex IV

Overview of ETC

• Site of entry for electrons from NADH and FADH₂ (succinate) from the CAC and other catabolic pathways

• Four multienzyme complexes connected by two mobile carriers: ubiquinone (Q) and cytochrome c

• Electrons flow from -E^º’ to + -E^º’

• During electron flow, H⁺ is pumped from the matrix to the cytosolic side of the mitochondrial inner membrane