L9-chpt5 part 2-spring 2025

Mitochondria II: Oxidative Phosphorylation

• Concepts include:

  • Redox reactions

  • The electron transport chain

  • ATP synthesis

Glycolysis Key Points

• Location: Cytosol

• Starting Reactant: 1 glucose (6 carbons)

• End Product: 2 pyruvate (3 carbons)

• Energy: 4 ATP formed – 2 ATP used = 2 ATP net gain

• Co-enzymes Reduced: 2 NADH + 2H+

Fates of Pyruvate

• Aerobic:

  • Pyruvate crosses mitochondrial membranes

  • Further oxidized via the TCA cycle

• Anaerobic:

  • Different metabolic pathways, typically involving fermentation.

The Warburg Effect in Cancer

• Definition: Tumors and proliferating cells often skip oxidative phosphorylation, favoring high rates of aerobic glycolysis even in the presence of oxygen and functional mitochondria.

• Advantages: Investigate if the Warburg effect offers any metabolic advantages to cancer cells.

Anaerobic Metabolism and NAD+ Production

• Cells regenerate NAD+ required for glycolysis via:

  • C: From fermentation of pyruvate to lactic acid.

ATP Production Experiment

• If mitochondria from muscle cells are purified and glucose is added, will ATP be produced?

  • A: yes

  • B: no

Learning Goals for Oxidative Phosphorylation

By the end of today's topic, you should be able to:

• Describe energy transfer/storage during oxidative phosphorylation.

• Explain the importance of redox potentials.

• Calculate free energy changes with redox potentials.

• Know major electron transport chain complexes and their functions.

• Describe mitochondrial ATP synthase function and its coupling with the proton-motive force.

TCA Cycle Key Points

• Reaction: Acetyl CoA + 2 H2O + FAD + 3 NAD+ + GDP + Pi → 2 CO2 + FADH2 + 3 NADH + 3 H+ + GTP + HS-CoA

• Location: Mitochondrial matrix

• Starting Reactant: 2-carbon Acetyl CoA

• End Product: Cycle maintains continuity with 4-carbon oxaloacetate and releases CO2.

• Energy: 1 GTP formed, 4 NADH formed, 1 FADH2 formed, producing 30+ ATP.

Oxidative Phosphorylation Overview

• Process: Making ATP using energy released from electron oxidation.

• Accounts for over 2 x 10^26 ATP molecules produced daily.

Summary of Oxidative Phosphorylation Process

1. High-energy electrons from FADH2 or NADH pass through the electron transport chain in the inner membrane.

2. Protons (H+) move inward through ATP-synthesizing enzyme, driving ATP synthesis. This energy-coupling is called chemiosmosis.

Oxidation-Reduction Potentials

• Redox Potential:

  • Strong oxidizing agents: High electron affinity

  • Strong reducing agents: Low electron affinity.

• Redox reactions involve electron transfer causing charge separation measurable as redox potential against H+-H2 standard.

• Oxidation: Loss of electrons

• Reduction: Gain of electrons.

Redox Potential of Reaction Couples

• Better reducing agents (electron donors): Strong reducing agents like NADH versus better oxidizing agents like NAD.

Free Energy Change Calculation for TCA Reaction

• Equation: DG0’ = -nF DE’0 where:

  • n: Number of electrons transferred

  • F: Faraday constant (23.063 kcal/V·mol)

  • E: Voltage difference in standard redox potentials.

TCA Cycle Redox Potentials

• Three TCA reactions with high negative redox potential transfer electrons to 3 NAD+.

• Lower redox potential reaction transferring electrons to FAD is catalyzed by succinate dehydrogenase.

Electron Transport Chain (ETC)

• Electrons move through inner membrane carriers with increasing positive redox potential, losing energy downhill until reaching O2, which becomes water.

Electron Carriers in the ETC

• Types of carriers:

  • Flavoproteins: Bound to FAD or FMN.

  • Cytochromes: Contain heme groups with Fe/Cu ions.

  • Ubiquinone (coenzyme Q): Lipid-soluble, consists of isoprenoid units.

  • Iron-sulfur proteins: Involves inorganic sulfur.

Arrangement of Electron Carriers

• Electrons lose energy while moving down the chain, affecting redox potentials.

• The final electron acceptor is O2.

Identifying Electron Transport Complexes

• The specific sequence of carriers in the ETC established via inhibitors that block transport at various sites.

Reduction Potential Favoring Electron Flow

• Flow Sequence:

  • Complex I: NADH dehydrogenase, electrons to ubiquinone, transports 4 H+.

  • Complex II: Succinate oxidized to FAD to ubiquinone, no H+ transport.

  • Complex III: Transfers electrons from ubiquinone to cytochrome c, transports 4 H+.

  • Complex IV: Transfers electrons to O2, transporting 4 H+.

The Proton-Motive Force

• Proton concentration gradients create pH gradients and an electric potential across the mitochondrial membrane, constituting the proton-motive force (Δp).

ATP Synthase Structure and Mechanism

• ATP synthase has F1 particle as catalytic subunit (α3β3δγε) with 3 catalytic sites for ATP synthesis.

• F0 particle is membrane-embedded, facilitating proton movement.

Binding Change Mechanism of ATP Formation

• The mechanism specifies that energy from proton movement changes binding affinities, allowing for ATP synthesis by successive conformational changes in active sites (L, T, O conformations).

Rotational Catalysis in ATP Synthase

• Provides evidence for ATP synthesis; a rotating mechanism is observed aiding ATP creation as protons diffuse.

Additional Roles of Proton-Motive Force

• Drives ADP and Pi uptake, promotes Ca2+ import into mitochondria, and supports mitochondrial fusion activities.

Peroxisomes

• Membrane-bound vesicles containing oxidative enzymes for:

  • Oxidizing very-long-chain fatty acids.

  • Synthesizing plasmalogens (phospholipids).

  • Engaging in oxidative metabolism and importing proteins.

• Hydrogen peroxide (H2O2) produced is decomposed by catalase.

Additional Resources

• Video on mitochondria: YouTube Mitochondria

• Other potentially useful videos:

  • Video 1

  • Video 2