22 oxidative phosphorylation

Oxidative Phosphorylation

Review Overview

  • The lecture focuses on oxidative phosphorylation, which is a fundamental process in aerobic respiration.

  • Key previous topics include:

    • The electron transport chain (ETC) generates a proton gradient across the inner mitochondrial membrane.

    • ATP synthase: structure and mechanism.

    • Coupling and uncoupling of ATP synthesis, proton pumping, and electron transport.

    • Overall energetics of aerobic respiration.

Outline of Lecture 22

  1. ATP Synthase Mechanism

    • ATP synthase utilizes the proton gradient generated by electron transport to synthesize ATP.

    • This reaction stores the energy extracted from previous metabolic stages in the ATP molecule.

    • Oxidative phosphorylation culminates in the major ATP-yielding step of aerobic respiration.

  2. Mitochondrial Function

    • Mitochondria convert reduction potential (electrons) into a proton gradient across the inner membrane, then into ATP.

  3. Proton Gradient and Energy

    • Respiratory complexes in the inner mitochondrial membrane form "supercomplexes" or "respirasomes".

    • The concentration gradient facilitates the transport of protons back to the matrix, making it thermodynamically favorable.

    • The electrical gradient adds to this effect, leading to a significant free energy change of approximately 21 kJ/mol for this process.

    • The electrochemical gradient of H+ across the inner membrane, known as the protonmotive force, is a substantial reservoir of potential energy.

  4. Historical Context

    • Initially, it was not clear that the proton gradient was the source of energy for ATP production; researchers searched for high-energy intermediates.

    • Peter Mitchell won the Nobel Prize in 1978 for his work on the proton motive force.

  5. Gibbs Free Energy Equations

    • The change in Gibbs free energy (ΔG) associated with the movement of protons can be described by the equation:
      ΔG = RT ext{ln} rac{C2}{C1} + ZFΔψ

    • Where:

      • pmf = 2.3 RT ΔpH + FΔψ

      • Free energy components:

      • Concentration (4.5 kJ/mol)

      • Electrical gradient (14.5-19.3 kJ/mol)

  6. Proton Pumping in ETC

    • For each pair of electrons from NADH entering the electron transport chain, 10 protons are pumped out of the matrix.

    • Energy associated:

      • 10 H+ per NADH ≈ 200 kJ/mol

      • ATP synthesis requires approximately 50 kJ/mol.

ATP Synthase Structure and Function

  • ATP synthase (complex V) catalyzes ATP formation from ADP and inorganic phosphate (Pi) using the proton gradient.

  • Structure:

    • Contains a transmembrane Fo sub-complex and F1 "knobs" on the matrix side of the inner membrane.

    • Configurations include:

    • Stator (non-rotating): consisting of b, d, F6, and OSCP subunits.

    • Rotor (rotating): includes c-ring (composed of 8-15 subunits), and γ, δ, ε subunits.

    • The motor-like design with stationary and rotating parts functions similarly to an electric motor.

  • Mechanism:

    • Protonation and re-ionization of glutamate residues in c-ring driving rotation.

  • Flow of protons occurs through channels in the a subunit and allows protons to cross the membrane leading to ATP production.

ATP Synthase Mechanism Detailed

  1. Conformational Changes

    • As ATP synthase rotates, the asymmetry of the coiled-coil rotor causes conformational changes in the α3β3 catalytic domains.

    • Each catalytic subunit transitions through three conformations during each γ-subunit rotation.

  2. Conformational Phases

    • Loose: ADP and Pi binding.

    • Tight: ATP synthesis.

    • Open: Release of ATP.

    • One full rotation (360°) of the γ-subunit generates three ATP molecules.

    • Each 120° rotation alters the conformation of the αβ dimers accordingly.

Summary of Mechanistic Insights

  • The process of moving H+ down an electrochemical gradient (exergonic) is coupled to ATP synthesis (endergonic).

  • Reconstituting oxidative phosphorylation in isolated mitochondria allows detailed study of the pathway and energy flux.

  • Electron transport is directly linked to ATP synthesis; lack of ADP halts electron transport due to elevated proton gradient.

Effect of Inhibitors on ATP Synthase

  • Oligomycin:

    • Used by bacteria to inhibit ATP synthase, binding to c-ring subunits, preventing protons from re-entering the mitochondrial matrix.

    • Inhibition of ATP synthesis leads to a steep proton gradient, making electron transport unsustainable.

Coupling and Uncoupling in Respiration

  • Respiratory Control:

    • When cellular ATP usage increases, ADP levels rise, enhancing ATP synthesis.

  • Uncouplers:

    • Chemical uncouplers (e.g., 2,4-dinitrophenol) allow protons to re-enter the matrix without producing ATP, dissipating the protonmotive force.

    • Different from inhibition, protons are still pumped out, but energy is lost as heat.

Historical Notes on Dinitrophenol

  • Briefly used as a weight-loss agent in the 1930s but banned by the FDA due to severe side effects including risk of overdose leading to energy depletion and potentially fatal consequences.

Physiological Relevance of Uncoupling

  • Regulated uncoupling can be beneficial in organisms like babies or hibernating mammals where brown adipose tissue expresses a proton channel that dissipates energy as heat.

Energy Yield in Aerobic Respiration

  • Each NADH yields approximately 2.5 ATP, while each FADH2 yields around 1.5 ATP based on experimental data.

  • Significant ATP yield from aerobic respiration compared to glycolysis alone.

  • Conclusively, oxidative phosphorylation is responsible for ATP production, and its processes are tightly linked to cellular energy needs.

Final Summary

  • Key points of oxidative phosphorylation discussed include ATP synthesis mechanisms, proton gradients, coupling/uncoupling effects, and the substantial yield of ATP through aerobic respiration compared to glycolysis.

Instructor Q&A

  • Weekly sessions are held for student inquiries regarding lecture material.

    • When: Wednesdays, 4-5 PM

    • Location: Health Sciences Building I132

Office Hours

  • Reader/Grader availability:

    • Sanjay: Wednesdays 9:30-10:20 AM at Health Sciences J375

    • Sidhant: Fridays 1:30-2:20 PM at Health Sciences J375