Citric Acid Cycle and Fatty Acid Oxidation Flashcards

Overview of Glucose Oxidation and Fatty Acid Metabolism

• Glucose Oxidation Stages:

  • Stage I: Glycolysis (occurs in the cytosol).

  • Stage II: Pyruvate is oxidized to generate one molecule each of $CO_2$, $NADH$, and acetyl CoA. The acetyl CoA is subsequently oxidized to $CO_2$ via the citric acid cycle.

  • Energy Storage: Most energy released during Stages I and II is temporarily stored in reduced electron carriers, $NADH$ and $FADH_2$. These carriers deliver high-energy electrons to the electron-transport chain (Stage III).

• Fatty Acid Oxidation:

  • Short- to Long-Chain Fatty Acids: Oxidation occurs within the mitochondria, resulting in the production of $ATP$.

  • Very Long Chain Fatty Acids: Oxidation occurs primarily in peroxisomes. This process generates heat rather than $ATP$.

Mitochondrial Structure and Compartmentalization

• The mitochondrion consists of four distinct compartments:

  1. Outer Membrane: Contains large pores that allow for the passive diffusion of molecules up to a size of $500\,kDa$.

  2. Inner Membrane: A single continuous membrane characterized by three distinct domains:

     • Boundary Membrane: The flat domain located just beneath the outer membrane.

     • Cristae: Sheetlike and tubelike invaginations that extend from the boundary membrane into the center of the organelle. Dimensions are noted as approximately $1-2\,mm$ and $0.1-0.5\,\mu m$.

     • Crista Junctions: Sharp bends connecting the boundary membrane to the cristae.

  3. Intermembrane Space: This space is continuous with the lumen of every crista.

  4. Matrix: The central space surrounded by the inner membrane.

     • Contains the enzymes for the citric acid cycle.

     • Contains mitochondrial DNA (represented as orange spheres in diagrams), ribosomes, and granules.

Pyruvate Transport into the Mitochondrion

• Mitochondrial Pyruvate Carrier (MPC): A hetero-dimer consisting of two small homologous membrane proteins, MPC1 and MPC2 in humans.

• Mechanism: Pyruvate binds to the MPC and is co-transported into the matrix alongside a proton ($H^+$).

Net Yields of Glycolysis and the Citric Acid Cycle

• Glycolysis (1 Glucose to 2 Pyruvate):

  • $CO_2$ Produced: $0$

  • $NAD^+$ reduced to $NADH$: $2$

  • $FAD$ reduced to $FADH_2$: $0$

  • $ATP$ (or $GTP$): $2$

• Pyruvate Oxidation (2 Pyruvates to 2 Acetyl CoA):

  • $CO_2$ Produced: $2$

  • $NAD^+$ reduced to $NADH$: $2$

  • $FAD$ reduced to $FADH_2$: $0$

  • $ATP$ (or $GTP$): $0$

• Citric Acid Cycle (2 Acetyl CoA to 4 $CO_2$):

  • $CO_2$ Produced: $4$

  • $NAD^+$ reduced to $NADH$: $6$

  • $FAD$ reduced to $FADH_2$: $2$

  • $ATP$ (or $GTP$): $2$

• Total Combined Yield:

  • $CO_2$: $6$

  • $NADH$: $10$

  • $FADH_2$: $2$

  • $ATP/GTP$: $4$

The Mitochondrial Electron-Transport Chain (ETC)

• Core Components: Electrons flow through four major multiprotein complexes labeled I–IV.

• Mobile Electron Carriers:

  • Coenzyme Q (CoQ): Lipid-soluble; exists in an oxidized form (CoQ) and a reduced form ($CoQH_2$).

  • Cytochrome c (cyt c): Water-soluble protein carrier.

• The Electron Path: $NADH$ → Complex I → $CoQH_2$ → Complex III → cyt c → Complex IV → $O_2$ (forming $H_2O$).

• Complex I (NADH Dehydrogenase):

  • $NADH$ donates two electrons via Flavin Mononucleotide (FMN) and iron-sulfur clusters to CoQ.

  • Conformational Changes: Electron flow induces a piston-like horizontal movement of the t-helix.

  • Proton Pumping: This mechanical movement drives the pumping of four protons ($4\,H^+$) from the matrix into the intermembrane space per electron pair.

• Reduction Potentials ($E^{\circ \prime}$):

  • The reduction potential of electron carriers increases as electrons move down the chain, favoring spontaneous flow.

  • $NADH$: $-320\,mV$

  • $O_2$: $+860\,mV$ (highest potential, terminal acceptor).

• The Q Cycle (Complex III):

  • An evolutionarily conserved mechanism that optimizes proton transport.

  • Net result: Oxidation of one $CoQH_2$ molecule transfers four protons into the intermembrane space and two electrons to two cytochrome c molecules.

Chemiosmosis and the Proton-Motive Force (PMF)

• Definition: Chemiosmosis is the process where a proton-motive force, generated by proton pumping across a membrane, powers $ATP$ synthesis.

• Proton-Motive Force Components:

  1. Membrane Potential ($\Delta V$): A large electrical gradient.

  2. Concentration Gradient ($\Delta pH$): A smaller chemical gradient of $H^+$ ions.

• Pumping Directions:

  • Bacteria: Protons pumped from cytosolic face to exoplasmic face.

  • Mitochondria: Protons pumped from the matrix to the intermembrane space.

• ATP Synthesis: Protons flow down their electrochemical gradient through the $F_0F_1$ complex (ATP synthase).

Structure and Mechanism of ATP Synthase

• $F_0F_1$ Complex Components:

  • $F_0$: The rotating portion embedded in the membrane; includes the c ring and a subunit.

  • $F_1$: The stationary head (ATPase), containing alternating $\alpha$ and $\beta$ subunits and the central $\gamma$ subunit.

• Proton Path through $F_0$:

  1. Proton enters Half-channel I from the intermembrane space.

  2. Arg-210 is displaced.

  3. Proton binds to a negative charge on Asp-61 within the c ring.

  4. The c ring rotates.

  5. The proton exits through Half-channel II into the matrix.

• Binding-Change Mechanism (Viewed from the membrane surface):

  • Proton movement through $F_0$ drives the rotation of the $F_1$ $\gamma$ subunit, which forces conformational changes in the $\beta$ subunits.

  • O (Open) State: Binds $ATP$ very poorly; binds $ADP$ and $P_i$ weakly.

  • L (Loose) State: Binds $ADP$ and $P_i$ more strongly; cannot bind $ATP$.

  • T (Tight) State: Binds $ADP$ and $P_i$ tightly enough to spontaneously catalyze the formation of $ATP$.

• Reversibility: ATP synthase can act as a pump and hydrolyze $ATP$ to rebuild the proton gradient if the electrochemical gradient falls below a specific threshold.

Product Yields and Efficiency of Glucose Oxidation

• Glycolysis (Direct): $2\,ATP$ + $2\,NADH$ (cytosolic). Yield: $3-5\,ATP$.

  • Note: Cytosolic $NADH$ yields fewer $ATP$ molecules ($1.5$ to $2$) because the inner membrane is impermeable to $NADH$; transport (e.g., malate-aspartate shuttle) requires energy.

• Pyruvate Oxidation: $2\,NADH$ (matrix). Yield: $5\,ATP$.

• Acetyl Group Oxidation (CAC): $6\,NADH$ (matrix) + $2\,FADH_2$ + $2\,GTP$. Yield: $15\,ATP$ (from $NADH$) + $3\,ATP$ (from $FADH_2$) + $2\,ATP$ (from $GTP$).

• Total Yield: Approximately $30\,ATP$ per glucose molecule.

Uncoupling and Toxicity

• Uncoupling Proteins (UCPs): These proteins allow protons to leak back across the inner mitochondrial membrane, bypassing ATP synthase. The energy from the gradient is released as heat rather than captured as $ATP$.

• 2,4-Dinitrophenol (DNP):

  • A chemical uncoupling agent and highly toxic industrial chemical.

  • Historically and illegally used as a diet pill; it is extremely dangerous with "no safe dose."

  • Causes health professionals to issue warnings due to increasing related deaths.

Principles of Free Energy ($G$)

• Definition: Free energy ($G$) measures the energy of a molecule that can be used to do work at constant temperature.

• Spontaneous Reactions: Occur when \Delta G < 0. Disorder in the universe increases.

• Coupled Reactions: An unfavorable reaction (\Delta G^{\circ} > 0) can be driven by a favorable reaction (\Delta G^{\circ} < 0) if they share intermediates.

  • Example: Making sucrose ($\Delta G^{\circ} = +23\,kJ/mol$) is driven by $ATP$ hydrolysis ($\Delta G^{\circ} = -30.5\,kJ/mol$), resulting in a net $\Delta G^{\circ} = -7.5\,kJ/mol$.

• Equilibrium Constant ($K$): Fixed relationship to $\Delta G^{\circ}$. At $37^{\circ}C$:

  • $\Delta G^{\circ} = -5.94 \log_{10}(K)$.

• High-Energy Bonds: Bonds in compounds like acetyl phosphate ($\Delta G^{\circ} = -43.1\,kJ/mol$) and $ATP$ ($\Delta G^{\circ} = -30.5\,kJ/mol$) that release significant energy upon hydrolysis.

Photosynthesis: Four-Stage Overview

• Light Reactions (Stages 1-3): Absorption of light, generation of high-energy electrons, formation of $O_2$, electron transport (creating PMF), and $ATP$/$NADPH$ synthesis. Occurs in the thylakoid membrane.

• Dark Reactions (Stage 4): Carbon fixation and carbohydrate synthesis (sucrose and starch). Occurs in the stroma.

• Structure: Chloroplasts have a double membrane, thylakoid lumen (low pH), and stroma.

Sample Exam Question & Discussion

• Question: Which of the following is NOT true of chemiosmosis?

  • Options:

    • ATP synthase generates ATP from ADP and Pi

    • It produces majority of ATP in a cell

    • The proton-motive force is used to synthesize ATP

    • It requires oxygen (This is the correct answer for the "NOT true" prompt, as chemiosmosis itself is the movement of ions; while respiratory chemiosmosis uses oxygen as a terminal acceptor, the process of chemiosmosis/PMF utilization can occur in anaerobic or photosynthetic conditions).

Exam 1 Study Guide Topics

• Compare oxidative phosphorylation in eukaryotes vs. bacteria.

• Contrast electron sources for respiration (organic molecules) vs. photosynthesis (water/sunlight).

• Explain why $NADH$ does not donate electrons directly to $O_2$ (energy would be lost as heat rather than captured).

• Identify the direction of proton pumping (matrix to intermembrane space).

• Distinguish substrate-level phosphorylation from oxidative phosphorylation.