9.4 Mitochondrial Architecture & the Respiratory Complexes

9.4 Mitochondrial Architecture & the Respiratory Complexes

The reduced coenzymes generated during glycolysis, the pyruvate dehydrogenase complex, and the citric acid cycle hold high-energy electron pairs. Oxidative phosphorylation liberates this chemical free energy through a series of sequential oxidation-reduction reactions, using the energy to establish a transmembrane electrochemical proton gradient that powers ATP synthesis.

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Mitochondrial Architecture & Thermodynamic Principles

Oxidative phosphorylation relies strictly on the compartmentalized anatomy of the mitochondrion:

• Outer Mitochondrial Membrane (OMM): Highly permeable to small molecules and ions ($<5\text{ kDa}$) due to abundant pore-forming proteins called porins. The solute composition of the intermembrane space is essentially equivalent to the cytosol.
• Intermembrane Space (IMS): The compartment between the outer and inner membranes into which respiratory complexes pump protons. During active respiration, the IMS exhibits a lower pH ($\approx 7.0–7.2$) and a positive electrical potential relative to the matrix.
• Inner Mitochondrial Membrane (IMM): A specialized, protein-dense lipid bilayer folded into extensive invaginations called cristae to maximize functional surface area. Uniquely rich in cardiolipin (diphosphatidylglycerol) and virtually impermeable to all ions (including $\text{H}^+$) and polar solutes. Maintaining this impermeability is an absolute biological requirement for preserving the electrochemical proton gradient.
• Mitochondrial Matrix: The internal compartment containing the PDC, TCA cycle enzymes, $\beta$-oxidation machinery, mitochondrial DNA, and ribosomes. During respiration, it maintains an alkaline pH ($\approx 7.8–8.0$) and a net negative electrical charge.

The Thermodynamic Driving Force: Reduction Potentials

Electrons flow spontaneously through the electron transport chain because each successive carrier possesses a more positive standard reduction potential ($E^{\circ\prime}$) than the preceding donor:

• $\text{NAD}^+/\text{NADH}$: $E^{\circ\prime} = -0.32\text{ V}$ (strong electron donor)
• $\text{FAD}/\text{FADH}_2$: $E^{\circ\prime} \approx -0.22\text{ V}$
• $\text{Ubiquinone (CoQ)}/\text{CoQH}_2$: $E^{\circ\prime} = +0.04\text{ V}$
• $\text{Cytochrome } c\text{ (Fe}^{3+}/\text{Fe}^{2+}\text{)}$: $E^{\circ\prime} = +0.25\text{ V}$
• $\text{O}_2/\text{H}_2\text{O}$: $E^{\circ\prime} = +0.82\text{ V}$ (terminal electron acceptor)

The overall free energy change for the transfer of two electrons from NADH to molecular oxygen is massive:

$$\Delta E^{\circ\prime} = +0.82\text{ V} - (-0.32\text{ V}) = +1.14\text{ V}$$ $$\Delta G^{\circ\prime} = -nF\Delta E^{\circ\prime} = -(2)(96.485\text{ kJ/V}\cdot\text{mol})(1.14\text{ V}) = -220\text{ kJ/mol}$$

This exergonic free energy drop is harnessed by Complexes I, III, and IV to pump protons across the inner membrane against their electrochemical gradient.

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The Respiratory Complexes and Proton-Pumping Stoichiometry

Complex I: NADH-Ubiquinone Oxidoreductase

• Cofactors: Flavin mononucleotide (FMN) and 8 to 9 Iron-Sulfur (Fe-S) centers.
• Electron Pathway: NADH in the matrix binds Complex I, transferring 2 electrons to FMN (forming $\text{FMNH}_2$). Electrons traverse a chain of Fe-S clusters to reduce lipid-soluble Ubiquinone (Coenzyme Q) to Ubiquinol ($\text{QH}_2$).
• Proton Pumping: The exergonic electron transfer induces long-range conformational changes across the membrane-spanning arm, pumping $4\text{ H}^+$ from the matrix into the intermembrane space per electron pair.

Complex II: Succinate-Ubiquinone Reductase (Succinate Dehydrogenase)

• Cofactors: Covalently bound FAD, three Fe-S centers ($[2\text{Fe}-2\text{S}]$, $[4\text{Fe}-4\text{S}]$, $[3\text{Fe}-4\text{S}]$), and a heme $b$.
• Electron Pathway: Succinate is oxidized to fumarate in the matrix, transferring 2 electrons to FAD, through the Fe-S clusters, and onto Ubiquinone to form $\text{QH}_2$.
• Proton Pumping: Because the standard reduction potential difference between succinate/fumarate ($+0.03\text{ V}$) and ubiquinone ($+0.04\text{ V}$) is extremely small, insufficient free energy is released to translocate protons. Complex II pumps $0\text{ H}^+$.

Mobile Carrier 1: Ubiquinone / Coenzyme Q (CoQ)

Ubiquinone is a small, hydrophobic benzoquinone containing an isoprenoid side chain (10 units in humans, $\text{CoQ}_{10}$) that enables it to diffuse freely within the hydrophobic core of the inner mitochondrial membrane. It functions as a flexible two-electron and two-proton collector, accepting reducing equivalents from Complex I, Complex II, glycerol-3-phosphate dehydrogenase, and electron-transferring flavoprotein (ETF) from fatty acid $\beta$-oxidation.

Complex III: Ubiquinol-Cytochrome c Oxidoreductase (Cytochrome $bc_1$ Complex)

• Cofactors: Cytochrome $b$ (hemes $b_L$ and $b_H$), Rieske iron-sulfur protein ($[2\text{Fe}-2\text{S}]$), and cytochrome $c_1$.
• The Q-Cycle Mechanism: $\text{QH}_2$ carries two electrons, but the downstream carrier, Cytochrome $c$, can accept only one electron at a time. Complex III resolves this obligate stoichiometric mismatch via the Q-cycle:
  • In round one, a $\text{QH}_2$ binds the $Q_o$ site. One electron reduces a Rieske Fe-S center and passes to cytochrome $c_1$ and then to a mobile Cytochrome $c$. The second electron passes through hemes $b_L$ and $b_H$ to reduce a bound ubiquinone at the $Q_i$ site, forming a transient semiquinone radical ($\text{Q}^{\bullet-}$).
  • In round two, a second $\text{QH}_2$ binds the $Q_o$ site; one electron reduces a second Cytochrome $c$, while the second electron reduces the semiquinone radical at the $Q_i$ site completely, regenerating one molecule of $\text{QH}_2$.
• Proton Pumping: Across the full Q-cycle, two $\text{QH}_2$ are consumed and one is regenerated (net 1 $\text{QH}_2$ oxidized), two Cytochrome $c$ molecules are reduced, and $4\text{ H}^+$ are pumped into the intermembrane space.

Mobile Carrier 2: Cytochrome $c$

Cytochrome $c$ is a water-soluble, peripheral membrane hemeprotein situated on the outer (IMS-facing) surface of the inner membrane. It carries a single electron via reversible cycling of its central iron atom between ferric ($\text{Fe}^{3+}$) and ferrous ($\text{Fe}^{2+}$) oxidation states, shuttling from Complex III to Complex IV.

Complex IV: Cytochrome c Oxidase

• Cofactors: Heme $a$, heme $a_3$, and two copper centers ($\text{Cu}_A$ and $\text{Cu}_B$).
• Reaction: Catalyzes the 4-electron reduction of molecular oxygen to two water molecules: $$4\text{ Cyt } c\text{ (Fe}^{2+}\text{)} + 4\text{ H}^+\text{ (matrix)} + \text{O}_2 \longrightarrow 4\text{ Cyt } c\text{ (Fe}^{3+}\text{)} + 2\text{ H}_2\text{O}$$
• Proton Pumping: Translocates $2\text{ H}^+$ into the intermembrane space per pair of electrons transferred (equivalent to $4\text{ H}^+$ pumped per $\text{O}_2$ reduced).

Cumulative Proton Pumping per Electron Pair

• Per NADH (Complex I $\to$ III $\to$ IV): $4\text{ H}^+\text{ (I)} + 4\text{ H}^+\text{ (III)} + 2\text{ H}^+\text{ (IV)} = \mathbf{10\text{ H}^+}$
• Per $\mathbf{FADH_2}$ (Complex II $\to$ III $\to$ IV): $0\text{ H}^+\text{ (II)} + 4\text{ H}^+\text{ (III)} + 2\text{ H}^+\text{ (IV)} = \mathbf{6\text{ H}^+}$

Summary of Electron Transport Chain Complexes & Prosthetic Groups