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13.1 Overview of Pyruvate Oxidation and the Citric Acid Cycle

Oxidative Processes in the Generation of Metabolic Energy

The citric acid cycle (CAC), also known as the Krebs cycle or TCA cycle, is the central pathway for oxidizing metabolic fuels in aerobic organisms. Most of the energy yield from substrate oxidation in the CAC is stored in reduced electron carriers such as NADH.

• Central Pathway: The CAC is the main route for the complete oxidation of carbohydrates, fats, and proteins.

• Energy Storage: Energy is captured in the form of reduced electron carriers (NADH, FADH2).

• Example: NADH generated in the CAC is later used in the electron transport chain to produce ATP.

Stages of Cellular Respiration

Cellular respiration is divided into three main stages, each with distinct biochemical roles:

• Stage 1: Carbon from metabolic fuels is incorporated into acetyl-CoA.

• Stage 2: The citric acid cycle oxidizes acetyl-CoA to produce CO2, reduced electron carriers, and a small amount of ATP.

• Stage 3: Reduced electron carriers are reoxidized, providing energy for the synthesis of additional ATP via oxidative phosphorylation.

In eukaryotes, all three stages occur in the mitochondria.

Structure of Mitochondria

The mitochondrion is the organelle where cellular respiration takes place. It has distinct compartments:

• Mitochondrial Matrix: Location of stages 1 and 2 reactions (acetyl-CoA formation and CAC).

• Inner Mitochondrial Membrane: Location of stage 3 reactions (electron transport and ATP synthesis).

Example: The enzymes of the CAC are soluble in the matrix, while the electron transport chain complexes are embedded in the inner membrane.

13.2 Pyruvate Oxidation: A Major Entry Route for Carbon into the Citric Acid Cycle

The Pyruvate Dehydrogenase (PDH) Multienzyme Complex

Pyruvate, the end product of glycolysis, is transported into mitochondria and converted to acetyl-CoA by the pyruvate dehydrogenase (PDH) complex. This is a key regulatory step linking glycolysis and the CAC.

• Enzyme Components:

  1. Pyruvate dehydrogenase (E1)

  2. Dihydrolipoamide transacetylase (E2)

  3. Dihydrolipoamide dehydrogenase (E3)

• Function: Converts pyruvate to acetyl-CoA, producing NADH and CO2.

Example: The PDH complex is a large multienzyme assembly, facilitating substrate channeling and efficient catalysis.

Mechanistic Overview of PDH Complex

The PDH complex catalyzes a multi-step reaction involving decarboxylation, oxidation, and transfer of the acetyl group to CoA.

• Step 1: Decarboxylation of pyruvate.

• Step 2: Oxidation and transfer to lipoamide.

• Step 3: Transfer of acetyl group to CoA.

• Step 4: Regeneration of oxidized lipoamide and production of NADH.

Equation:

$\text{Pyruvate} + \text{CoA} + \text{NAD}^+ \rightarrow \text{Acetyl-CoA} + \text{CO}_2 + \text{NADH} + \text{H}^+$

13.3 The Citric Acid Cycle

The Fate of Carbon in the Citric Acid Cycle

The CAC consists of eight sequential reactions. The product of the eighth reaction, oxaloacetate, and acetyl-CoA (from PDH complex) are reactants for the first reaction, making the cycle continuous.

• Products per Turn: 2 CO2, 3 NADH, 1 FADH2, and 1 ATP (or GTP).

• Carbon Tracking: Carbon atoms from acetyl-CoA are released as CO2 during the cycle.

Citric Acid Cycle Reactions

• Reaction 1 – Citrate Synthase: Acetyl-CoA + Oxaloacetate + H2O → Citrate + CoA-SH Hydrolysis of the thioester bond makes this step highly exergonic ($\Delta G^\circ' = -32.2$ kJ/mol).

• Reaction 2 – Aconitase: Citrate → cis-Aconitate → D-Isocitrate Two-step isomerization ($\Delta G^\circ' = +6.3$ kJ/mol).

• Reaction 3 – Isocitrate Dehydrogenase: Isocitrate + NAD+ → α-Ketoglutarate + CO2 + NADH Oxidative decarboxylation ($\Delta G^\circ' = -11.6$ kJ/mol).

• Reaction 4 – α-Ketoglutarate Dehydrogenase Complex: α-Ketoglutarate + NAD+ + CoA-SH → Succinyl-CoA + CO2 + NADH Analogous to PDH complex ($\Delta G^\circ' = -33.5$ kJ/mol).

• Reaction 5 – Succinyl-CoA Synthetase: Succinyl-CoA + GDP (or ADP) + Pi → Succinate + GTP (or ATP) + CoA-SH Substrate-level phosphorylation ($\Delta G^\circ' = -3.8$ kJ/mol).

• Reaction 6 – Succinate Dehydrogenase: Succinate + FAD → Fumarate + FADH2 Delivers electrons directly to the electron transport chain ($\Delta G^\circ' = -11.6$ kJ/mol).

• Reaction 7 – Fumarase: Fumarate + H2O → L-Malate Hydration reaction, highly stereospecific ($\Delta G^\circ' = -3.8$ kJ/mol).

• Reaction 8 – Malate Dehydrogenase: L-Malate + NAD+ → Oxaloacetate + NADH + H+ Endergonic, but driven forward by low oxaloacetate and NADH levels ($\Delta G^\circ' = +29.7$ kJ/mol).

13.4 Stoichiometry and Energetics of the Citric Acid Cycle

Overview of One Turn of the Citric Acid Cycle

One complete turn of the CAC yields:

• 1 high-energy phosphate (ATP or GTP)

• 3 NADH

• 1 FADH2

The energy stored in NADH and FADH2 is used to drive ATP synthesis in the electron transport chain.

Overall Reaction:

$\text{Acetyl-CoA} + 2\text{H}_2\text{O} + 3\text{NAD}^+ + \text{FAD} + \text{ADP} + \text{P}_i \rightarrow 2\text{CO}_2 + 3\text{NADH} + 3\text{H}^+ + \text{FADH}_2 + \text{ATP} + \text{CoA}$

14.1 The Mitochondrion: Scene of the Action

Overview of Oxidative Energy Generation

Most ATP is generated during the reoxidation of NADH and FADH2 in the electron transport chain (stage 3 of respiration). Glycolysis and the CAC (stages 1 and 2) produce only a small amount of ATP directly.

• NADH and FADH2 Production: 10 NADH and 2 FADH2 per glucose molecule.

• ATP Synthesis: Reoxidation of these carriers provides the energy for most ATP synthesis.

Mitochondrial Localization of Citric Acid Cycle and Oxidative Phosphorylation

Respiratory processes are compartmentalized within the mitochondrion:

• Matrix: CAC and fatty acid oxidation.

• Inner Membrane: Electron transport chain and ATP synthase.

• Intermembrane Space: Contains enzymes for nucleotide metabolism.

14.3 Electron Transport

Electron Carriers in the Respiratory Chain

The electron transport chain (ETC) catalyzes the flow of electrons from low to high reduction potential carriers, ultimately reducing oxygen to water.

• Flavoproteins: Contain FMN or FAD, participate in redox reactions.

• Iron–Sulfur Proteins: Contain nonheme iron clusters (FeS, Fe4S4), act as single electron carriers.

• Coenzyme Q (Ubiquinone): Lipid-soluble, can transfer two electrons in one step via a semiquinone intermediate.

• Cytochromes: Proteins containing heme groups, classified by absorption spectra (b, c, a types).

Iron–Sulfur Clusters

Iron–sulfur clusters are prosthetic groups in proteins, consisting of nonheme iron complexed with cysteine thiol sulfurs. Their reduction potential varies with cluster type and protein environment.

• Function: Single electron transfer in the ETC.

Coenzyme Q

Coenzyme Q is a mobile electron carrier in the inner mitochondrial membrane.

• Electron Transfer: Can transfer two electrons in one step, bridging two-electron and one-electron carriers.

Cytochromes

Cytochromes are classified by their absorption spectra and contain heme prosthetic groups.

• Types: Cytochromes b, c, and a, each with distinct spectral properties.

Electron Carrier	Structure/Group	Function
Flavoproteins	FMN, FAD	Redox reactions, initial electron acceptors
Iron–Sulfur Proteins	FeS, Fe4S4	Single electron transfer
Coenzyme Q	Ubiquinone	Mobile carrier, two-electron transfer
Cytochromes	Heme groups	Single electron transfer, classified by spectra

Additional info: The notes above expand on the brief points in the slides, providing definitions, context, and examples for each major concept. The table summarizes the main electron carriers in the mitochondrial respiratory chain for comparison.