Electron Transport Chain (ETC) – Components & Inhibitors
Section A: Core Components of the ETC
Q1. Explain how the structure and arrangement of ETC components facilitate ATP synthesis.
Answer:
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ETC is located in the inner mitochondrial membrane.
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Electrons from NADH and FADH₂ pass through complexes I–IV, ultimately reducing O₂ to H₂O.
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Energy released during electron transfer is used by complexes I, III, and IV to pump H⁺ into the intermembrane space, creating a proton gradient.
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This electrochemical gradient drives ATP synthase (Complex V) to generate ATP from ADP + Pi.
Clinical Insight: Disruption in the gradient or complex function impairs oxidative phosphorylation, leading to energy failure in tissues.
Q2. Compare the roles of Complex I and Complex II in electron transport. Why does FADH₂ produce less ATP than NADH?
Answer:
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Complex I (NADH dehydrogenase) accepts electrons from NADH and pumps 4 H⁺ into the intermembrane space.
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Complex II (Succinate dehydrogenase) transfers electrons from FADH₂ directly to coenzyme Q but does not pump protons.
Thus:
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NADH → More H⁺ pumped → More ATP (~3 ATP)
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FADH₂ → Less H⁺ pumped → Less ATP (~2 ATP)
Conclusion: FADH₂ bypasses Complex I, resulting in lower ATP yield.
Q3. Why is coenzyme Q (ubiquinone) important in the ETC, and how is it different from cytochrome c?
Answer:
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Coenzyme Q (CoQ) is a lipid-soluble mobile carrier that shuttles electrons between Complexes I/II and III.
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Cytochrome c is a water-soluble protein that transfers electrons from Complex III to Complex IV.
Difference:
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CoQ moves within the inner membrane, while cytochrome c moves in the intermembrane space.
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CoQ accepts 2 electrons, cytochrome c carries 1 electron.
Clinical significance: CoQ deficiency impairs ETC flow, leading to myopathies and encephalopathies.
Section B: Inhibitors of the ETC
Q4. A cyanide poisoning victim presents with lactic acidosis and high venous O₂ content. Explain the biochemical mechanism behind these findings.
Answer:
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Cyanide inhibits Complex IV (cytochrome c oxidase) → prevents electron transfer to oxygen → halts ATP production.
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Cells switch to anaerobic glycolysis → lactic acid accumulates → lactic acidosis.
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O₂ is not consumed → high venous O₂ content (tissue hypoxia despite adequate oxygen supply).
Q5. A patient accidentally ingests rotenone, a pesticide. Which ETC complex is inhibited, and what is the consequence on ATP production?
Answer:
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Rotenone inhibits Complex I, blocking electron transfer from NADH to CoQ.
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This stops proton pumping at Complex I → reduced proton gradient → decreased ATP synthesis.
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Electrons from FADH₂ via Complex II can still enter, but ATP production is diminished.
Q6. Explain how oligomycin affects the ETC and ATP production.
Answer:
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Oligomycin inhibits ATP synthase (Complex V).
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Proton gradient builds up but can’t be used to synthesize ATP.
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Eventually, ETC slows down due to back pressure from the high proton gradient.
Clinical insight: Oligomycin is a research tool, not typically encountered clinically.
Q7. How do uncouplers like 2,4-dinitrophenol (DNP) disrupt oxidative phosphorylation?
Answer:
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Uncouplers dissipate the proton gradient by allowing H⁺ to flow back into the matrix without passing through ATP synthase.
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ETC continues → oxygen consumption increases
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But ATP production stops
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Energy is released as heat → hyperthermia
DNP was used as a weight loss agent, but caused fatal hyperthermia.
Section C: Clinical Integration & Problem-Solving
Q8. A child presents with exercise intolerance and muscle biopsy shows ragged red fibers. Which mitochondrial function is likely impaired and why?
Answer:
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Likely a mitochondrial myopathy affecting ETC function.
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Ragged red fibers = accumulated defective mitochondria
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Defective ETC → impaired ATP production → muscle weakness, especially during exercise.
Common cause: mtDNA mutations affecting ETC complexes, especially Complex I or IV.
Q9. How does ischemia (e.g., in myocardial infarction) affect ETC function and lead to cell injury?
Answer:
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Ischemia → ↓O₂ delivery → ETC can’t transfer electrons to O₂.
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ATP production stops → ion pumps fail → cell swelling, lysis
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Accumulation of NADH → anaerobic glycolysis and lactic acidosis
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Release of cytochrome c from mitochondria may trigger apoptosis
Q10. Why is the inner mitochondrial membrane impermeable to protons, and how is this critical to ETC function?
Answer:
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The inner mitochondrial membrane is highly selective, maintaining the proton gradient created by ETC complexes.
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This gradient is the driving force for ATP synthesis via chemiosmosis.
If membrane integrity is lost:
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Proton leak → no ATP formation
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Seen in mitochondrial diseases, uncoupling, or membrane damage.
Oxidative Phosphorylation & Uncouplers
Section A: Conceptual Understanding
Q1. What is oxidative phosphorylation and how is it linked to the electron transport chain (ETC)?
Answer:
Oxidative phosphorylation is the final stage of cellular respiration, where ATP is synthesized as electrons pass through the ETC and generate a proton gradient across the inner mitochondrial membrane.
Process:
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Electrons from NADH/FADH₂ move through ETC complexes I–IV.
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This drives proton (H⁺) pumping into the intermembrane space.
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The resulting proton gradient (electrochemical potential) powers ATP synthase (Complex V) to convert ADP + Pi → ATP.
Linkage: Electron transport provides the energy for phosphorylation of ADP – hence the term “oxidative phosphorylation.”
Q2. Why does NADH generate more ATP than FADH₂?
Answer:
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NADH donates electrons at Complex I, which pumps 4 protons, contributing more to the proton gradient.
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FADH₂ donates electrons at Complex II, which does not pump protons.
As a result:
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NADH → ~2.5 to 3 ATP
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FADH₂ → ~1.5 to 2 ATP
So NADH contributes more energy toward ATP synthesis than FADH₂.
Q3. How does the proton motive force drive ATP synthesis in mitochondria?
Answer:
The proton motive force (PMF) consists of:
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Chemical gradient (difference in proton concentration)
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Electrical gradient (membrane potential)
Protons flow downhill through ATP synthase, causing conformational changes in the F₁ subunit, which facilitates ATP formation from ADP + Pi.
Section B: Uncouplers and Their Effects
Q4. Define uncouplers and explain their mechanism of action.
Answer:
Uncouplers are substances that dissipate the proton gradient across the inner mitochondrial membrane without ATP synthesis.
Mechanism:
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They carry protons back into the matrix independent of ATP synthase.
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ETC continues (electrons move, O₂ is consumed), but ATP is not produced.
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Energy is released as heat instead.
Q5. What are the physiological and pathological effects of uncoupling oxidative phosphorylation?
Answer:
Effects:
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Increased oxygen consumption
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No ATP generation
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Heat production → can cause hyperthermia
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Leads to cellular energy crisis, lactic acidosis
Clinical relevance:
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Seen in 2,4-dinitrophenol (DNP) poisoning
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Also occurs physiologically in brown adipose tissue via thermogenin
Q6. How do uncouplers differ from inhibitors of the electron transport chain?
Answer:
| Feature | Uncouplers | ETC Inhibitors |
|---|---|---|
| Mechanism | Dissipate proton gradient | Block electron flow |
| ETC activity | Continues | Stops |
| ATP synthesis | Decreases | Stops |
| O₂ consumption | Increases | Decreases |
| Example | DNP, thermogenin | Cyanide, rotenone, oligomycin |
Q7. Why does oxygen consumption increase in the presence of an uncoupler like DNP?
Answer:
DNP dissipates the proton gradient, so ATP synthase cannot operate.
To compensate:
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ETC speeds up to maintain the gradient
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More oxygen is consumed to accept electrons at Complex IV
But since protons bypass ATP synthase, ATP is not formed, and energy is lost as heat.
Section C: Clinical & Experimental Contexts
Q8. A man consumes 2,4-dinitrophenol (DNP) for weight loss and presents with fever, rapid breathing, and lactic acidosis. Explain the biochemical basis.
Answer:
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DNP is an uncoupler → proton gradient collapses
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ETC and O₂ consumption increase
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ATP not produced, cells rely on anaerobic glycolysis → lactic acidosis
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Energy is released as heat → hyperthermia
Outcome: Potentially fatal unless promptly treated.
Q9. What is the role of thermogenin in brown adipose tissue, and how does it function as a physiological uncoupler?
Answer:
Thermogenin (UCP-1) is a natural uncoupling protein in brown fat.
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Allows protons to re-enter mitochondrial matrix without ATP production
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Generates heat, important in neonates and hibernating animals
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Maintains body temperature
Q10. Why does oligomycin cause a decrease in oxygen consumption, unlike DNP?
Answer:
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Oligomycin inhibits ATP synthase, preventing proton flow back into the matrix.
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Proton gradient builds up → ETC slows down due to back pressure
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Result: ↓ oxygen consumption and ↓ ATP production
Contrast with DNP: DNP destroys the gradient, so ETC accelerates, but ATP still isn’t made.
Theories of Oxidative Phosphorylation & Substrate-Level Phosphorylation
Section A: Theories of Oxidative Phosphorylation
Q1. What is oxidative phosphorylation, and why is it considered the most efficient method of ATP generation?
Answer:
Oxidative phosphorylation is the process by which ATP is formed as electrons are transferred through the electron transport chain (ETC) to oxygen, coupled with the phosphorylation of ADP.
It is the most efficient because:
- It yields ~32–34 ATP molecules per glucose molecule.
- It occurs in the mitochondrial inner membrane using the proton gradient.
Q2. Describe the chemiosmotic theory of oxidative phosphorylation. Why is it widely accepted?
Answer:
The chemiosmotic theory, proposed by Peter Mitchell, states that:
- Electron transfer through ETC pumps protons (H⁺) into the intermembrane space.
- This creates a proton gradient and membrane potential (proton motive force).
- Protons flow back into the matrix via ATP synthase, driving ATP synthesis.
Why accepted:
- Supported by experimental evidence (e.g., ATP synthesis in artificial vesicles with a pH gradient)
- Explains the coupling of electron transport and ATP synthesis
Q3. Compare the chemiosmotic theory with the chemical coupling hypothesis.
Answer:
| Feature | Chemiosmotic Theory | Chemical Coupling Hypothesis |
|---|---|---|
| Mechanism | Uses proton gradient | Proposes chemical intermediates |
| ATP Synthesis | Driven by H⁺ flow through ATP synthase | Driven by energy-rich phosphorylated intermediates |
| Evidence | Strong experimental support | Largely theoretical |
| Acceptance | Widely accepted | Obsolete |
Conclusion: The chemiosmotic theory best explains mitochondrial ATP generation.
Q4. How does the binding change mechanism of ATP synthase explain ATP synthesis?
Answer:
Proposed by Paul Boyer, the binding change mechanism states:
- The F₁ subunit of ATP synthase has 3 sites that cycle through:
- Loose (L): binds ADP + Pi
- Tight (T): converts ADP + Pi to ATP
- Open (O): releases ATP
- Rotation of the γ subunit due to proton flow drives these changes.
Significance: Explains how mechanical energy from proton flow leads to chemical energy in ATP.
Q5. A patient has a mutation affecting ATP synthase. What energy-related symptoms would you expect and why?
Answer:
- Muscle weakness
- Exercise intolerance
- Lactic acidosis
Why?
ATP synthase mutation → impaired ATP production → cells switch to anaerobic glycolysis → increased lactate → energy deficit in high-demand tissues (e.g., brain, muscle).
Section B: Substrate-Level Phosphorylation (SLP)
Q6. Define substrate-level phosphorylation. How does it differ from oxidative phosphorylation?
Answer:
Substrate-level phosphorylation (SLP) is the direct transfer of a phosphate group from a high-energy substrate to ADP to form ATP, without involving the ETC.
| Feature | SLP | Oxidative Phosphorylation |
|---|---|---|
| Location | Cytosol & Mitochondria | Inner mitochondrial membrane |
| Oxygen Required | ❌ No | ✅ Yes |
| ATP Yield | Low (1 ATP per reaction) | High (up to 3 ATP per NADH) |
| Enzymes | Kinases (e.g., pyruvate kinase) | ETC complexes + ATP synthase |
Q7. Identify two reactions in human metabolism where substrate-level phosphorylation occurs.
Answer:
- Glycolysis
- Phosphoglycerate kinase:1,3-BPG + ADP → 3-PG + ATP
- Pyruvate kinase:PEP + ADP → Pyruvate + ATP
- TCA Cycle
- Succinyl-CoA synthetase:Succinyl-CoA + GDP → Succinate + GTP (→ ATP)
Q8. Why is substrate-level phosphorylation especially important in hypoxic or anaerobic conditions?
Answer:
Under low oxygen, the ETC cannot operate.
Cells rely on anaerobic glycolysis, where SLP is the only source of ATP.
Clinical relevance:
- In ischemia (e.g., MI or stroke), tissues shift to SLP for survival, but ATP yield is low → cell injury ensues.
Q9. A red blood cell has no mitochondria. How does it generate ATP?
Answer:
- RBCs depend entirely on glycolysis for ATP.
- ATP is generated through substrate-level phosphorylation during glycolysis.
Key enzymes:
- Phosphoglycerate kinase
- Pyruvate kinase
This makes SLP essential for RBC survival and ion balance (e.g., Na⁺/K⁺-ATPase function).
Q10. How can inhibition of oxidative phosphorylation lead to lactic acidosis even in the presence of oxygen?
Answer:
This condition is called histotoxic hypoxia, caused by ETC poisons (e.g., cyanide).
Mechanism:
- ETC is blocked → oxidative phosphorylation halts
- Cells switch to anaerobic glycolysis
- Excess pyruvate → lactate → lactic acidosis
Clinical presentation: Normal oxygen levels, but tissues cannot use it → mitochondrial dysfunction.