outline glycolysis as phosphorylation of glucose and the subsequent splitting of fructose 1,6-bisphosphate (6C) into two triose phosphate molecules (3C), which are then further oxidised to pyruvate (3C), with the production of ATP and reduced NAD

Respiration – Glycolysis and the Aerobic Pathway
(Cambridge International AS & A Level Biology 9700)

Why cells need respiration – ATP generated by respiration supplies the energy required for active transport, movement, macromolecular synthesis (proteins, nucleic acids) and cell division. The Cambridge syllabus (Topic 12 – Energy & Respiration) expects students to describe how the four‑stage aerobic pathway provides this ATP.

1. Overview – The Four‑Stage Aerobic Pathway

• Stage 1 – Glycolysis: cytosolic breakdown of one glucose (C₆) to two pyruvate (C₃) molecules.
• Stage 2 – Link reaction (pyruvate oxidation): conversion of each pyruvate to acetyl‑CoA in the mitochondrial matrix.
• Stage 3 – Krebs (citric‑acid) cycle: series of oxidations in the mitochondrial matrix that release CO₂ and generate reduced co‑enzymes.
• Stage 4 – Oxidative phosphorylation: electron‑transport chain (ETC) and chemiosmosis in the inner mitochondrial membrane produce the bulk of ATP.

When oxygen is limiting the pathway can terminate in anaerobic fermentation (lactate or ethanol + CO₂).

2. Glycolysis – From Glucose to Pyruvate

2.1. General features

• Location: Cytosol (no membrane barrier).
• Net products per glucose:

  Product	Yield
  ATP (substrate‑level phosphorylation)	+2 (4 produced – 2 invested)
  NADH (cytosolic)	+2 (equivalent to ≈ 3 ATP via the malate‑aspartate shuttle or ≈ 2 ATP via the glycerol‑3‑phosphate shuttle)
  Pyruvate	2 molecules

• P/O ratios (used in Stage 4): ≈ 2.5 ATP per NADH, ≈ 1.5 ATP per FADH₂.

2.2. Energy‑investment (phosphorylation) phase

1. Hexokinase (or glucokinase in liver)

   Glucose + ATP → Glucose‑6‑phosphate (G6P) + ADP

2. Phosphoglucose isomerase – G6P ⇌ fructose‑6‑phosphate (F6P).
3. Phosphofructokinase‑1 (PFK‑1) – key regulatory step

   F6P + ATP → Fructose‑1,6‑bisphosphate (F1,6BP) + ADP

Two ATP molecules are consumed per glucose.

2.3. Cleavage phase

• Aldolase splits F1,6BP into:

  • Glyceraldehyde‑3‑phosphate (G3P)
  • Dihydroxyacetone phosphate (DHAP)

• Triose‑phosphate isomerase (TPI) rapidly interconverts DHAP ⇌ G3P, ensuring that both three‑carbon molecules continue through the pathway.

2.4. Energy‑pay‑off phase (oxidation & substrate‑level phosphorylation)

4. Glyceraldehyde‑3‑phosphate dehydrogenase (GAPDH)

   G3P + NAD⁺ + Pi → 1,3‑Bisphosphoglycerate (1,3‑BPG) + NADH + H⁺

5. Phosphoglycerate kinase (PGK) (substrate‑level phosphorylation)

   1,3‑BPG + ADP → 3‑Phosphoglycerate (3‑PG) + ATP

6. Phosphoglycerate mutase – 3‑PG ⇌ 2‑Phosphoglycerate (2‑PG).
7. Enolase – dehydration of 2‑PG → phosphoenolpyruvate (PEP).
8. Pyruvate kinase (PK) (substrate‑level phosphorylation)

   PEP + ADP → Pyruvate + ATP

2.5. Regulation of glycolysis (AO2)

• PFK‑1 – allosteric control:

  • Inhibited by high ATP (energy signal) and citrate (TCA‑cycle intermediate).
  • Activated by AMP (low energy) and fructose‑2,6‑bisphosphate (hormonal regulation).

• Hexokinase – feedback inhibition by its product G6P.
• GAPDH – slowed when the NADH/NAD⁺ ratio is high (e.g., anaerobic conditions), diverting pyruvate to fermentation.

3. Link Reaction (Pyruvate Oxidation)

• Location: Mitochondrial matrix.
• Enzyme complex: Pyruvate dehydrogenase complex (PDC) – requires thiamine‑pyrophosphate, lipoic acid, CoA, FAD, NAD⁺.
• Overall reaction (per glucose):

  2 Pyruvate + 2 CoA + 2 NAD⁺ → 2 Acetyl‑CoA + 2 CO₂ + 2 NADH + 2 H⁺

• Energy yield: Each NADH ≈ 2.5 ATP in oxidative phosphorylation → ≈ 5 ATP per glucose.

4. Krebs (Citric‑Acid) Cycle

• Location: Mitochondrial matrix.
• Per acetyl‑CoA (per turn) – see table.

Per glucose (2 acetyl‑CoA): 6 NADH, 2 FADH₂, 2 GTP (≈ 2 ATP), 4 CO₂.

5. Oxidative Phosphorylation – Electron‑Transport Chain & Chemiosmosis

• Location: Inner mitochondrial membrane (ETC complexes I–IV) and the inter‑membrane space (proton gradient).
• Key principle: Electrons from NADH (Complex I) and FADH₂ (Complex II) are passed to O₂, pumping protons across the membrane. The resulting electro‑chemical gradient drives ATP synthase (Complex V).
• P/O ratios (syllabus expectation):

  • ≈ 2.5 ATP per NADH.
  • ≈ 1.5 ATP per FADH₂.

• Overall aerobic yield per glucose (typical values):

  • Glycolysis substrate‑level ATP … 2 ATP
  • Glycolytic NADH (via shuttle) … 3–5 ATP
  • Link‑reaction NADH … 5 ATP
  • Krebs cycle: 6 NADH → 15 ATP, 2 FADH₂ → 3 ATP, 2 GTP → 2 ATP
  • Total ≈ 30–32 ATP per glucose (depends on shuttle).

6. Anaerobic Fermentation (when O₂ is limiting)

6.1. Lactic‑acid fermentation (muscle cells)

1. Pyruvate + NADH → Lactate + NAD⁺ (lactate dehydrogenase).
2. Regenerates NAD⁺ so glycolysis can continue, yielding only the 2 ATP from substrate‑level phosphorylation.

6.2. Alcoholic fermentation (yeast & some plants)

1. Pyruvate → Acetaldehyde + CO₂ (pyruvate decarboxylase).
2. Acetaldehyde + NADH → Ethanol + NAD⁺ (alcohol dehydrogenase).
3. Again, only the 2 ATP from glycolysis are produced.

7. Respiratory Quotient (RQ)

RQ = CO₂ produced / O₂ consumed.

• For glucose: C₆H₁₂O₆ + 6 O₂ → 6 CO₂ + 6 H₂O → RQ = 1.0 (carbohydrate metabolism).
• Fats give RQ ≈ 0.7; proteins ≈ 0.8 – useful for interpreting respirometry data.

8. Practical Investigations (AO2 / AO3)

9. Summary Table – Stages of Aerobic Respiration

10. Flow‑Diagram (suggested illustration)