Cellular Respiration

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Senior 6 · Biology

Cellular Respiration

5 lessons

Lesson 3 of 5

The Krebs cycle and oxidative phosphorylation

25 min

Where does most of the ATP come from?

After glycolysis and the link reaction, one glucose has given only 2 ATP. Yet a cell with oxygen makes about fifteen times as much. The rest comes from the last two stages, both inside the mitochondrion: the Krebs cycle, which strips the remaining hydrogen and carbon from the acetyl groups, and oxidative phosphorylation, which uses that hydrogen to drive a molecular turbine, ATP synthase. Oxygen is needed only at the very end, but without it the whole system stops.

Definition

Krebs cycle (citric acid cycle): a cycle of reactions in the mitochondrial matrix that oxidises acetyl groups to CO₂, reducing NAD and FAD and making a little ATP.

Decarboxylation: removal of carbon dioxide from a molecule. Dehydrogenation: removal of hydrogen.

Electron transport chain (ETC): a series of carrier proteins in the inner mitochondrial membrane that pass electrons from reduced NAD and reduced FAD to oxygen, releasing energy in small steps.

Chemiosmosis: the synthesis of ATP using the energy of a gradient of H⁺ ions flowing through ATP synthase across a membrane (Peter Mitchell, 1961).

The Krebs cycle

Figure 1: One turn of the Krebs cycle, for one acetyl group. The cycle turns twice per glucose.

The 2-carbon acetyl group from acetyl CoA joins oxaloacetate (4C) to form citrate (6C). Coenzyme A is released to collect another acetyl group.
Citrate is decarboxylated and dehydrogenated to a 5-carbon compound: one CO₂ and one reduced NAD.
The 5-carbon compound is decarboxylated and dehydrogenated to a 4-carbon compound: one CO₂, one reduced NAD, and one ATP made by substrate-level phosphorylation.
The 4-carbon compound is converted back to oxaloacetate in further dehydrogenations: one reduced FAD and one reduced NAD.

Per turn: 2 CO₂, 3 reduced NAD, 1 reduced FAD, 1 ATP. The cycle turns twice per glucose, once for each acetyl group. Oxaloacetate is regenerated, so a small amount keeps the cycle running.

Oxidative phosphorylation

Figure 2: The electron transport chain and chemiosmosis on the inner mitochondrial membrane (simplified).

Reduced NAD and reduced FAD deliver hydrogen atoms to the electron transport chain on the inner membrane (the cristae). Each hydrogen atom splits into a proton (H⁺) and an electron.
The electrons pass from carrier to carrier, each at a lower energy level. The energy released is used by the carriers to pump H⁺ ions from the matrix into the intermembrane space.
H⁺ ions build up in the narrow intermembrane space, creating an electrochemical gradient (a difference in both concentration and charge) across the inner membrane, which is impermeable to H⁺.
H⁺ ions can flow back into the matrix only through ATP synthase. As they pass, the enzyme rotates and joins ADP and $\mathrm{P_i}$ to make ATP: chemiosmosis.
At the end of the chain, oxygen is the final electron acceptor: it combines with electrons and H⁺ ions to form water. Without oxygen, the carriers stay reduced, the chain stops, NAD and FAD are not regenerated, and the Krebs cycle and link reaction stop too.

Reduced FAD passes its hydrogen to the chain at a later carrier than reduced NAD, so fewer H⁺ ions are pumped and it yields less ATP.

Counting the ATP

Two sets of numbers are used. The traditional count assumes 3 ATP for each reduced NAD and 2 for each reduced FAD; many exams, and many textbooks, still use it. Careful measurements show the real yields are lower, about 2.5 and 1.5, because the H⁺ gradient is also used to move pyruvate, ADP and phosphate into the mitochondrion, and some H⁺ leaks back across the membrane.

Table 1: ATP from one molecule of glucose

Stage	ATP made directly	Reduced NAD	Reduced FAD
Glycolysis (cytoplasm)	2 (net)	2	0
Link reaction $\times 2$ (matrix)	0	2	0
Krebs cycle $\times 2$ (matrix)	2	6	2
Total	4	10	2

Traditional count: $4 + (10 \times 3) + (2 \times 2) = 4 + 30 + 4 = 38$ ATP.

Modern estimate: $4 + (10 \times 2.5) + (2 \times 1.5) = 4 + 25 + 3 = 32$ ATP, reduced to about 30 in cells where the reduced NAD from glycolysis enters the mitochondrion by a less efficient shuttle.

Either way, about 90 % of the ATP comes from oxidative phosphorylation.

Worked example 1: what share of the ATP comes from each stage?

Using the traditional count of 38 ATP per glucose, calculate the percentage made (a) by substrate-level phosphorylation, (b) by oxidative phosphorylation, and (c) from the reduced NAD of the Krebs cycle alone.

(a) Substrate-level: 4 ATP. $\frac{4}{38} \times 100 = 10.5\,\%$ .
(b) Oxidative: $38 - 4 = 34$ ATP. $\frac{34}{38} \times 100 = 89.5\,\%$ .
(c) Krebs cycle reduced NAD: $6 \times 3 = 18$ ATP. $\frac{18}{38} \times 100 = 47.4\,\%$ .

Conclusion: almost nine tenths of the ATP depends on oxygen, through the electron transport chain.

Worked example 2: how cyanide kills

Cyanide binds to the last carrier of the electron transport chain (cytochrome oxidase) and stops it passing electrons to oxygen. Explain why a person poisoned by cyanide stops using oxygen, and why death follows within minutes.

Electrons cannot leave the chain, so every carrier stays reduced and no more electrons are accepted from reduced NAD or reduced FAD.
No H⁺ ions are pumped, the gradient disappears, and ATP synthase stops: oxidative phosphorylation ends.
With no oxidised NAD and FAD returned to the matrix, the link reaction and Krebs cycle stop too. Oxygen is not used, even though the blood is full of it.
Only glycolysis can continue, giving 2 ATP per glucose instead of about 30 to 38. Tissues with high energy demand, such as the brain and heart, fail within minutes.

Warning

Common mistakes

"Oxygen is needed in the Krebs cycle." Oxygen is used only at the end of the electron transport chain. The Krebs cycle stops without oxygen only because NAD and FAD are not regenerated.
"ATP synthase pumps H⁺." The carriers pump H⁺ out; ATP synthase lets H⁺ flow back, and uses that flow.
"The CO₂ we breathe out comes from oxygen." The oxygen atoms you breathe in end up in water; the CO₂ comes from the carbon of the food, released in the link reaction and Krebs cycle.

Cognito: "A-Level Biology - Oxidative Phosphorylation". Watch for the electron transport chain, the pumping of H⁺ ions, ATP synthase and the ATP yield, and compare the numbers with Table 1.

Tip

Memory hook for one turn of the Krebs cycle: "3-1-1-2": 3 reduced NAD, 1 reduced FAD, 1 ATP, 2 CO₂. Double everything for one glucose. For the electron transport chain, remember the order: electrons flow, protons are pumped, protons return through ATP synthase, oxygen accepts the electrons.

✦

Practice: the Krebs cycle and oxidative phosphorylation

Name the 4-carbon compound that accepts the acetyl group in the Krebs cycle, and the 6-carbon compound it forms.
For one glucose molecule, state the number of (a) CO₂ molecules released in the Krebs cycle, (b) reduced NAD formed in the Krebs cycle.
Explain why the inner mitochondrial membrane is folded into cristae.
Explain the role of oxygen in aerobic respiration.
Using the modern values (2.5 ATP per reduced NAD, 1.5 per reduced FAD), calculate the ATP yield from the Krebs cycle products of one glucose, including the ATP made directly.
The drug 2,4-dinitrophenol (DNP) makes the inner membrane leaky to H⁺. Predict and explain its effects on (a) ATP production, (b) oxygen use, (c) body temperature.
Explain why reduced FAD yields less ATP than reduced NAD.

Show answersHide answers

Oxaloacetate (4C); citrate (6C).
(a) 4 (2 per turn × 2 turns). (b) 6 (3 per turn × 2 turns).
To give a large surface area for many electron transport chains and ATP synthase molecules, so more ATP can be made.
Oxygen is the final electron acceptor: it combines with electrons and H⁺ at the end of the chain to form water, keeping electrons flowing so that H⁺ can be pumped and NAD and FAD regenerated.
Krebs products per glucose: 2 ATP, 6 reduced NAD, 2 reduced FAD: $2 + (6 \times 2.5) + (2 \times 1.5) = 2 + 15 + 3 = 20$ ATP.
(a) ATP production falls: H⁺ leaks back without passing through ATP synthase, so the gradient is lost. (b) Oxygen use rises or continues: the chain keeps running, often faster, since nothing holds it back. (c) Body temperature rises: the energy of the gradient is released as heat instead of being trapped in ATP. (DNP was once sold as a slimming drug and caused deaths from overheating.)
Reduced FAD passes its electrons to the chain at a later carrier, so they pass fewer H⁺-pumping carriers, fewer H⁺ ions are pumped, and less ATP is made.

Key points

Krebs cycle (matrix), per turn: 2 CO₂, 3 reduced NAD, 1 reduced FAD, 1 ATP; twice per glucose.
Electron transport chain (inner membrane): electrons flow to oxygen, H⁺ is pumped into the intermembrane space, and H⁺ flowing back through ATP synthase makes ATP (chemiosmosis).
Per glucose: 4 ATP directly, 10 reduced NAD, 2 reduced FAD; total 38 ATP (traditional) or about 30 to 32 (modern).

Sources and credits

2.4.4: Oxidation of Pyruvate and the Citric Acid Cycle, from Biology 2e by OpenStax (Mary Ann Clark, Matthew Douglas, Jung Choi), via LibreTexts Biology, licensed under CC BY 4.0. Used for the Krebs cycle and its products. Adapted by Ganzaa.
2.4.5: Oxidative Phosphorylation, from Biology 2e by OpenStax (Mary Ann Clark, Matthew Douglas, Jung Choi), via LibreTexts Biology, licensed under CC BY 4.0. Used for the electron transport chain, chemiosmosis, ATP yield and the effects of poisons. Adapted by Ganzaa.
Video: A-Level Biology - Oxidative Phosphorylation, Cognito, embedded from YouTube.

Figures 1 and 2, Table 1, the worked examples and the practice questions are written by Ganzaa.

1Where does most of the ATP come from?

2The Krebs cycle

3Oxidative phosphorylation

4Counting the ATP

Where does most of the ATP come from?

The Krebs cycle

Oxidative phosphorylation

Counting the ATP