UNIT 2.2.2

The Krebs Cycle (Citric Acid Cycle)

Completing the oxidation of organic molecules

🎯 After this unit, you will be able to:

Describe the location and overview of the Krebs cycle
Explain the key steps where NADH, FADH₂, and ATP are produced
Calculate the energy yield per acetyl-CoA and per glucose
Understand the role of the Krebs cycle in plant metabolism

🔄 What is the Krebs Cycle?

The Krebs cycle (also called the citric acid cycle or tricarboxylic acid cycle) is the second stage of cellular respiration. It takes place in the mitochondrial matrix (the fluid-filled space inside mitochondria) and completes the oxidation of organic molecules, producing reducing power (NADH and FADH₂) and a small amount of ATP .

Acetyl-CoA + 3 NAD⁺ + FAD + ADP + Pi → 2 CO₂ + 3 NADH + FADH₂ + ATP + CoA

Key concept: The Krebs cycle is named after Hans Krebs, who discovered it in 1937 (and won a Nobel Prize). It's a true cycle—the starting molecule (oxaloacetate) is regenerated at the end, allowing it to turn continuously .

🧪 Did you know? Hans Krebs discovered the cycle while working in Germany, but had to flee to England because he was Jewish. He completed his work at the University of Sheffield, and the cycle was initially called the "citric acid cycle" or "tricarboxylic acid cycle" .

📍 Location and Preparation

Where Does It Happen?

The Krebs cycle occurs in the mitochondrial matrix—the semi-fluid substance inside the inner mitochondrial membrane. This is where all the enzymes of the cycle are located .

🔬 [Diagram: Mitochondrion showing matrix, cristae, and location of Krebs cycle — to be inserted]

Getting Ready: Pyruvate → Acetyl-CoA

Before entering the Krebs cycle, pyruvate (from glycolysis) must be converted to acetyl-CoA. This reaction is catalyzed by the pyruvate dehydrogenase complex, a huge multi-enzyme complex .

Pyruvate + CoA + NAD⁺ → Acetyl-CoA + CO₂ + NADH

This step is irreversible and highly regulated. It produces:

1 CO₂ (first carbon removed from glucose)
1 NADH
1 Acetyl-CoA (2 carbons) that enters the cycle

For each glucose molecule (which produced 2 pyruvate), this step happens twice, producing 2 NADH before the cycle even begins .

📋 The Krebs Cycle: Step by Step

The Krebs cycle consists of 8 enzyme-catalyzed steps. Here are the key reactions where energy carriers are produced:

🔄 [Diagram: Complete Krebs cycle showing 8 steps with enzymes and products — to be inserted]

Step 1

Citrate formation

Acetyl-CoA (2C) + oxaloacetate (4C) → citrate (6C). Catalyzed by citrate synthase.

Step 3

First oxidation

Isocitrate → α-ketoglutarate. Produces 1 NADH and releases 1 CO₂.

Step 4

Second oxidation

α-ketoglutarate → succinyl-CoA. Produces 1 NADH and releases 1 CO₂.

Step 5

Substrate-level ATP

Succinyl-CoA → succinate. Produces 1 ATP (or GTP in some organisms).

Step 6

Third oxidation

Succinate → fumarate. Produces 1 FADH₂ (via succinate dehydrogenase).

Step 8

Fourth oxidation

Malate → oxaloacetate. Produces 1 NADH, regenerating the starting molecule.

💰 Energy Accounting: Per Acetyl-CoA

For each acetyl-CoA that enters the Krebs cycle, we get:

Product	Quantity per acetyl-CoA	From which steps
NADH	3	Steps 3, 4, and 8
FADH₂	1	Step 6
ATP (or GTP)	1	Step 5
CO₂	2	Steps 3 and 4

Remember: Each glucose produces 2 acetyl-CoA (after glycolysis and pyruvate oxidation), so these numbers double when accounting for one glucose molecule .

📊 Complete Accounting: From Glucose through Krebs

Let's track all energy carriers from one glucose molecule through glycolysis, pyruvate oxidation, and the Krebs cycle:

Stage	ATP (net)	NADH	FADH₂
Glycolysis	2	2	0
Pyruvate oxidation (2×)	0	2	0
Krebs cycle (2×)	2	6	2
TOTAL per glucose	4 ATP	10 NADH	2 FADH₂

These NADH and FADH₂ molecules will now enter the electron transport chain to produce much more ATP through oxidative phosphorylation .

⚡ Did you know? The 10 NADH and 2 FADH₂ from one glucose will ultimately produce about 30-32 ATP in the electron transport chain—most of the energy from glucose! Glycolysis and Krebs directly produce only 4 ATP .

🎛️ Regulation of the Krebs Cycle

The Krebs cycle is tightly regulated to match the cell's energy needs:

Substrate availability: Needs acetyl-CoA and oxaloacetate
Feedback inhibition: High ATP and NADH inhibit key enzymes (isocitrate dehydrogenase, α-ketoglutarate dehydrogenase)
Calcium signaling: In animals (less so in plants), calcium activates several enzymes
Product inhibition: Accumulation of products slows the cycle

🌿 Plant-Specific Regulation

In plants, the Krebs cycle is also regulated by light/dark cycles and interacts with photosynthesis. During the day, mitochondria may run the Krebs cycle differently, using alternative oxidases and pathways to balance energy and carbon needs .

Some plant enzymes have isoforms that are light-regulated, connecting mitochondrial metabolism to chloroplast function .

🔄 The Krebs Cycle is Amphibolic

The Krebs cycle is amphibolic—it functions in both catabolism (breaking down molecules) and anabolism (building molecules). Intermediates are siphoned off for biosynthesis:

🧬 Biosynthetic roles

Citrate → fatty acid synthesis
α-ketoglutarate → amino acids (glutamate)
Succinyl-CoA → chlorophyll synthesis
Oxaloacetate → aspartate, other amino acids

⚡ Energy roles

Complete oxidation of acetyl-CoA
Production of NADH and FADH₂
Supply reducing power for ATP synthesis

When intermediates are removed for biosynthesis, they must be replenished by anaplerotic reactions (like PEP carboxylase fixing CO₂ into oxaloacetate) .

🧑‍🌾 Horticultural Implications

Fruit Ripening and Senescence

During fruit ripening and senescence, respiration rates change dramatically. The Krebs cycle activity affects:

Climacteric fruits: A burst of respiration (including Krebs cycle activity) accompanies ripening
Senescence: Mitochondrial function declines, reducing energy availability

Post-Harvest Storage

Controlled atmosphere storage (low O₂, high CO₂) slows the Krebs cycle by:

Reducing substrate availability (less pyruvate from glycolysis)
Direct inhibition of some Krebs enzymes by CO₂
Slowing NADH re-oxidation in the electron transport chain

Respiratory Quotient (RQ)

The respiratory quotient (CO₂ released / O₂ consumed) indicates which substrates are being oxidized:

RQ ≈ 1.0: Carbohydrates (glucose) — typical for most fruits
RQ > 1.0: Organic acids (citrus, apples during storage)
RQ < 1.0: Fats or proteins (seeds, nuts)

🍎 Measuring RQ in Apple Storage

Apple storage facilities monitor CO₂ production and O₂ consumption to calculate RQ. A sudden increase in RQ might indicate that apples are switching from carbohydrate oxidation to organic acid oxidation—a sign of stress or senescence. This helps managers adjust storage conditions .

🔗 Connections to Other Pathways

With Photosynthesis: During the day, mitochondria in photosynthetic tissues may run a truncated Krebs cycle, exporting intermediates for nitrogen assimilation and other processes. The photorespiratory pathway also connects to the Krebs cycle through glycine metabolism .

With Nitrogen Metabolism: α-ketoglutarate is the carbon skeleton for glutamate synthesis, the entry point for ammonium assimilation. This connects the Krebs cycle directly to nitrogen metabolism in plants .

With Fatty Acid Oxidation: When seeds germinate, stored fats are converted to acetyl-CoA (via β-oxidation), which enters the Krebs cycle to provide energy and carbon skeletons for the growing seedling .

🔗 [Diagram: Krebs cycle connections to other metabolic pathways — to be inserted]

📌 Unit Summary

Krebs cycle occurs in mitochondrial matrix, oxidizes acetyl-CoA (from pyruvate)
Per acetyl-CoA: produces 3 NADH, 1 FADH₂, 1 ATP, and 2 CO₂
Per glucose (after glycolysis + pyruvate oxidation): 10 NADH, 2 FADH₂, 4 ATP (from Krebs + glycolysis)
Cycle is amphibolic—provides intermediates for biosynthesis
Regulated by energy status (ATP, NADH)
Horticultural relevance: Fruit ripening, post-harvest storage, respiratory quotient monitoring

Reflection question: A post-harvest physiologist measures the respiratory quotient of stored apples and finds it has increased from 1.0 to 1.2 over several weeks. What does this tell you about what's happening inside the apples? What adjustments might be made to storage conditions?

📌 Key terms introduced

Krebs cycle Citric acid cycle Mitochondrial matrix Acetyl-CoA Pyruvate dehydrogenase NADH FADH₂ Substrate-level phosphorylation Amphibolic Anaplerotic reactions Respiratory quotient (RQ)

✅ Check your understanding

Where in the cell does the Krebs cycle occur?
For each acetyl-CoA that enters the Krebs cycle, how many NADH, FADH₂, and ATP are produced?
From one glucose molecule, what is the total yield of NADH and FADH₂ after glycolysis, pyruvate oxidation, and the Krebs cycle?
Explain what "amphibolic" means and give an example of a Krebs cycle intermediate used for biosynthesis.
Why might a post-harvest facility monitor the respiratory quotient of stored fruits?

Discuss your answers in the course forum.

Plant Biochemistry for Horticulture · HORT 202 · Dilla University · Last updated March 2026