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8 Steps of Citric acid Cycle (Krebs cycle) and Enzymes involved in each Step

The Krebs cycle, Citric acid cycle or TCA cycle is an eight step cyclic reactions in which acetyl CoA is oxidized producing CO2, reduced coenzymes (NADH + H+ and FADH2), and ATP.

Site of Reaction: Mitochondrial matrix in Eukaryotes

                              Cytoplasm in Prokaryotes

All enzymes are present in mitochondrial matrix except succinate dehydrogenase which is bound to inner mitochondrial membrane

Pyruvate formed in Glycolysis enters mitochondrion and is converted to acetyl CoA which enters Krebs cycle.

8 Steps of Citric acid Cycle (Krebs cycle) and Enzymes involved in each Step

Step 1: Condensation reaction

Acetyl CoA (C2) is added to oxaloacetate (C4) to form citrate (C6).

Step1 Enzyme: Citrate synthase

Coenzyme A (CoA-SH) is removed in the process.

Step 2: Isomerization

Citrate (C6) is isomerized forming isocitrate (C6).

Step2 Enzyme: Aconitase

2 step process: First step involves dehydration of citrate to cis-aconitase (unstable product)

Second step involving rehydration of cis-aconitase into isocitrate.


Step 3: Dehydrogenation (1st NADH synthesis) and decarboxylation (reduction in C atom: C6 to C5)

Isocitrate (C6) is converted into alphaketoglutarate (C5).

Step3 Enzyme: Isocitrate dehydrogenase

2 step process: First step isocitrate is dehydrogenated to oxalosuccinate (C6) (intermediate product) NAD+ is reduced, forming NADH + H+

Second step: Decarboxylation of oxalosuccinate(C6) to α-ketoglutarate (C5). Carbondioxide is removed.


Step 4: Oxidative decarboxylation (2nd NADH synthesis)

(reduction in C atom: C5 to C4)

α-ketoglutarate (C5) is oxidatively decarboxylated to form succinyl-CoA (C4), high energy compound.

Step 4 Enzyme: α-ketoglutarate dehydrogenase enzyme complex

During this step coenzyme A is added, CO2 is removed, and NAD+ is reduced, forming NADH + H+


Step 5: Substrate level phosphorylation (GTP/ATP synthesis)

Succinyl CoA(C4) is converted to succinate (C4)

Step 5 Enzyme: succinyl-CoA synthase

Coenzyme A is released, Substrate level phosphorylation of guanosine diphosphate (GDP) to guanosine triphosphate (GTP). GTP is then hydrolyzed to form ATP


Step 6: Dehydrogenation (FADH2 synthesis)

Succinate(C4) is converted to fumarate (C4).

Step 6 Enzyme: succinate dehydrogenase in inner mitochondrial membrane

During this step, FAD is reduced forming FADH2.


Step 7: Hydration

Fumarate(C4) is converted to malate(C4) with the addition of water.

Step 7 Enzyme: Fumarase (Fumaric acid hydratase)


Step 8: Dehydrogenation (3rd NADH synthesis)

Final step: Malate (C4) is converted to oxaloacetate(C4)

Step 8 Enzyme: Malate dehydrogenase

In the process, NAD+ is reduced to form NADH + H+.

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