Citric acid cycle

The citric acid cycle, also known as the tricarboxylic acid cycle (TCA cycle) or the Krebs cycle, (On rare occasions the citric acid cycle is known by a fourth name, the Szent-Györgyi-Krebs cycle) is a series of Hans Krebs.

In fermentation.

Overview

Two carbons are coenzyme Q, an intermediate in the electron transfer chain.^[1]

The citric acid cycle is continuously supplied new carbons in the form of acetyl-CoA, entering at step 1 below.^[2]

S t e p	Substrates	Products	Enzyme	Reaction type	Comment
1	Acetyl CoA + H₂O	CoA-SH	Citrate synthase	Aldol condensation	rate limiting stage, extends the 4C oxaloacetate to a 6C molecule
2	Citrate	cis-Aconitate + H₂O	Aconitase	Dehydration	reversible isomerisation
3	cis-Aconitate + H₂O	Isocitrate	Aconitase	Hydration	reversible isomerisation
4	Isocitrate + NAD⁺	Oxalosuccinate + NADH + H ⁺	Isocitrate dehydrogenase	Oxidation	generates NADH (equivalent of 3 ATP)
5	Oxalosuccinate	α-Ketoglutarate + CO₂	Isocitrate dehydrogenase	Decarboxylation	irreversible stage, generates a 5C molecule
6	α-Ketoglutarate + NAD⁺ + CoA-SH	Succinyl-CoA + NADH + H⁺ + CO₂	α-Ketoglutarate dehydrogenase	Oxidative decarboxylation	generates NADH (equivalent of 3 ATP), regenerates the 4C chain (CoA excluded)
7	Succinyl-CoA + GDP + P_i	Succinate + CoA-SH + GTP	Succinyl-CoA synthetase	substrate level phosphorylation	or ADP->ATP,^[1] generates 1 ATP or equivalent
8	Succinate + ubiquinone (Q)	Fumarate + ubiquinol (QH₂)	Succinate dehydrogenase	Oxidation	uses FAD as a prosthetic group (FAD->FADH₂ in the first step of the reaction) in the enzyme,^[1] generates the equivalent of 2 ATP
9	Fumarate + H₂O	L-Malate	Fumarase	H₂O addition (hydration)
10	L-Malate+ NAD⁺	Oxaloacetate + NADH + H⁺	Malate dehydrogenase	Oxidation	generates NADH (equivalent of 3 ATP)

Mitochondria in animals including humans possess two succinyl-CoA synthetases, one that produces GTP from GDP, and another that produces ATP from ADP.^[3] Plants have the type that produces ATP (ADP-forming succinyl-CoA synthetase).^[2]

The GTP that is formed by GDP-forming succinyl-CoA synthetase may be utilized by nucleoside-diphosphate kinase to form ATP (the catalyzed reaction is GTP + ADP -> GDP + ATP).^[1]

A simplified view of the process

The citric acid cycle begins with acetyl-CoA transferring its two-carbon acetyl group to the four-carbon acceptor compound (oxaloacetate) to form a six-carbon compound (citrate).
The citrate then goes through a series of chemical transformations, losing first one, then a second carboxyl group as CO₂. The carbons lost as CO₂ originate from what was oxaloacetate, not directly from acetyl-CoA. The carbons donated by acetyl-CoA become part of the oxaloacetate carbon backbone after the first turn of the citric acid cycle. Loss of the acetyl-CoA-donated carbons as CO₂ requires several turns of the citric acid cycle. However, because of the role of the citric acid cycle in anabolism, they may not be lost since many TCA cycle intermediates are also used as precursors for the biosynthesis of other molecules.^[4]
Most of the energy made available by the oxidative steps of the cycle is transferred as energy-rich electrons to NAD⁺, forming NADH. For each acetyl group that enters the citric acid cycle, three molecules of NADH are produced.
Electrons are also transferred to the electron acceptor FAD, forming FADH₂.
At the end of each cycle, the four-carbon oxaloacetate has been regenerated, and the cycle continues.

Products

Products of the first turn of the cycle are: one GTP, three NADH, one FADH₂, two CO₂.

Because two acetyl-CoA molecules are produced from each glucose molecule, two cycles are required per glucose molecule. Therefore, at the end of all cycles, the products are: two GTP, six NADH, two FADH₂, and four CO₂

Description	Reactants	Products
The sum of all reactions in the citric acid cycle is:	Acetyl-CoA + 3 NAD⁺ + FAD + GDP + P_i + 2 H₂O	→ CoA-SH + 3 NADH + 3 H⁺ + FADH₂ + GTP + 2 CO₂
Combining the reactions occurring during the pyruvate oxidation with those occurring during the citric acid cycle, the following overall pyruvate oxidation reaction is obtained:	Pyruvic acid + 4 NAD⁺ + FAD + GDP + P_i + 2 H₂O	→ 4 NADH + 4 H⁺ + FADH₂ + GTP + 3 CO₂
Combining the above reaction with the ones occurring in the course of glycolysis, the following overall glucose oxidation reaction (excluding reactions in the respiratory chain) is obtained:	Glucose + 10 NAD⁺ + 2 FAD + 2 ADP + 2 GDP + 4 P_i + 2 H₂O	→ 10 NADH + 10 H⁺ + 2 FADH₂ + 2 ATP + 2 GTP + 6 CO₂

(the above reactions are equilibrated if P_i represents the H₂PO₄^- ion, ADP and GDP the ADP^2- and GDP^2- ions, respectively, and ATP and GTP the ATP^3- and GTP^3- ions, respectively).

Considering the future conversion of GTP to ATP and the maximum 32 ATP produced by the 10 NADH and the 2 FADH₂ (see the theoretical yields for cellular respiration), it follows that each glucose molecule is able to produce a maximum of 32 ATP.

Regulation

Although pyruvate dehydrogenase is not technically a part of the citric acid cycle, its regulation is included here.

The regulation of the TCA cycle is largely determined by substrate availability and product inhibition. NADH, a product of all dehydrogenases in the TCA cycle with the exception of allosteric effector whose concentration changes less than 10% ^[5].

Calcium is used as a regulator. It activates α-ketoglutarate dehydrogenase.^[6] This increases the reaction rate of many of the steps in the cycle, and therefore increases flux throughout the pathway.

Citrate is used for feedback inhibition, as it inhibits phosphofructokinase, an enzyme involved in fructose 1,6-bisphosphate, a precursor of pyruvate. This prevents a constant high rate of flux when there is an accumulation of citrate and a decrease in substrate for the enzyme.

Recent work has demonstrated an important link between intermediates of the citric acid cycle and the regulation of E3 ubiquitin ligase complex which targets them for rapid degradation. This reaction is calalysed by prolyl 4-hydroxylases. Fumarate and succinate have been identified as potent inhibitors of prolyl hydroxylases thus leading to the stabilisation of HIF.^[7]

Major metabolic pathways converging on the TCA cycle

Most of the body's catabolic pathways converge on the TCA cycle, as the diagram shows. Reactions that form intermediates of the TCA cycle in order to replenish them (especially during the scarcity of the intermediates) are called anaplerotic reactions.

The citric acid cycle is the third step in carbohydrate catabolism (the breakdown of sugars). Glycolysis breaks glucose (a six-carbon-molecule) down into pyruvate (a three-carbon molecule). In eukaryotes, pyruvate moves into the decarboxylation and enters the citric acid cycle.

In amino acids can become a source of energy by being converted to Acetyl-CoA and entering into the citric acid cycle.

In beta oxidation which results in acetyl-CoA which can be used in the citric acid cycle. Sometimes beta oxidation can yield propionyl CoA which can result in further glucose production by gluconeogenesis in the liver.

The citric acid cycle is always followed by oxidative phosphorylation. This process extracts the energy (as electrons) from NADH and FADH₂, oxidizing them to NAD⁺ and FAD, respectively, so that the cycle can continue. Whereas the citric acid cycle does not use oxygen, oxidative phosphorylation does.

The total energy gained from the complete breakdown of one molecule of glucose by glycolysis, the citric acid cycle and oxidative phosphorylation equals about 36 ATP molecules. The citric acid cycle is called an amphibolic pathway because it participates in both anabolism.

References

^ ^a ^b ^c ^d Berg, JM; JL Tymoczko, L Stryer (2002). Biochemistry - 5th Edition. WH Freeman and Company, 465-484, 498-501. ISBN 0-7167-4684-0.
^ ^a ^b Buchanan; Gruissem, Jones (2000). Biochemistry & molecular biology of plants, 1st Edition, American society of plant physiology. ISBN 0-943088-39-9.
^ Johnson JD, Mehus JG, Tews K, Milavetz BI, Lambeth DO (1998). "Genetic evidence for the expression of ATP- and GTP-specific succinyl-CoA synthetases in multicellular eucaryotes". J Biol Chem 273 (42): 27580-6.
^ Wolfe RR, Jahoor F. (1990) Recovery of labeled CO₂ during the infusion of C-1- vs C-2-labeled acetate: implications for tracer studies of substrate oxidation. Am J Clin Nutr. 51(2):248-52. PMID 2106256
^ Voet, D. & Voet, J. G. (2004) Biochemistry 3rd Edition (John Wiley & Sons, Inc., New York) p. 615
^ Denton RM; Randle PJ, Bridges BJ, Cooper RH, Kerbey AL, Pask HT, Severson DL, Stansbie D, Whitehouse S. (Oct 1975). "Regulation of mammalian pyruvate dehydrogenase". Mol Cell Biochem 9 (1): 27-53.
^ Koivunen P, Hirsilä M, Remes AM, Hassinen IE, Kivirikko KI, Myllyharju J (2007). "Inhibition of hypoxia-inducible factor (HIF) hydroxylases by citric acid cycle intermediates: possible links between cell metabolism and stabilization of HIF". J. Biol. Chem. 282 (7): 4524–32. PMID 17182618.

Neil A. Campbell; Jane B. Reece (Dec 2005). Biology, 7th ed., Benjamin Cummings. ISBN 978-0805371468.
Solomon, E.P.; Berg, L.R., Martin, D.W. (Mar 2005). Biology. Brooks Cole. ISBN 978-0534495480.