Citric acid cycle

The citric acid cycle—also known as the Krebs cycle, Szent–Györgyi–Krebs cycle, or TCA cycle (tricarboxylic acid cycle)^[1]^[2]—is a series of biochemical reactions that release the energy stored in nutrients through acetyl-CoA oxidation. The energy released is available in the form of ATP. The Krebs cycle is used by organisms that generate energy via respiration, either anaerobically or aerobically (organisms that ferment use different pathways). In addition, the cycle provides precursors of certain amino acids, as well as the reducing agent NADH, which are used in other reactions. Its central importance to many biochemical pathways suggests that it was one of the earliest metabolic components.^[3]^[4] Even though it is branded as a "cycle", it is not necessary for metabolites to follow a specific route; at least three alternative pathways of the citric acid cycle are recognized.^[5]

Its name is derived from the citric acid (a tricarboxylic acid, often called citrate, as the ionized form predominates at biological pH^[6]) that is consumed and then regenerated by this sequence of reactions. The cycle consumes acetate (in the form of acetyl-CoA) and water and reduces NAD⁺ to NADH, releasing carbon dioxide. The NADH generated by the citric acid cycle is fed into the oxidative phosphorylation (electron transport) pathway. The net result of these two closely linked pathways is the oxidation of nutrients to produce usable chemical energy in the form of ATP.

In eukaryotic cells, the citric acid cycle occurs in the mitochondrial matrix. In prokaryotic cells, such as bacteria, which lack mitochondria, the citric acid cycle reaction sequence is performed in the cytosol with the proton gradient for ATP production being across the cell's surface (plasma membrane) rather than the inner membrane of the mitochondrion.

For each pyruvate molecule (from glycolysis), the overall yield of energy-containing compounds from the citric acid cycle is three NADH, one FADH₂, and one GTP or ATP.^[7]

Discovery

Several of the components and reactions of the citric acid cycle were established in the 1930s by the research of Albert Szent-Györgyi, who received the Nobel Prize in Physiology or Medicine in 1937 specifically for his discoveries pertaining to fumaric acid, a component of the cycle.^[8] He made this discovery by studying pigeon breast muscle. Because this tissue maintains its oxidative capacity well after breaking down in the Latapie mincer and releasing in aqueous solutions, breast muscle of the pigeon was very well qualified for the study of oxidative reactions.^[9] The citric acid cycle itself was finally identified in 1937 by Hans Adolf Krebs and William Arthur Johnson while at the University of Sheffield,^[10]^[11] for which the former received the Nobel Prize for Physiology or Medicine in 1953, and for whom the cycle is sometimes named the "Krebs cycle".^[12] Independently citric acid cycle was identified in 1937 by German biochemists Carl Martius and Franz Knoop.

Overview

The citric acid cycle is a metabolic pathway that connects carbohydrate, fat, and protein metabolism. The reactions of the cycle are carried out by eight enzymes that completely oxidize acetate (a two carbon molecule), in the form of acetyl-CoA, into two molecules each of carbon dioxide. Through catabolism of sugars, fats, and proteins, the two-carbon organic product acetyl-CoA is produced which enters the citric acid cycle. The reactions of the cycle also convert three equivalents of nicotinamide adenine dinucleotide (NAD⁺) into three equivalents of reduced NAD (NADH), one equivalent of flavin adenine dinucleotide (FAD) into one equivalent of FADH₂, and one equivalent each of guanosine diphosphate (GDP) and inorganic phosphate (P_i) into one equivalent of guanosine triphosphate (GTP). The NADH and FADH₂ generated by the citric acid cycle are, in turn, used by the oxidative phosphorylation pathway to generate energy-rich ATP.

One of the primary sources of acetyl-CoA is from the breakdown of sugars by glycolysis which yield pyruvate that in turn is decarboxylated by the pyruvate dehydrogenase complex generating acetyl-CoA according to the following reaction scheme:

+ HSCoA + NAD⁺ → + NADH + CO₂

The product of this reaction, acetyl-CoA, is the starting point for the citric acid cycle. Acetyl-CoA may also be obtained from the oxidation of fatty acids. Below is a schematic outline of the cycle:

The citric acid cycle begins with the transfer of a two-carbon acetyl group from acetyl-CoA 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 two carboxyl groups 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 might not be lost, since many citric acid cycle intermediates are also used as precursors for the biosynthesis of other molecules.^[13]
Most of the electrons made available by the oxidative steps of the cycle are transferred to NAD⁺, forming NADH. For each acetyl group that enters the citric acid cycle, three molecules of NADH are produced. The citric acid cycle includes a series of redox reactions in mitochondria.^{[clarification needed]}^[14]
In addition, electrons from the succinate oxidation step are transferred first to the FAD cofactor of succinate dehydrogenase, reducing it to FADH₂, and eventually to ubiquinone (Q) in the mitochondrial membrane, reducing it to ubiquinol (QH₂) which is a substrate of the electron transfer chain at the level of Complex III.
For every NADH and FADH₂ that are produced in the citric acid cycle, 2.5 and 1.5 ATP molecules are generated in oxidative phosphorylation, respectively.
At the end of each cycle, the four-carbon oxaloacetate has been regenerated, and the cycle continues.

Steps

There are ten basic steps in the citric acid cycle, as outlined below. The cycle is continuously supplied with new carbon in the form of acetyl-CoA, entering at step 0 in the table.^[15]

	Reaction type	Substrates	Enzyme	Products	Comment
0 / 10	Aldol condensation	Oxaloacetate + Acetyl CoA + H₂O	Citrate synthase	Citrate + CoA-SH	irreversible, extends the 4C oxaloacetate to a 6C molecule
1	Dehydration	Citrate	Aconitase	cis-Aconitate + H₂O	reversible isomerisation
2	Hydration	cis-Aconitate + H₂O	Aconitase	Isocitrate	reversible isomerisation
3	Oxidation	Isocitrate + NAD⁺	Isocitrate dehydrogenase	Oxalosuccinate + NADH + H ⁺	generates NADH (equivalent of 2.5 ATP)
4	Decarboxylation	Oxalosuccinate	Isocitrate dehydrogenase	α-Ketoglutarate + CO₂	rate-limiting, irreversible stage, generates a 5C molecule
5	Oxidative decarboxylation	α-Ketoglutarate + NAD⁺ + CoA-SH	α-Ketoglutarate dehydrogenase, Thiamine pyrophosphate, Lipoic acid, Mg++,transsuccinytase	Succinyl-CoA + NADH + H ⁺ + CO₂	irreversible stage, generates NADH (equivalent of 2.5 ATP), regenerates the 4C chain (CoA excluded)
6	Substrate-level phosphorylation	Succinyl-CoA + GDP + P_i	Succinyl-CoA synthetase	Succinate + CoA-SH + GTP	or ADP→ATP instead of GDP→GTP,^[16] generates 1 ATP or equivalent. Condensation reaction of GDP + P_i and hydrolysis of succinyl-CoA involve the H₂O needed for balanced equation.
7	Oxidation	Succinate + ubiquinone (Q)	Succinate dehydrogenase	Fumarate + ubiquinol (QH₂)	uses FAD as a prosthetic group (FAD→FADH₂ in the first step of the reaction) in the enzyme.^[16] These two electrons are later transferred to QH₂ during Complex II of the ETC, where they generate the equivalent of 1.5 ATP
8	Hydration	Fumarate + H₂O	Fumarase	L-Malate	Hydration of C-C double bond
9	Oxidation	L-Malate + NAD⁺	Malate dehydrogenase	Oxaloacetate + NADH + H⁺	reversible (in fact, equilibrium favors malate), generates NADH (equivalent of 2.5 ATP)
10 / 0	Aldol condensation	Oxaloacetate + Acetyl CoA + H₂O

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