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Krebs cycle

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Columbia Encyclopedia

Krebs cycle, series of chemical reactions carried out in the living cell; in most higher animals, including humans, it is essential for the oxidative metabolism of glucose and other simple sugars. The breakdown of glucose to carbon dioxide and water is a complex set of chemical interconversions called carbohydrate catabolism, and the Krebs cycle is the second of three major stages in the process, occurring between glycolysis and oxidative phosphorylation. This cycle, also known as the citric acid cycle, was named in recognition of the German chemist Hans Krebs, whose research into the cellular utilization of glucose contributed greatly to the modern understanding of this aspect of metabolism. The name citric acid cycle is derived from the first product generated by the sequence of conversions, i.e., citric acid. The reactions are seen to comprise a cycle inasmuch as citric acid is both the first product and the final reactant, being regenerated at the conclusion of one complete set of chemical rearrangements. Citric acid is a so-called tricarboxylic acid, containing three carboxyl groups (COOH). Hence the Krebs cycle is sometimes referred to as the tricarboxylic acid (TCA) cycle. The Krebs cycle begins with the condensation of one molecule of a compound called oxaloacetic acid and one molecule of acetyl CoA (a derivative of coenzyme A; see coenzyme). The acetyl portion of acetyl CoA is derived from pyruvic acid, which is produced by the degradation of glucose in glycolysis. After condensation, the oxaloacetic acid and acetyl CoA react to produce citric acid, which serves as a substrate for seven distinct enzyme-catalyzed reactions that occur in sequence and proceed with the formation of seven intermediate compounds, including succinic acid, fumaric acid, and malic acid. Malic acid is converted to oxaloacetic acid, which, in turn, reacts with yet another molecule of acetyl CoA, thus producing citric acid, and the cycle begins again. Each turn of the citric acid cycle produces, simultaneously, two molecules of carbon dioxide and eight atoms of hydrogen as byproducts. The carbon dioxide generated is an ultimate end product of glucose breakdown and is removed from the cell by the blood. The hydrogen atoms are donated as hydride ions to the system of electron transport molecules, which allow for oxidative phosphorylation. In most higher plants, in certain microorganisms, such as the bacterium Escherichia coli, and in the algae, the citric acid cycle is modified to a form called the glyoxylate cycle, so named because of the prominent intermediate, glyoxylic acid.

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Wikipedia

The citric acid cycle, also known as the tricarboxylic acid cycle (TCA cycle) or the Krebs cycle, (or rarely, the Szent-Györgyi-Krebs cycle) is a series of enzyme-catalysed chemical reactions of central importance in all living cells that use oxygen as part of cellular respiration. In eukaryotes, the citric acid cycle occurs in the matrix of the mitochondrion. The components and reactions of the citric acid cycle were established by seminal work from both Albert Szent-Györgyi and Hans Krebs.

In aerobic organisms, the citric acid cycle is part of a metabolic pathway involved in the chemical conversion of carbohydrates, fats and proteins into carbon dioxide and water to generate a form of usable energy. Other relevant reactions in the pathway include those in glycolysis and pyruvate oxidation before the citric acid cycle, and oxidative phosphorylation after it. In addition, it provides precursors for many compounds including some amino acids and is therefore functional even in cells performing fermentation.

Overview

Two carbons are oxidized to CO₂, and the energy from these reactions is transferred to other metabolic processes by GTP (or ATP), and as electrons in NADH and QH₂. The NADH generated in the TCA cycle may later donate its electrons in oxidative phosphorylation to drive ATP synthesis; FADH₂ is covalently attached to succinate dehydrogenase, an enzyme functioning both in the TCA cycle and the mitochondrial electron transport chain in oxidative phosphorylation. FADH₂ thereby facilitates transfer of electrons to coenzyme Q, which is the final electron acceptor of the reaction catalyzed by the Succinate:ubiquinone oxidoreductase complex, also acting as an intermediate in the electron transport chain.

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

	Substrates	Products	Enzyme	Reaction type	Comment
1	Oxaloacetate + Acetyl CoA + H₂O	Citrate + 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 2.5 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 2.5 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, 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, generates the equivalent of 1.5 ATP
9	Fumarate + H₂O	L-Malate	Fumarase	H₂O addition (hydration)
10	L-Malate+ NAD⁺	Oxaloacetate + NADH + H⁺	Malate dehydrogenase	Oxidation	generates NADH (equivalent of 2.5 ATP)

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

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).

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.
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 Q, forming QH₂.
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 (or ATP), three NADH, one QH₂, 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 QH₂, and four CO₂''

Description	Reactants	Products
The sum of all reactions in the citric acid cycle is:	Acetyl-CoA + 3 NAD⁺ + Q + GDP + P_i + 2 H₂O	→ CoA-SH + 3 NADH + 3 H⁺ + QH₂ + 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⁺ + Q + GDP + P_i + 2 H₂O	→ 4 NADH + 4 H⁺ + QH₂ + 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 Q + 2 ADP + 2 GDP + 4 P_i + 2 H₂O	→ 10 NADH + 10 H⁺ + 2 QH₂ + 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).

Estimates for the total number of ATP obtained after complete oxidation of one glucose in glycolysis, citric acid cycle, and oxidative phosphorylation given in the literature range from 30-38 molecules of ATP. A recent assessment of the total ATP yield obtained in these distinct reaction cycles, taking into account updated proton-to-ATP ratios, has arrived at an estimate of 29.85 ATP per glucose molecule.

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 succinate dehydrogenase, inhibits pyruvate dehydrogenase, isocitrate dehydrogenase and α-ketoglutarate dehydrogenase, and also citrate synthase. Acetyl-CoA inhibits pyruvate dehydrogenase, while succinyl-CoA inhibits succinyl-CoA synthase and citrate synthase. When tested in vitro with TCA enzymes, ATP inhibits citrate synthase and α-ketoglutarate dehydrogenase; however, ATP levels do not change more than 10% in vivo between rest and vigorous exercise. There is no known allosteric mechanism that can account for large changes in reaction rate from an allosteric effector whose concentration changes less than 10% .

Calcium is used as a regulator. It activates pyruvate dehydrogenase, isocitrate dehydrogenase and α-ketoglutarate dehydrogenase. 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 glycolysis that catalyses formation of 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 hypoxia inducible factors (HIF). HIF plays a role in the regulation of oxygen haemostasis, and is a transcription factor which targets angiogenesis, vascular remodelling, glucose ulitisation, iron transport and apoptosis. HIF is synthesized consititutively and hydroxylation of at least one of two critical proline residues mediates their interaction with the von Hippel Lindau 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.

Major metabolic pathways converging on the TCA cycle

Several catabolic pathways converge on the TCA cycle. 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 mitochondria. It is converted into acetyl-CoA by decarboxylation and enters the citric acid cycle.

In protein catabolism, proteins are broken down by protease enzymes into their constituent amino acids. The carbon backbone of these amino acids can become a source of energy by being converted to Acetyl-CoA and entering into the citric acid cycle.

In fat catabolism, triglycerides are hydrolyzed to break them into fatty acids and glycerol. In the liver the glycerol can be converted into glucose via dihydroxyacetone phosphate and glyceraldehyde-3-phosphate by way of gluconeogenesis. In many tissues, especially heart tissue, fatty acids are broken down through a process known as beta oxidation which results in acetyl-CoA which can be used in the citric acid cycle. Beta oxidation of odd chain fatty acids 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 QH₂, oxidizing them to NAD⁺ and Q, 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 30 ATP molecules, in eukaryotes. The citric acid cycle is called an amphibolic pathway because it participates in both catabolism and anabolism.