AccessScience from McGraw-Hill Education Page 1 of 6 www.accessscience.com Citric acid cycle Contributed by: Gerhard W. E. Plaut Publication year: 2014 In aerobic cells from animal and certain other species, the major pathway for the complete oxidation of acetyl coenzyme A (the thioester of acetic acid with coenzyme A); also known as the Krebs cycle or tricarboxylic acid cycle. Reduced electron carriers generated in the cycle are reoxidized by oxygen via the electron transport system; water is formed, and the energy liberated is conserved by the phosphorylation of adenosine diphosphate (ADP) to adenosine triphosphate (ATP). Reactions of the cycle also function in metabolic processes other than energy generation. The role of the cycle in mammalian tissues will be emphasized in this article. See also: ADENOSINE TRIPHOSPHATE (ATP); COENZYME; ENZYME. Reactions The first step in the cycle involves the condensation of the acetyl portion of acetyl coenzyme A (CoA) with the four-carbon compound oxaloacetate to form citrate, a tricarboxylate containing six carbons (see illustration). A shift of the hydroxyl group of citrate to an adjacent carbon results in the formation of D-threo-isocitrate, which in α turn is oxidized to the five-carbon compound -ketoglutarate and carbon dioxide (CO,2). In a second oxidative decarboxylation reaction, α-ketoglutarate, in the presence of CoA, is converted to succinyl CoA and another molecule of CO,2. In the subsequent formation of the four-carbon compound succinate and CoA, the energy in the thioester bond of succinyl CoA is conserved by the formation of guanosine triphosphate (GTP) from guanosine diphosphate (GDP) and inorganic phosphate. Fumarate is formed from succinate by the removal of two atoms of hydrogen, and the unsaturated compound is then hydrated to L-malate. The dehydrogenation of malate forms oxaloacetate, the starting four-carbon compound of the metabolic cycle. Thus, beginning with the two-carbon acetyl group, one completion of the cycle results in the formation of two molecules of carbon dioxide. The obligatory role of the intact cycle for the complete oxidation of acetyl CoA has been demonstrated by inhibiting enzymes required for specific reactions (for example, inhibition of succinate dehydrogenase by the substrate analog malonate) or by altering the regeneration or depletion of an intermediate of the cycle. Electron transport and oxidative phosphorylation The oxidation of acetyl CoA to CO,2 in the cycle occurs without direct reaction with molecular oxygen. The oxidations occur at dehydrogenation reactions in which hydrogen atoms and electrons are transferred from intermediates of the cycle to the electron carriers nicotinamide adenine dinucleotide (NAD,+) and flavin adenine dinucleotide (FAD). The NAD-specific dehydrogenase reactions are isocitrate dehydrogenase [reaction (1)], α-ketoglutarate dehydrogenase (2), and malate dehydrogenase (3). FAD is the coenzyme for succinate AccessScience from McGraw-Hill Education Page 2 of 6 www.accessscience.com UnlabelledCitric acid cycle. image dehydrogenase, (4). Unlabelled Image AccessScience from McGraw-Hill Education Page 3 of 6 www.accessscience.com Image of Chem Equation(1) 1 Unlabelled Image Image of Chem Equation(2) 2 Image of Chem(3) Equation 3 Image of Chem(4) Equation 4 The electrons from NADH and FADH,2 are transferred to molecular oxygen via a series of electron transport carriers, with regeneration of NAD,+ and FAD. The energy liberated in the electron transport chain is partially conserved by the formation of ATP from ADP and inorganic phosphate, by a process called oxidative phosphorylation. The energy generated as oxygen accepts electrons from the reduced coenzymes generated in one turn of the cycle results in the maximal formation of 11 molecules of ATP. Because GTP obtained by phosphorylation of GDP at the succinyl CoA to succinate step of the cycle is readily converted to ATP by nucleotide diphosphokinase, as in reaction (5), Image of Chem Equation 5(5) the yield is 12 molecules of ATP per molecule of acetyl CoA metabolized. See also: NICOTINAMIDE ADENINE DINUCLEOTIDE (NAD). The electron transport and oxidative phosphorylation systems and the enzymes required for the citric acid cycle are located in the mitochondria of cells. These mitochondrial systems are the major source of ATP for energy-consuming reactions in most tissues. The citric acid cycle does not occur in all cells. For example, mature human red blood cells do not contain mitochondria and the cycle is absent. In these cells, ATP is formed by the anaerobic conversion of glucose to lactate (anaerobic glycolysis). See also: MITOCHONDRIA; PHOSPHATE METABOLISM. AccessScience from McGraw-Hill Education Page 4 of 6 www.accessscience.com Regulation A major rate-limiting step of the citric acid cycle in aerobic mammalian tissues (for example, heart) is at NAD-specific isocitrate dehydrogenase [reaction (1)]. The activity of the enzyme is dependent on the concentration of the substrate (magnesium isocitrate): it is activated by ADP, calcium (Ca,2+), and citrate, and it is inhibited by NADH and reduced nicotinamide adenine dinucleotide phosphate (NADPH). Differences in energy demand affect the rate of citric acid flux and concentrations of modulators of NAD-dependent isocitrate dehydrogenase. At rest, the flux through the cycle is slowed, the cellular concentration of the isocitrate dehydrogenase inhibitor NADH is raised; and the activator ADP is lowered; the opposite occurs during high energy demand. The rate of cycle oxidation in liver mitochondria increases with decreased NADPH concentration; increased mitochondrial free Ca,2+ enhances cycle flux. Formation of acetyl CoA and intermediates Acetyl CoA is formed from carbohydrates, fats, and the carbon skeleton of amino acids. The origin of a precursor and the extent of its utilization depend on the metabolic capability of a specific tissue and on the physiological state of the organism. For example, most mammalian tissues have the capacity to convert glucose to pyruvate in a reaction called glycolysis. Pyruvate is then taken up from cellular cytosol by mitochondria and oxidatively decarboxylated to acetyl CoA and carbon dioxide by pyruvate dehydrogenase, as in reaction (6). Unlabelled Image Image of Chem Equation(6 6) Acetyl CoA is also the end product of fatty acid oxidation in mitochondria. However, the fatty acid oxidation pathway occurs in fewer tissues than does glycolysis or the citric acid cycle. For example, nervous tissue utilizes the complete oxidation of glucose but not of fatty acids to maintain energy needs. In prolonged starvation, the level of blood glucose declines to a concentration inadequate to support the optimal energy needs of brain, whereas the level of blood ketone bodies (formed from fatty acids in liver) rises. In this case, the oxidation of ketone bodies (acetoacetate and 3-hydroxybutyrate) via acetyl CoA supplements the diminished oxidation of glucose to fulfill the energy requirements of the brain. The amino acids follow varied pathways for forming compounds that can enter the citric acid cycle. For example, the paths of degradation of leucine and lysine lead directly to formation of acetyl CoA; only part of the carbons of aromatic amino acids and of isoleucine are converted to acetyl CoA. Pyruvate, formed by transamination of AccessScience from McGraw-Hill Education Page 5 of 6 www.accessscience.com alanine with the citric acid cycle intermediate α-ketoglutarate [reaction (7)], Image of Chem(7) Equation 7 can be oxidatively decarboxylated to acetyl CoA by pyruvate dehydrogenase [reaction (6)] or carboxylated to the cycle intermediate oxaloacetate by pyruvate carboxylase [reaction (8)]. Image of Chem(8) Equation 8 Oxaloacetate can also be formed by aspartate aminotransferase [reaction (9)]. Unlabelled Image Image of Chem Equation 9 (9) Portions of the carbon skeletons of valine, isoleucine, methionine, and threonine are converted to methylmalonyl CoA, which rearranges to the citric acid cycle intermediate succinyl CoA. See also: AMINO ACIDS. Role in lipogenesis and gluconeogenesis In addition to the cycle’s role in yielding catabolic energy, portions of it can supply intermediates for synthetic processes, such as the synthesis of the fatty acid moiety of triglycerides from glucose (lipogenesis), and formation of glucose from the carbon skeletons of certain amino acids, lactate, or glycerol (gluconeogenesis). Lipogenesis. When dietary intake exceeds the energy needs of the body, the excess calories are deposited as body fat. Under these conditions in humans, the synthesis of fatty acids from glucose occurs mainly in the liver. It is favored by metabolic changes resulting principally from a rise in blood insulin: (1) The rate of pyruvate formation increases because of the rise in tissue glucose and enhancement of glycolysis. (2) Increases in enzyme activities catalyzing the conversion of pyruvate to acetyl CoA at pyruvate dehydrogenase [reaction (6)] and to oxaloacetate at pyruvate carboxylase [reaction (8)] raise the concentrations of both of these substrates for citrate synthesis. When this occurs at a rate exceeding that of the citric acid cycle, the citrate concentration rises and citrate exits from the mitochondria to the cytosol. (3) In the cytosol, citrate is cleaved to oxaloacetate and acetyl CoA in a reaction requiring CoA and ATP. The acetyl CoA is converted to palmitate and other long-chain fatty acids in a series of condensation steps requiring CO,2, ATP as energy source, and NADPH as