Citric Acid Cycle Enzymes
The citric acid cycle, or CAC, was first proposed in 1937 by British biochemist Sir Hans Adolf Krebs. He acquired the 1953 Nobel Prize in Physiology or Medicine for his work, which elucidated most of the reactions in the cycle. However, his design had some gaps, so coenzyme A’s discovery was essential to fully unravel the process. Read on to learn about the enzymes responsible for these reactions and what they do.
Citrate synthase is one of eight enzymes involved in the citric acid cycle. It catalyzes the first step of the citric acid cycle, a chemical reaction that uses glucose to form alcohol called citrate. Glycolysis breaks glucose down into two carbon atoms, one of which is released as carbon dioxide. The remaining carbon atoms are carried inactivated form on special cofactor molecules and fully oxidized into carbon dioxide. Citrate synthase attaches acetate to oxaloacetate, which acts as a convenient handle for carbon atoms that are passed from one enzyme to the next.
In humans, citrate provides precursors for fatty acid synthesis. It also regulates glycolysis by negative modulation of phosphofructokinase activity. It exits the mitochondria by way of the tricarboxylate carrier. However, in bacteria, the citric acid cycle is not reversed in all cases. Nevertheless, some bacteria do manage to reverse the cycle.
This multi-enzyme complex comprises two subunits, CS and mMDH. CS catalyzes reversible NAD(H)-dependent conversion of l-malate to citrate and acetyl-CoA. Aco catalyzes the dehydration-rehydration of citrate to iso-citrate and the formation of cis-aconitate.
The action of citrate synthase is highly ordered. The enzyme is formed of two identical subunits, one of which starts in the open forum and then closes around its substrates, performing a linkage reaction. It then converts the substrates to acetyl-CoA. These products are then stored as glycogen. A newer study has revealed that the enzyme may also participate in cellular respiration.
The citrate synthase enzyme is present in all animals, plants, fungi, and archaebacteria. It consists of two monomeric subunits, each with a single amino acid chain containing 437 amino acids. These domains are connected via four pairs of helices. Hence, this enzyme is found in a spherical structure.
The first step in the citric acid cycle involves the conversion of pyruvate into acetyl-CoA. During the second step, acetyl-CoA binds to HIS 320 and hydrolyzes the oxaloacetate to form citrate. Then, citryl-coA is released from the enzyme through the PDH reaction.
It catalyzes the dehydrogenation of succinate and generates the reduced cofactor FADH2, which is covalently attached to the enzyme. The enzyme also possesses an iron-sulfur cluster that accepts electrons from FADH2 and passes them on to ubiquinone. This enzyme is part of the succinate-Q-reductase complex, which is associated with the inner mitochondrial membrane proteins. It forms an important link in the electron transport chain and is involved in the Krebs cycle.
The citric acid cycle consists of four steps, the first four of which involve two carbon atoms: acetyl and carbon dioxide. The remaining reactions use four carbon atoms from the succinyl group to resynthesize oxaloacetate, which combines with an incoming acetyl group.
Succinyl-CoA synthase activity was determined by using a purified bacterial succinyl-CoA synthetase. It hydrolyzes ATP, GTP, and phosphate, which increases its activity. Its activity in the presence of phosphate is the most significant of all the three enzymes.
Succinyl-CoA is an important part of the citric acid cycle. It converts propionyl-CoA to succinate in a three-step process. It then uses the oxidizing agent flavin adenine dinucleotide (FAD) to form succinate. The citric acid cycle enzymes are necessary for the production of ethanol.
It is vital to understand how the citric acid cycle works. The enzymes involved in the citric acid cycle must be activated to produce citric acid, and without these enzymes, the synthesis of ethanol will be impossible. Besides fructose, succinyl-CoA is a precursor to acetyl-CoA.
Succinyl-CoA synthase (SCS) catalyzes substrate-level phosphorylation in the forward direction. The enzyme replenishes the citric acid supply required for ketone body catabolism and porphyrin biosynthesis. The SCS protein consists of two isoforms: the a-subunit and the b subunit. Both determine the specificity of nucleotide phosphorylation and the direction of the reactions.
Primary succinate dehydrogenase deficiency results in tumors and tissue degeneration. This infection is caused by a mutation in the fumarase gene and is commonly found in leiomyomas. TCAC dysfunction is also caused by concurrent impairments in several steps of the cycle. For example, combined deficiencies of SDH and aconitase are associated with Friedreich’s ataxia.
This enzyme plays a major function in the metabolism of glucose. It is responsible for converting a large portion of glucose into acetyl-CoA. Pyruvate dehydrogenase facilitates the process by binding to pyruvate. This enzyme consists of three subunits and requires at least five cofactors to perform its function. In addition to being an essential regulator of glucose metabolism, pyruvate dehydrogenase is also required to produce acetyl-CoA.
Several studies have shown that fumarase and succinate dehydrogenase mutations act as tumor suppressors. Defects in fumarase or succinate dehydrogenase are associated with dominantly inherited uterine fibroids and skin leiomyomas. Mutations in fumarase and succinate dehydrogenase have been linked to a wide range of tumors, including brain and lung cancer. Studies of yeast have shown similar responses to mutations in isocitrate dehydrogenase. In addition, the relationship between isocitrate dehydrogenase dysfunction and genetic instability is consistent between human and yeast strains.
Isocitrate oxidase, also known as aconitase, is a crucial component of the citric acid cycle. The enzyme catalyzes the formation of one nmol of isocitrate per minute. O2* is measured to determine the rate constant of reactivation of aconitase.
The CAC is a series of steps in the metabolism of sugars. The first step involves the production of acetyl-CoA, which is the end product of glycolysis and b-oxidation. The second step, called oxidation, occurs when citrate is reduced to a lower level of acetyl-CoA, and the third step, isocitrate oxidase, is required to produce glutamate.
The other three TCA cycle enzymes, CIT1, FUM1, and iDH1, are encoded by the same gene, which is also found in two different genes: IDH1 and iDH2a. They have similar expression profiles and are the most responsive to TCA cycle defects. Some of the TCA cycle genes are also required for the synthesis of glutamate and a-ketoglutarate.
Guanosine diphosphate synthetase
The a-ketoglutarate molecule undergoes oxidative decarboxylation and is transferred from the citric acid’s 3′ to 4′ position by succinyl coenzyme A. The enzyme then catalyzes the oxidation of the isocitrate’s 4′ -OH group. This produces GTP and NAD+, which are both high-energy compounds.
The adenosine diphosphate molecule and oxaloacetate combine in the enzyme. The oxaloacetate group is stripped off of the CoA group, and the molecule is converted into a six-carbon molecule known as citrate. The citrate molecule undergoes an oxidation and rearrangement step, transferring electrons to NAD+ and releasing carbon dioxide.
The CAC is an important component of aerobic energy pathways. A-ketoglutarate is produced during the catabolism of sugars, fats, and amino acids. These two molecules then undergo the citric acid cycle, generating two equivalents of ATP and carbon dioxide. After these reactions, oxaloacetate is regenerated, ready to start the cycle again. NADH and FADH2 are then used in the final stages of cellular respiration, where they generate large amounts of ATP.
When this reaction takes place, the a-ketoglutarate is decarboxylated into succinyl-CoA, and another molecule of NAD+ is reduced to NADH. The enzyme catalyzes the conversion of the alpha-ketoglutarate to succinyl-CoA. The reaction is irreversible, as producing acetyl-CoA requires carbon dioxide.
The CAC is a sequence of chemical reactions that release energy from stored carbohydrates, fats, and proteins. The citric acid cycle breaks down the acetate and produces NADH, reduced nicotinamide adenine dinucleotide (NADH), and amino acids. The cycle is the primary metabolic pathway for aerobic processes in animal tissue. It takes place within the mitochondrial matrix in eukaryotes and prokaryotes’ cytosol. The first reaction is the oxidation of acetyl-CoA, and the other reaction is the conversion of oxaloacetate to citrate.
The 2nd in the cycle is the synthesis of GTP. GDP and ADP are formed by sequential oxidation of acetyl carbons in the Krebs cycle. The electron transport chain then synthesizes ATP. Finally, guanosine triphosphate (GTP) is produced as the unoxidized product of the entire cycle.