Acetoacetic acid, activation coenzyme

In extraliepatic tissues, d-/3-hydroxybutyrate is oxidized to acetoacetate by o-/3-hydroxybutyrate dehydrogenase (Fig. 17-19). The acetoacetate is activated to its coenzyme A ester by transfer of CoA from suc-cinyl-CoA, an intermediate of the citric acid cycle (see Fig. 16-7), in a reaction catalyzed by P-ketoacyl-CoA transferase. The acetoacetyl-CoA is then cleaved by thiolase to yield two acetyl-CoAs, which enter the citric acid cycle. Thus the ketone bodies are used as fuels. 

As the study of CoA developed, it became apparent that the coenzyme was involved in reactions whereby acetate was activated by ATP and subsequently transferred to various acetyl acceptors. In pigeon liver extracts it was shown that acetate could be activated by ATP in the presence of CoA to acetylate sulfanilamide, PABA, histamine, glucosamine, to synthesize acetoacetic acid and citrate. Acetyl phosphate, which has been demonstrated to be a product of pyruvate metabolism in several bacteria and could theoretically be considered to be an intermediate in these reactions, was found to be unable to replace acetate and ATP in animal tissues. Eventually it was shown that there is present in certain bacteria an enzyme, phosphotransacetylase, which could convert acetyl phosphate to a reactive product which was thought to be acetyl-CoA.i 194 isolation of acetyl-CoA from yeast extract by Lynen and Reichert confirmed the idea that acetyl-CA is the reactive 2-carbon unit in these reactions. Stadtman has demonstrated that acetyl-CoA is indeed the product of the action of phosphotransacetylase. Lipmann has recently... 

Isotope experiments of Weinhouse, Medes, and Floyd and of Buchanan, Sakami, and Gurin, have shown that acetoacetic acid and other /3-ketonic acids can supply active acetic acid for the synthesis of citric acid. Since the synthesis of citrate requires acetyl coenzyme A, the breakdown of 8-ketonic acids must lead to the formation of acetyl coenzyme A. Lynen and Reichert assumed a thioclastic fission, as formulated in Scheme 8. Observations which support this... 

The activating group, coenzyme A, may be lost from acetoacetyl-CoA. Hydrolysis was formerly assumed, but now it seems more probable that the CoA moiety is transferred onto succinate, forming succinyl-CoA. Actually, however, the liberation of free acetoacetic acid appears to proceed primarily via a detour, an attempt at synthesis ... 

Biotin is a growth factor for many bacteria, protozoa, plants, and probably all higher animals. In the absence of biotin, oxalacetate decarboxylation, oxalosuccinate carboxylation, a-ketoglutarate decarboxylation, malate decarboxylation, acetoacetate synthesis, citrulline synthesis, and purine and pyrimidine syntheses, are greatly depressed or absent in cells (Mil, Tl). All of these reactions require either the removal or fixation of carbon dioxide. Together with coenzyme A, biotin participates in carboxylations such as those in fatty acid and sterol syntheses. Active C02 is thought to be a carbonic acid derivative of biotin involved in these carboxylations (L10, W10). Biotin has also been involved in... 

In nature, the biologically active form of acetic acid is acetyl-coenzyme A (acetyl-CoA) (see Box 7.18). Two molecules of acetyl-CoA may combine in a Claisen-type reaction to produce acetoacetyl-CoA, the biochemical equivalent of ethyl acetoacetate. This reaction features as the start of the sequence to mevalonic acid (MVA), the precursor in animals of the sterol cholesterol. Later, we shall see another variant of this reaction that employs malonyl-CoA as the nucleophile (see Box 10.17). 

In muscle, most of the fatty acids undergoing beta oxidation are completely oxidized to C02 and water. In liver, however, there is another major fate for fatty acids this is the formation of ketone bodies, namely acetoacetate and b-hydroxybutyrate. The fatty acids must be transported into the mitochondrion for normal beta oxidation. This may be a limiting factor for beta oxidation in many tissues and ketone-body formation in the liver. The extramitochondrial fatty-acyl portion of fatty-acyl CoA can be transferred across the outer mitochondrial membrane to carnitine by carnitine palmitoyltransferase I (CPTI). This enzyme is located on the inner side of the outer mitochondrial membrane. The acylcarnitine is now located in mitochondrial intermembrane space. The fatty-acid portion of acylcarnitine is then transported across the inner mitochondrial membrane to coenzyme A to form fatty-acyl CoA in the mitochondrial matrix. This translocation is catalyzed by carnitine palmitoyltransferase II (CPTII Fig. 14.1), located on the inner side of the inner membrane. This later translocation is also facilitated by camitine-acylcamitine translocase, located in the inner mitochondrial membrane. The CPTI is inhibited by malonyl CoA, an intermediate of fatty-acid synthesis (see Chapter 15). This inhibition occurs in all tissues that oxidize fatty acids. The level of malonyl CoA varies among tissues and with various nutritional and hormonal conditions. The sensitivity of CPTI to malonyl CoA also varies among tissues and with nutritional and hormonal conditions, even within a given tissue. Thus, fatty-acid oxidation may be controlled by the activity and relative inhibition of CPTI. 

In normal liver, only relatively small amounts of ketone bodies are formed. Their concentration in the blood is 0.5-D.8 mg per 100 ml plasma. The acetoacetate produced by this physiological K. is degraded in the peripheral musculature. Coenzyme A from succi-nyl-CoA is transferred to the acetoacetate by aceto-acetate succinyl-CoA transferase. Direct activation of acetoacetate by coenzyme A and ATP can also occur (Fig, 2). The acetoacetyl-CoA produced in either case is thioclastically cleaved into two molecules of acetyl-CoA, consuming a CoA molecule in the process. In carbohydrate deficiency (starvation, ketone-mia in ruminants), or deficient carbohydrate utilization (diabetes mellitus), K. is greatly increased. The cause of this pathological K. is a disturbance of the equilibrium between the degradation of fatty acids to acetyl-CoA and its utilization in the tricarboxylic acid cycle. The several-fold increase in the oxidation of the fatty acids leads under these conditions to an increase in the intracellular acetyl-CoA concentration. This leads to the condensation of 2 molecules of... 

Coenzyme A was discovered by Lipmann in 1945 as necessary for the acetylation of sulfanilamide by pigeon liver extracts. Shortly thereafter this same coenzyme was identified as the activator of choline acetylation earlier observed by Nachmansohn and Berman, as well as Feldberg and Mann. Subsequently, work from Lipmann s laboratory, as well as from other laboratories, extended the role of CoA to a large variety of transacetylation reactions, i.e., acetylation of aromatic amines, synthesis of acetylcholine, of citrate, of acetoacetate, of fatty acids, of sterols, and of phospholipids. ... 

The oxidation of acetate, acetoacetate, and fatty acids by a nonmito-chondrial system from pig heart has been described. The heart system consists of three parts (1) a particulate nonmitochondrial fraction (2) a group of soluble enzymes and (3) a coenzyme concentrate, incompletely characterized, but which is known to contain di- and triphospho-pyridine nucleotides, coenzyme A, and ATP. The system reflects many of the properties of intact mitochondria and lends itself to a study of the reaction sequences in the activation and oxidation of fatty acids. 

---