Isoleucine enzymic oxidation

In the degradation of isoleucine, (3 oxidation proceeds to completion in the normal way with generation of acetyl-CoA and propionyl-CoA. However, in the catabolism of leucine after the initial dehydrogenation in the (3-oxidation sequence, carbon dioxide is added using a biotin enzyme (Chapter 14). The double bond conjugated with the carbonyl of the thioester makes this carboxylation analogous to a standard (3-carboxylation reaction. Why add the extra C02 ... 

As indicated above, the reaction catalyzed by the general d- and l-amino acid oxidases has been represented by a dehydrogenation of an amino acid by a flavoenzyme to jneld reduced flavoenzyme and the corresponding imino acid [reactions (2) - - (4)]. Indirect support for the formation of the hypothetical imino acid has been provided by a number of studies which exclude a,/3-unsaturation in the course of the reaction. For example, it has been shown that (a) the four isomers of isoleucine are enzymically oxidized by the appropriate amino acid oxidase to the corresponding optically active a-keto-/3-methylvaleric acids (5, 6), (b) the l-isomers of /3-phenylserine are converted by L-amino acid oxidase to the respective isomers of mandelic acid (7), (c) the l- and D-isomers of a-aminophenylacetic acid, which have no /3-hydrogen atom, are attacked by the amino acid oxidases 8, 9), aod (d) on the oxidation of L-leucine in the presence of D,0 by L-amino acid oxidase, no deuterium is found in the isolated a-ketoisocaproic acid (10). More direct evidence for the formation of the a-imino acid as an intermediate has been provided by Pitt (10a) in studies on the oxidation of aromatic amino aci by ophto-n-amino acid oxidase in the presence of a tautomerase. 

Fatty acids with odd numbers of carbon atoms are rare in mammals, but fairly common in plants and marine organisms. Humans and animals whose diets include these food sources metabolize odd-carbon fatty acids via the /3-oxida-tion pathway. The final product of /3-oxidation in this case is the 3-carbon pro-pionyl-CoA instead of acetyl-CoA. Three specialized enzymes then carry out the reactions that convert propionyl-CoA to succinyl-CoA, a TCA cycle intermediate. (Because propionyl-CoA is a degradation product of methionine, valine, and isoleucine, this sequence of reactions is also important in amino acid catabolism, as we shall see in Chapter 26.) The pathway involves an initial carboxylation at the a-carbon of propionyl-CoA to produce D-methylmalonyl-CoA (Figure 24.19). The reaction is catalyzed by a biotin-dependent enzyme, propionyl-CoA carboxylase. The mechanism involves ATP-driven carboxylation of biotin at Nj, followed by nucleophilic attack by the a-carbanion of propi-onyl-CoA in a stereo-specific manner. 

Figure 9-4. Metabolism of the branched-chain amino acids. The first two reactions, transamination and oxidative decarboxylation, are catalyzed by the same enzyme in all cases. Details are provided only for isoleucine. Further metabolism of isoleucine and valine follows a common pathway to propionyl CoA. Subsequent steps in the leucine degradative pathway diverge to yield acetoacetate. An intermediate in the pathway is 3-hydroxy-3-methylglutaryl CoA (HMG-CoA), which is a precursor for cytosolic cholesterol biosynthesis.

NAD+ serves as the oxidant. The reaction is catalyzed by a complex of enzymes whose molecular mass varies from 4 to 10 x 106, depending on the species and exact substrate.297 Separate oxoacid dehydrogenase systems are known for pyruvate,298-300 2-oxoglut-arate,301 and the 2-oxoacids with branched side chains derived metabolically from leucine, isoleucine, and... 

In a rare autosomal recessive condition (discovered in 1954) the urine and perspiration has a maple syrup odor/ High concentrations of the branched-chain 2-oxoacids formed by transamination of valine, leucine, and isoleucine are present, and the odor arises from decomposition products of these acids. The branched-chain amino acids as well as the related alcohols also accumulate in the blood and are found in the urine. The biochemical defect lies in the enzyme catalyzing oxidative decarboxylation of the oxoacids, as is indicated in Fig. 24-18. Insertions, deletions, and substitutions may be present in any of the subunits (Figs. 15-14,15-15). The disease which may affect one person in 200,000, is usually fatal in early childhood if untreated. Children suffer seizures, mental retardation, and coma. They may survive on a low-protein (gelatin) diet supplemented with essential amino acids, but treatment is difficult and a sudden relapse is apt to prove fatal. Some patients respond to administration of thiamin at 20 times the normal daily requirement. The branched-chain oxoacid dehydrogenase from some of these children shows a reduced affinity for the essential coenzyme thiamin diphosphate.d... 

By this means, it has been found that the excess of L-isoleucine has two distinct effects—one that is relatively slow, and unothcr that is rapid. The slower effect is to repress production by the cell of all the enzymes required io catalyze the series of biochemical reactions in the metabolic pathway by which the cell synthesizes L-isoleucine. The Iasi effect is to inhibit production of the enzyme for the first reaction ill the series. This enzyme is L-thrconinc deaminase, which removes the amino group from L-threonine. as a preliminary step to iis oxidation and reimroduction of (he amino group, in order to produce L-isolcucine from it. 

The newest enzyme for use in beer is acetolactate decarboxylase, used to decrease the fermentation time, by avoiding the formation of diacetyl. Externally or internally produced a-acetolactate decarboxylase transforms the a-acetolactate to acetoin (acetylmethylcarbinol) without the enzyme, acetolactate goes to diacetyl, and then a secondary fermentation slowly reduces it to acetoin. Avoiding or reducing the secondary fermentation results in significant reduction in storage capacity and money tied up in inventory Q). Normally acetolactate forms by the thiaminepyrophosphate-catalyzed acyloin condensation of acetaldehyde and pyruvic acid (2) or by the condensation of two pyruvic acid molecules to yield acetolactate and CC. Acetolactate is important in the synthesis of isoleucine and valine by the yeast. The acetolactate left at the end of the primary fermentation is oxidized spontaneously in a nonenzymatic reaction to diacetvl and C0.> (Eqn. 1)... 

Cutler, A.J., Sternberg, M. and Corm, E.E. (1985) Properties of a microsomal enzyme system from Linum usitatissimum (linen flax), which oxidizes valine to acetone cyanohydrin and isoleucine to 2-methylbutanone cyanohydrin. Arch. Biochem. Biophys., 238,272-9. 

During the catabolism of fatty acids with an odd number of carbon atoms and the amino acids valine, isoleucine and threonine the resultant propionyl-CoA is converted to succinyl-CoA for oxidation in the TCA cycle. One of the enzymes in this pathway, methylmalonyl-CoA miitase, requires vitamin B12 as a cofactor in the conve sion of methylmalonyl-CoA to succinyl-CoA. The 5 -deoxyadenosine derivative of cobalamin is required for this reaction. 

Branched-Chain Oxo-acid Decarboxylase and Maple Syrup Urine Disease The third oxo-acid dehydrogenase catalyzes the oxidative decarboxylation of the hranched-chain oxo-acids that arise from the transamination of the hranched-chain amino acids, leucine, isoleucine, and valine. It has a similtu suhunit composition to pyruvate and 2-oxoglutarate dehydrogenases, and the E3 suhunit (dihydrolipoyl dehydrogenase) is the same protein as in the other two multienzyme complexes. Genetic lack of this enzyme causes maple syrup urine disease, so-called because the hranched-chain oxo-acids that are excreted in the urine have a smell reminiscent of maple syrup. 

D. The branched-chain amino acids (valine, isoleucine, and leucine) are transaminated and then oxidatively decarboxylated by an enzyme that requires thiamine, lipoic add, coenzyme A, FAD, and NAD. 

Propionyl-CoA is an intermediary product in the metabo-hsm of four essential amino acids (isoleucine, valine, threonine, and methionine), the aliphatic side-chain of cholesterol, pyrimidines (uracd and thymine), and the final product of the [3-oxidation of odd-chain fatty acids. Under normal circumstances, propionyl-CoA first is converted by a biotin-dependent carboxylase to methylmalonyi-CoA, then to succinyl-CoA by an adenosylcobalamin-dependent mutase, leading to oxidation in the tricarboxylic acid cycle. Primary or secondary defects of these two enzymes were among the first organic acidurias to be discovered, and their natural history has been characterized perhaps better than any other inborn error of organic acid metabolism. 

MCM plays an essential role in propionate metabolism. Propionate and propionyl-CoA are intermediates in the catabolism of leucine and isoleucine and are further metabolized by carboxylation of propionyl-CoA to methylmalonyl-CoA. Isomerization to succinyl-CoA feeds the carbon chain into the tricarboxylic acid pathway of oxidative metabolism. For this reason, MCM is an important enzyme in bacterial and mammalian metabolism. It is one of the two vitamin Bj2-dependent enzymes known to be important in human metabolism. 

Valine, leucine, and isoleucine - The synthetic pathway from threonine and pyruvate to valine, leucine and isoleucine is outlined in Figure 21.26. The last four reactions in the biosynthesis of valine and isoleucine are catalyzed by the same four enzymes. Threonine dehydratase, which catalyzes the first step in conversion of threonine to isoleucine, is inhibited by isoleucine. Leucine, isoleucine, and valine are all catabolized via transamination followed by oxidative decarboxylation of the respective keto-acids (see here) and oxidation. The oxidation is similar to fatty acid oxidation, except for a debranching reaction for each intermediate. 

The oxidative pathways of the BCAA convert the carbon skeleton to either suc-cinyl CoA or acetyl CoA (see Chapter 39 and Fig. 42.9). The pathways generate NADH and FAD(2H) for ATP synthesis before the conversion of carbon into intermediates of the TCA cycle, thus providing the muscle with euergy without loss of carbou as CO2. Leucine is ketogenic in that it is converted to acetyl CoA and ace-toacetate. Skeletal muscle, adipocytes, and most other tissues are able to use these products aud, therefore, directly oxidize leuciue to CO2. The portiou of isoleucine converted to acetyl CoA is also oxidized directly to CO2. For the portion of valine and isoleucine that enters the TCA cycle as succinyl CoA to be completely oxidized to CO2, it must first be converted to acetyl CoA. To form acetyl CoA, succinyl CoA is oxidized to malate in the TCA cycle, and malate is then converted to pyruvate by malic enzyme (malate + NADP pyruvate + NADPH + H ) (see Fig. 42.9). Pyruvate can then be oxidized to acetyl CoA. Alternatively, pyruvate can form alanine or lactate. 

Fig. 42.9. Metabolism of the carbon skeletons of BCAA in skeletal muscle. 1. The first step in the metabolism of BCAA is transamination (TA). 2. Carbon from valine and isoleucine enters the TCA cycle as succinyl CoA and is converted to pyruvate by decarboxylating malate dehydrogenase (malic enzyme). 3. The oxidative pathways generate NADH and FAD(2H) even before the carbon skeleton enters the TCA cycle. The rate-limiting enzyme in the oxidative pathways is the a-keto acid dehydrogenase complex. The carbon skeleton also can be converted to glutamate and alanine, shown in blue.

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