Mutases, coenzymes

Coenzyme B12-Dependent Mutases Coenzyme B12-Dependent Eliminases Methyltransferases... 

R)-Methylmalonyl CoA mutase (coenzyme Bi2"d P dent), methylmalonyl CoA racemase ... 

Adenosylcobalamin (coenzyme B 2) is required in a number of rearrangement reactions that occurring in humans is the methylmalonyl-Co A mutase-mediated conversion of (R)-methylmalonyl-Co A (6) to succinjl-CoA (7) (eq. 1). The mechanism of this reaction is poorly understood, although probably free radical in nature (29). The reaction is involved in the cataboHsm of valine and isoleucine. In bacterial systems, adenosylcobalamin drives many 1,2-migrations of the type exemplified by equation 1 (30). 

There is one exception to the rule that requires bulky hydrophobic residues to fill the interior of eight-stranded a/p barrels in order to form a tightly packed hydrophobic core. The coenzyme Biz-dependent enzyme methylmalonyl-coenzyme A mutase, the x-ray structure of which was determined by Phil Evans and colleagues at the MRC Laboratory of Molecular... 

Figure 4.4 Schematic diagram of the structure of the a/p-barrel domain of the enzyme methylmalonyl-coenzyme A mutase. Alpha helices are red, and p strands are blue. The inside of the barrel is lined by small hydrophilic side chains (serine and threonine) from the p strands, which creates a hole in the middle where one of the substrate molecules, coenzyme A (green), binds along the axis of the barrel from one end to the other. (Adapted from a computer-generated diagram provided by P. Evans.)...

Mancia, E., et al. How coenzyme Biz radicals are generated the crystal structure of methylmalonyl-coen-zyme A mutase at 2 A resolution. Strueture 4 339-350, 1996. 

Methylmalonyl-CoA mutase 5 -deoxyadenosylco-balamin is part of dimethylbenzimidazolecobamide coenzyme, a constituent of methylmalonyl-CoA mutase. This mutase catalyses the isomerization of methylmalonyl-CoA to succinyl-CoA (anaplerotic reaction of the citric acid cycle). 

Ratnatilleke A, JW Vrijbloed, JA Robinson (1999) Cloning and sequencing of the coenzyme B,2-binding domain of isobutyryl-CoA mutase from Streptomyces cinnamonensis. Reconstitution of mutase activity and characterization of the recombinant enzyme produced in Escherichia coli. J Biol Chem 274 31679-31685. 

In mammals and in the majority of bacteria, cobalamin regulates DNA synthesis indirectly through its effect on a step in folate metabolism, catalyzing the synthesis of methionine from homocysteine and 5-methyltetrahydrofolate via two methyl transfer reactions. This cytoplasmic reaction is catalyzed by methionine synthase (5-methyltetrahydrofolate-homocysteine methyl-transferase), which requires methyl cobalamin (MeCbl) (253), one of the two known coenzyme forms of the complex, as its cofactor. 5 -Deoxyadenosyl cobalamin (AdoCbl) (254), the other coenzyme form of cobalamin, occurs within mitochondria. This compound is a cofactor for the enzyme methylmalonyl-CoA mutase, which is responsible for the conversion of T-methylmalonyl CoA to succinyl CoA. This reaction is involved in the metabolism of odd chain fatty acids via propionic acid, as well as amino acids isoleucine, methionine, threonine, and valine. 

Carboxylation of propionyl-CoA is accomplished by propionyl-CoA carboxylase (biotin, which is the carboxyl group carrier, serves as a coenzyme for this enzyme) the presence of ATP is also required. The methylmalonyl-CoA formed is converted by methylmalonyl-CoA mutase (whose coenzyme, deoxyadenosylcobalamin, is a derivative of vitamin B]2) to succinyl-CoA the latter enters the Krebs cycle. 

Biochemical findings are variable. The blood cobala-min and folate levels often are normal. Patients often have homocysteinemia with hypomethioninemia, the latter finding discriminating this group from homocystinuria secondary to cystathionine- P-synthase deficiency. Urinary excretion of methylmalonic acid may be high, reflecting the fact that vitamin B12 serves as a cofactor for the methyl-malonyl-CoA (coenzyme A) mutase reaction. 

Hydrogen Radical Transfer Catalyzed by a Coenzyme Bn-Dependent Mutase... 

Table 11.10 Transmission coefficients and their effect on KIE for H/D radical transfer at 278 K catalyzed by a coenzyme B -dependent mutase (Dybala-Defratyka, A., Paneth, P., Baneijee, R. and Truhlar, D. G., Proc. Natl. Acad. 5c/.104, 10774 (2007) Chowdhury, S. and Baner-jee, R.,./. Am. Chem. Soc. 122, 5417 (2000)) ...

Various coenzymes are involved in these reactions. The carboxylase [3] requires biotin, and the mutase [4] is dependent on coenzyme Bj2 (5 -deoxyadenosyl cobalamin see p. 108). Succinyl-CoA is an intermediate in the tricar-... 

R) -2-Methyl-3-oxopropanoyl-coenzyme A, METHYLMALONYL-CoA EPIMERASE l-METHYLMALONYL-CoA MUTASE... 

Propionyl-CoA is first carboxylated to form the d stereoisomer of methylmalonyl-CoA (Pig. 17—11) by propionyl-CoA carboxylase, which contains the cofactor biotin. In this enzymatic reaction, as in the pyruvate carboxylase reaction (see Pig. 16-16), C02 (or its hydrated ion, HCO ) is activated by attachment to biotin before its transfer to the substrate, in this case the propionate moiety. Formation of the carboxybiotin intermediate requires energy, which is provided by the cleavage of ATP to ADP and Pi- The D-methylmalonyl-CoA thus formed is enzymatically epimerized to its l stereoisomer by methylmalonyl-CoA epimerase (Pig. 17-11). The L-methylmal onyl -CoA then undergoes an intramolecular rearrangement to form succinyl-CoA, which can enter the citric acid cycle. This rearrangement is catalyzed by methylmalonyl-CoA mutase, which requires as its coenzyme 5 -deoxyadenosyl-cobalamin, or coenzyme Bi2, which is derived from vitamin B12 (cobalamin). Box 17—2 describes the role of coenzyme B12 in this remarkable exchange reaction. 

Oxidation of unsaturated fatty acids requires two additional enzymes enoyl-CoA isomerase and 2,4-dienoyl-CoA reductase. Odd-number fatty acids are oxidized by the /3-oxidation pathway to yield acetyl-CoA and a molecule of propionyl-CoA This is carboxylated to methylmalonyl-CoA, which is isomerized to succinyl-CoA in a reaction catalyzed by methylmalonyl-CoA mutase, an enzyme requiring coenzyme B12. 

A review of the biochemistry of coenzyme B12 reactions, including the methylmalonyl-CoA mutase reaction. 

The propionyl-CoA derived from these three amino acids is converted to succinyl-CoA by a pathway described in Chapter 17 carboxylation to methylmalonyl-CoA, epimerization of the methylmalonyl-CoA, and conversion to succinyl-CoA by the coenzyme B independent methylmalonyl-CoA mutase (see Fig. 17-11). In the rare genetic disease known as methylmalonic acidemia, methylmalonyl-CoA mutase is lacking—with serious metabolic consequences (Table 18-2 Box 18-2). 

For this group of aminomutases PLP is required as a second coenzyme. Third, X is attached via a carbon atom the enzymes are called mutases. Methyl-malonyl-Co mutase is required for catabolism of propionate in the human body, and is one of only two known vitamin B12-... 

Figure 16-23 Three-dimensional structure of methylmalonyl-CoA mutase from Propionobacterium. shermanii. The B12 coenzyme is deeply buried, as is the active site. A molecule of bound desulfo-coenzyme A, a substrate analog, blocks the active site entrance on the left side. From Mancia et al.i05 Courtesy of Philip R. Evans.

In this hereditary disease up to 1 - 2 g of methylmalonic acid per day (compared to a normal of <5 mg/day) is excreted in the urine, and a high level of the compound is present in blood. Two causes of the rare disease are known/ One is the lack of functional vitamin B12-containing coenzyme. This can be a result of a mutation in any one of several different genes involved in the synthesis and transport of the cobalamin coenzyme.6 Cultured fibroblasts from patients with this form of the disease contain a very low level of the vitamin B12 coenzyme (Chapter 16), and addition of excess vitamin B12 to the diet may restore coenzyme synthesis to normal. Among elderly patients a smaller increase in methylmalonic acid excretion is a good indicator of vitamin B12 deficiency. A second form of the disease, which does not respond to vitamin B12, arises from a defect in the methylmalonyl mutase protein. Methylmalonic aciduria is often a very severe disease, frequently resulting in death in infancy. Surprisingly, some children with the condition are healthy and develop normally.3 1... 

Co corrinoids play central roles in the two classes of enzymic reactions, i.e. methyl transfer mediated by vitamin B,2 and mutase or isomerase reactions catalyzed by coenzyme B. 253 Though there remain many ambiguities, the former is considered to be a combination of Scheme 100, i and its reverse process, and the latter to be represented by Scheme 103. 

The product of acetyl-CoA carboxylase reaction, malonyl-CoA, is reduced via malonate semialdehyde to 3-hydroxypropionate, which is further reductively converted to propionyl-CoA. Propionyl-CoA is carboxylated to (S)-methylmalonyl-CoA by the same carboxylase. (S)-Methylmalonyl-CoA is isomerized to (R)-methylmal-onyl-CoA, followed by carbon rearrangement to succinyl-CoA by coenzyme B 12-dependent methylmalonyl-CoA mutase. Succinyl-CoA is further reduced to succinate semialdehyde and then to 4-hydroxybutyrate. The latter compound is converted into two acetyl-CoA molecules via 4-hydroxybutyryl-CoA dehydratase, a key enzyme of the pathway. 4-Hydroxybutyryl-CoA dehydratase is a [4Fe-4S] cluster and FAD-containing enzyme that catalyzes the elimination of water from 4-hydroxybutyryl-CoA by a ketyl radical mechanism to yield crotonyl-CoA [34]. Conversion of the latter into two molecules of acetyl-CoA proceeds via normal P-oxidation steps. Hence, the 3-hydroxypropionate/4-hydroxybutyrate cycle (as illustrated in Figure 3.5) can be divided into two parts. In the first part, acetyl-CoA and two bicarbonate molecules are transformed to succinyl-CoA, while in the second part succinyl-CoA is converted to two acetyl-CoA molecules. 

Vitamin B12 is a biologically active corrinoid, a group of cobalt-containing compounds with macrocyclic pyrrol rings. Vitamin B12 functions as a cofactor for two enzymes, methionine synthase and L-methylmalonyl coenzyme A (CoA) mutase. Methionine synthase requires methylcobalamin for the methyl transfer from methyltetrahydrofolate to homocysteine to form methionine tetrahy-drofolate. L-methylmalonyl-CoA mutase requires adenosylcobalamin to convert L-methylmalonyl-CoA to succinyl-CoA in an isomerization reaction. An inadequate supply of vitamin B12 results in neuropathy, megaloblastic anemia, and gastrointestinal symptoms (Baik and Russell, 1999). 

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