4- hydroxybutyryl-CoA dehydratase

H ydroxypropionate / 4- hydroxybutyrate cycle 9 6 NAD(P)H Acetyl-CoA/propionyl-CoA carboxylase HCOJ Acetyl-CoA, succinyl-CoA Acetyl-CoA/propionyl-CoA carboxylase", enzymes reducing malonyl-CoA to propionyl-CoA, methylmalonyl-CoA rnutase, 4-hydroxybutyryl-CoA dehydratase... 

Dicarboxylate/ 4-hydroxybutyrate cycle11 8 2 NAD(P)H, 3 ferredoxin, 1 unknown donor Pyruvate synthase, PEP carboxylase C02 HCOJ Acetyl-CoA, pyruvate, PEP, oxaloacetate, succinyl-CoA 4-Hydroxybutyryl-CoA dehydratase... 

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. 

Hydroxybutyryl-CoA Dehydrase. - 4-Hydroxybutyryl-CoA dehydratase catalyses the reversible dehydration of 4-hydroxybutyryl-CoA to crotonyl-CoA, which involves cleavage of an unactivated P-C-H bond. The enzyme also catalyses the apparently irreversible isomerization of vinylacetyl-CoA to crotonyl-CoA. The enzyme has been described as homotetramer with M, 56 kDa per subunit. It is thought to contain two molecules of FAD and one [4Fe-4S] cluster per homotetramer. 

Despite the fact that numerous enzymes have been characterized that catalyze the addition of water to unsaturated fatty acids that are coupled to CoA or ACP, such as methylglucatonyl-CoA hydratase (E.C. 4.2.1.18), lactoyl-CoA dehydratase (E.C. 4.2.1.54), 3-hydroxybutyryl-CoA dehydratase (E.C. 4.2.1.55), itaconyl-CoA dehydratase (E. C. 4.2.1.56), isohexenylglutaconyl-CoA hydratase (E. C. 4.2.1.57), farnesyl-CoA dehydratase (E.C. 4.2.1.57), long-chain enoyl-CoA hydratase (E.C. 4.2.1.74), 3-hydroxydecanoyl-ACP dehydratase (E.C. 4.2.1.60) and 3-hydroxypalmitoyl-ACP dehydratase (E.C. 4.2.1.61), these enzymes are seldomly applied in organic synthesis. 

Miih and co-workers have characterized 4-hydroxybutyryl-CoA dehydrase from Clostridium aminobutyricum by optical spectroscopy and EPR. The EPR spectrum of active dehydratase showed essentially no signal at 100 K. When the enzyme was reduced such that the optical spectrum indicated the presence of a flavin radical. An EPR signal, with g = 2.0014, was detected at the same temperature. It showed no hyperfine structure and had a linewidth of 2 mT, characteristic for a neutral flavin radical, and could be easily saturated at lower temperatures. When the FAD was photoreduced further to the hydroquinone the EPR signal disappeared. 

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