Oxidative decarboxylation of a-ketoglutarate

In the normal prolyl 4-hydroxylase reaction (Fig. 4a), one molecule of a-ketoglutarate and one of 02 bind to the enzyme. The a-ketoglutarate is oxidatively decarboxylated to form C02 and succinate. The remaining oxygen atom is then used to hydroxylate an appropriate Pro residue in procollagen. No ascorbate is needed in this reaction. However, prolyl 4-hydroxylase also catalyzes an oxidative decarboxylation of a-ketoglutarate that is not coupled to proline hydroxylation—and this is the reaction that requires ascorbate (Fig. 4b). During this reaction, the heme Fe2+ becomes oxidized, and the oxidized form of the enzyme is inactive—unable to hydroxylate proline. The ascorbate consumed in the reaction presumably functions to reduce the heme iron and restore enzyme activity. 

The formation of ATP (or GTP) at the expense of the energy released by the oxidative decarboxylation of a-ketoglutarate is a substrate-level phosphorylation, like the synthesis of ATP in the glycolytic reactions catalyzed by glyceraldehyde 3-phosphate dehydrogenase and pyruvate kinase (see Fig. 14-2). The GTP formed by succinyl-CoA synthetase can donate its terminal phosphoryl group to ADP to form ATP, in a reversible reaction catalyzed by nucleoside diphosphate kinase (p. 505) ... 

TPP also mediates the oxidative decarboxylation of a-ketoglutaric acid, another intermediate of carboxydrate metabolism in the citric acid cycle. The nutritional requirement for thiamine increases as dietary carbohydrate increases because of a greater demand for TPP. 

The oxidative decarboxylation of a-ketoglutarate closely resembles that of pyruvate, also an a-ketoacid. 

Succinyl Coenzyme A Ts Formed by the Oxidative Decarboxylation of a-Ketoglutarate 485... 

Within the TCA cycle, the oxidative decarboxylation of a-ketoglutarate is catalyzed by the multisubunit a-ketoglutarate dehydrogenase complex, which contains the coenzymes thiamine-pyrophosphate, lipoate, and FAD. A similar complex, the pyruvate dehydrogenase complex (PDC), catalyzes the oxidation of pyruvate to acetyl CoA, thereby providing a link between the pathways of glycolysis and the TCA cycle (see Fig. 20.1)... 

Fig. 20.8. Oxidative decarboxylation of a-ketoglutarate. The a-ketoglutarate dehydrogenase complex oxidizes a-ketoglutarate to succinyl CoA. The carboxyl group is released as COp. The keto group on the a-carbon is oxidized, and then forms the acyl CoA thioester, succinyl CoA. The a, p, 7, and 8 on succinyl CoA refer to the sequence of atoms in a-ketog-lutarate.

Of all the enzymes of the carnitine biosynthetic pathway, yBBH is the best-studied enzyme. Like e-N-trimethyllysine hydroxylase, y-BBH is a non-heme ferrous-iron dioxygenase that requires a-ketoglutarate, Fe and molecular oxygen as cofactors. In this class of enzymes, the hydroxylation of the substrate is linked to the oxidative decarboxylation of a-ketoglutarate. Ascorbate is needed to maintain iron in die reduced state. yBBHhas been isolated from various sources including human kidney, " catf and rat liver and the bacterium Pseudomonas AKl. A common problem in the purification of this enzyme from mammalian tissues is the poor stability of the protein. We, therefore, determined the optimal storage conditions and subsequently pmified ybutyrobetaine hydroxylase from rat liver. 

Coenzyme A has recently been implicated in the activation and transfer of acyl groups other than acetate. For example, CoA is required for the oxidative decarboxylation of a-ketoglutarate to form succinate (Kaufman, 1951 Sanadi and Littlefield, 1951). Succinyl-CoA was proposed as an intermediate in this reaction. Hippuric acid synthesis also requires CoA (Chantrenne, 1951). The postulated intermediate in this case was benzoyl-CoA. The formation of acyl-CoA complexes from acetyl-CoA and free fatty acids may also occur (Stadtman, 1950, 1951). 

In the R.c.c., two of the reactions of the TCA cycle are replaced by alternative reactions catalysed by non-TCA cycle enzymes (i) the citrate (si)-synthase (EC 4.1.3.7)-catalysed formation of citrate from ace-tyl-CoA and oxaloacetate is replaced by the ATP citrate (pro-3.S)-lyase (EC 4.1.3.8)-catalysed, ATP-dri-ven cleavage of citrate to acetyl-CoA and oxaloacetate (eq. 1 J, Fig.), and (ii) the a-ketoglutaiate dehydrogenase complex-catalysed oxidative decarboxylation of a-ketoglutarate to CO2 and succinyl-CoA is replaced by the a-ketoglutarate synthase (EC 1.2.73)-catalysed reductive carboxylation of succinyl CoA (eq. 2 O, Fig.), in which the reductant is reduced ferredoxin (Fd, ... 

Thiamin pyrophosphate (or thiamin diphosphate) is a coenzyme involved in (1) the oxidative decarboxylation of pyruvate to acetyl coenzyme A (enzyme pyruvate dehydrogenase), (2) the oxidative decarboxylation of a-ketoglutarate to succinyl coenzyme A (a-ketoglutarate dehydrogenase) in the tricarboxylic add cycle, (3) the pentose phosphate pathway (transketolase) and (4) the synthesis of branched-chain amino acids such as valine (branched-chain ketoacid dehydrogenase) in bacteria, yeasts and plants. 

This particular form of the system of dicarboxylic acids had to be abandoned when Krebs showed that a-ketoglutarate and citrate, in addition to succinate, fumarate, malate and oxaloacetate, also re-establish the respiration of a muscle pulp. The case of a-ketoglutaric acid, in the scheme of Szent-Gyorgyi, did not present an insurmountable difficulty since the oxidative decarboxylation of a-ketoglutarate pelds succinic acid. 

Recently George Gardinale has been able to show that prolyl hydroxylase can catalyze the oxidative decarboxylation of a-ketoglutarate in the absence of a prolyl substrate,>at a lower but measurable rate. The same cofactors, Fe and ascorbate, are required and optimal conditions are the same for both. This suggests that a-ketoglutarate is the first acceptor of the oxygen and that a peroxy compound, perhaps enzyme bound persuccinate, then attacks the 4-trans position of proline to yield the intermediate shown above. Gleavage of the peroxy compound would then yield equal amounts of succinate and hydroxyproline. 

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