Enzyme phosphoenolpyruvate carboxylation

The direct carboxylation of pyruvate to form oxaloacetate is thermodynamically unfavorable. The enzyme phosphoenolpyruvate (PEP) carboxylase solves this problem by starting instead with phosphoenolpyruvate. 

In order to progress from pyruvate to glucose, pyruvate is first carboxylated in mitochondria to form oxaloac-etate. The action of malate dehydrogenase converts oxaloacetate to malate, which is able to leave the mitochondria and enter the cytosol where the reverse reaction regenerates oxaloacetate. The enzyme phosphoenolpyruvate carboxykinase then catalyses the phosphorylation and decarboxylation of oxaloacetate to form phosphoenolpyruvate (31a). 

Oxaloacetate is an intermediate of many metabolic pathways. It also plays a role in the malate-aspartate shuttle, which transfers high energy electrons into mitochondria. Citrate is formed by the condensation of oxaloacetate with acetyl CoA. A transamination reaction transfers an amino group from an amino acid to an a-keto acid. Transfer of the amino group from aspartate to a-ketoglutarate forms oxaloacetate and glutamate. In gluconeogenesis, pyruvate is carboxylated in mitochondria to form oxaloacetate. After transfer to the cytosol, the enzyme phosphoenolpyruvate carboxykinase catalyses the conversion of oxaloacetate to phosphoenolpyruvate. 

Larsen, T.M., Wedeking, J.E., Rayment, I. and Reed, G.H. (1996) A carboxylate oxygen of the substrate bridges the magnesium ions at the active site of enolase structure of the yeast enzyme complexed with the equilibrium mixture of 2-phosphoglycerate and phosphoenolpyruvate at 1.8 A resolution, Biochemistry, 30, 4349-4358. 

In addition to the aforementioned allenic steroids, prostaglandins, amino acids and nucleoside analogs, a number of other functionalized allenes have been employed (albeit with limited success) in enzyme inhibition (Scheme 18.56) [154-159]. Thus, the 7-vinylidenecephalosporin 164 and related allenes did not show the expected activity as inhibitors of human leukocyte elastase, but a weak inhibition of porcine pancreas elastase [156], Similarly disappointing were the immunosuppressive activity of the allenic mycophenolic acid derivative 165 [157] and the inhibition of 12-lipoxygenase by the carboxylic acid 166 [158]. In contrast, the carboxyallenyl phosphate 167 turned out to be a potent inhibitor of phosphoenolpyruvate carboxylase and pyruvate kinase [159]. Hydrolysis of this allenic phosphate probably leads to 2-oxobut-3-enoate, which then undergoes an irreversible Michael addition with suitable nucleophilic side chains of the enzyme. 

C4 plants incorporate CO2 by the carboxylation of phosphoenolpyruvate (PEP) via the enzyme PEP carboxylase to make the molecule oxaloacetate which has 4 carbon atoms (hence C4). The carboxylation product is transported from the outer layer of mesophyll cells to the inner layer of bundle sheath cells, which are able to concentrate CO2, so that most of the CO2 is fixed with relatively little carbon fractionation. 

This cycle resembles the 3-hydroxypropionate/4-hydroxybutyrate cycle, but with pyruvate ferredoxin oxidoreductase (pyruvate synthase) and phosphoenolpyruvate (PEP) carboxylase as the carboxylating enzymes (Figure 3.6). 

Fig. 5.4. Two types of energy metabolism in cestodes. (a) Type 1 homolactate fermentation, (b) Type 2 Malate dismutation. Reaction 3 involves a carboxylation step decarboxylation occurs at 6, 7 and 10. Reducing equivalents are generated at reactions 6 and 7 one reducing equivalent is used at reaction 9. Thus, when the mitochondrial compartment is in redox balance and malate is the sole substrate, twice as much propionate as acetate is produced. Key 1, pyruvate kinase 2, lactate dehydrogenase 3, phosphoenolpyruvate carboxykinase 4, malate dehydrogenase 5, mitochondrial membrane 6 malic enzyme 7, pyruvate dehydrogenase complex 8, fumarase 9, fumarate reductase 10, succinate decarboxylase complex. indicates reactions at which ATP is synthesised from ADP cyt, cytosol mit, mitochondrion. (After Bryant Flockhart, 1986.)...

Today the metabolic network of the central metabolism of C. glutamicum involving glycolysis, pentose phosphate pathway (PPP), TCA cycle as well as anaplerotic and gluconeogenetic reactions is well known (Fig. 1). Different enzymes are involved in the interconversion of carbon between TCA cycle (malate/oxaloacetate) and glycolysis (pyruvate/phosphoenolpyruvate). For anaplerotic replenishment of the TCA cycle, C. glutamicum exhibits pyruvate carboxylase [20] and phosphoenol-pyruvate (PEP) carboxylase as carboxylating enzymes. Malic enzyme [21] and PEP carboxykinase [22,23] catalyze decarboxylation reactions from the TCA cycle... 

Carboxylation is, in the first place, a thermodynamic problem. Nature solves this problem by associating the carboxylation with some exergonic process. This may be reduction of a product (as in the case of malic enzyme and isocitrate dehydrogenase), hydrolysis of ATP (as in the case of pyruvate carboxylase), hydrolysis of phosphoenolpyruvate (as in the case of phosphoenolpyruvate carboxylase), or cleavage of a carbon-carbon bond (as in the case of ribulose-bisphosphate carboxylase). 

C4 plants green plants in which the primary product of CO2 fixation is not 3-phosphoglycerate (cf. C3 plants) but a C4 acid such as oxaloacetate, malate or aspartate. These plants possess two types of photi>-synthesizing cells. In mesophyll cells near the leaf surface, CO2 is fixed into C4-compounds. This prefixation of CO2 is due to the action of the cytosolic enzyme, phosphoeno/pyruvate carboxylase (EC 4.1.1.31), which carboxylates phosphoenolpyruvate to oxaloacetic acid (see Hatch-Slack-Kortschak cycle). The Calvin cycle (see) operates in the the vascular bundle cells of C4 plants, and CO2 for the Calvin cycle is derived from the decarboxylation C4 compounds rather than directly from the atmosphere. This Kranz anatomy , i.e. photosynthetically active bundle sheath cells with a photosynthetically active layer... 

Pyruvate may enter the Krebs cycle by a pathway different from that involving the formation of acetyl-CoA. Indeed, the keto acid can be converted to a dicarboxylic acid by at least two different biochemical processes. One of them involves the carboxylation of pyruvic acid leading to the formation of oxaloacetic acid. The actual substrate in that reaction is not pyruvic acid, but phosphoenolpyruvic acid. The enzyme involved requires manganese, and IDP acts as the phosphorus acceptor. The reaction is coupled with the pyrophosphate transfer (catalyzed by nucleoside diphosphokinase) from ITP to ADP. 

In mammalian tissues, excluding muscle, the most important anaplerotic reaction employs pyruvate carboxylase which contains a biotin prosthetic group (Figure 5.3b) responsible for the transfer of a carboxyl group. ATP provides the energy to bond covalently the carboxyl group from HCO, to the biotin which transfers it when pyruvate binds to the enzyme. In muscle cells, the major pathway utilizes phosphoenolpyruvate and phos-phoenolpyruvate carboxykinase which occurs both in the cytosol and mitochondrial matrix. Two routes are therefore possible in oxaloacetate syn-... 

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