Glycolysis Phosphoenolpyruvic carboxykinase

Biotin acts to induce glucokinase, phosphofructokinase, and pyruvate kinase (key enzymes of glycolysis), phosphoenolpyruvate carboxykinase (a key enzyme of gluconeogenesis), and holocarboxylase synthetase, acting via a cell-surface receptor linked to formation of cGMP and increased activity of RNA polymerase. The activity of holocarboxylase synthetase (Section 11.2.2) falls in experimental biotin deficiency and increases with a parallel increase in... 

Pyruvate is converted to oxaloacetate (by pyruvate carboxylase). The oxaloacetate is decarboxylated and phosphorylated to phosphoenolpyruvate (PEP) by phosphoenolpyruvate carboxykinase (PEP carboxykinase). PEP is converted to fructose 1,6-bisphosphate by a direct reversal of several reactions in glycolysis. Next, fructose 1,6-bisphosphate is dephosphorylated to fructose 6-phosphate (by fructose 1,6-bisphosphatase) and this is then converted to glucose 6-phosphate (by phosphoglucoisomerase). Finally, glucose 6-phosphate is dephosphorylated (by glucose 6-phosphatase) to yield glucose. 

Most cestodes which have been investigated, however, conform to the second category, type 2, which is characterised by a C02-fixation step. Carbohydrate is degraded to the level of PEP by glycolysis, the steps involved being similar to those in mammalian tissue. At this point, the enzymes pyruvate kinase and phosphoenolpyruvate carboxykinase (PEPCK) compete for available substrate and a branch-point occurs (Fig. 5.4). The relative activities of these two enzymes determine the fate of the PEP and the subsequent types and amounts of end-products formed (see below). 

Finally, oxaloacetate is simultaneously decarboxylated andphosphorylated by phosphoenolpyruvate carboxykinase in the cytosol. The CO2 that vv as added to pyruvate by pyruvate carboxylase comes off in this step. Recall that, in glycolysis, the presence of a phosphoryl group traps the unstable enol isomer of pyruvate as phosphoenolpyruvate (Section 16.1.7). In gluconeogenesis, the formation of the unstable enol is driven by decarboxylation—the oxidation of the carboxylic acid to CO2 —and trapped by the addition of a phosphate to carbon 2 from GTP. The two-step pathway... 

Phosphoenolpyruvate carboxykinase converts oxaloacetate to phospho-enolpyruvate, Phosphoenolpyruvate forms fructose 1,6-bisphosphate by reversal of the steps of glycolysis. 

Comparison of the reactions of glycolysis and gluconeogenesis. All the reactions of glycolysis occur in the cytoplasm of the cell. However, in many human cells, pyruvate carboxylase is found in the mitochondria and phosphoenolpyruvate carboxykinase is located in the cytoplasm. Oxaloacetate, the product of the reaction catalyzed by pyruvate carboxylase, is shuttled out of the mitochondria and into the cytoplasm by a complex set of reactions. 

Finally, oxaloacetate is simultaneously decarboxylated and phosphorylated by phosphoenolpyruvate carboxykinase in the cytosol. The CO2 that was added to pyruvate by pyruvate carboxylase comes off in this step. Recall that, in glycolysis, the presence of a phosphoryl group traps the unstable enol isomer of pyruvate as phosphoenolpyruvate (Section 16.1.7). In gluconeogenesis, the formation of the unstable enol is driven by decarboxylation—the oxidation of the carboxylic acid to CO2—and trapped by the addition of a phosphate to carbon 2 from GTP. The two-step pathway for the formation of phosphoenolpyruvate from pyruvate has a AG° of + 0.2 kcal mol ( + 0.13 kj moP ) in contrast with +7.5 kcal mol ( + 31 kj mol ) for the reaction catalyzed by pyruvate kinase. The much more favorable AG° for the two-step pathway results from the use of a molecule of ATP to add a molecule of CO2 in the carboxylation step that can be removed to power the formation of phosphoenolpyruvate in the decarboxylation step. Decarboxylations often drive reactions otherwise highly endergonic. This metabolic motif is used in the citric acid cycle (Section IS.x.x), the pentose phosphate pathway (Section 17.x.x), and fatty acid synthesis (Section 22.x.x). 

Enzymes that participate in gluconeogenesis, but not in glycolysis, are active under fasting conditions. Pyruvate carboxylase is activated by acetyl CoA, derived from oxidation of fatty acids. Phosphoenolpyruvate carboxykinase, fructose 1,6-bisphosphatase, and glucose 6-phosphatase are induced that is, the quantity of the enzymes increases. Eructose 1,6-bisphosphatase is also active because levels of fructose 2,6-bisphosphate, an inhibitor of the enzyme, are low. 

Gluconeogenesis is stimulated because the synthesis of phosphoenolpyruvate carboxykinase, fructose 1,6-bisphosphatase, and glucose 6-phosphatase is induced and because there is an increased availability of precursors. Fructose 1,6-bisphos-phatase is also activated because the levels of its inhibitor, fructose 2,6-bisphos-phate, are low (Fig. 36.9). During fasting, the activity of the corresponding enzymes of glycolysis is decreased. 

Figure 2.4. Interconnection of the citric acid cycle and the pathways of glycolysis and gluconeogenesis. MDH = malate dehydrogenase. ME= malate NADP dehydrogenase. PEPCK = phosphoenolpyruvate carboxykinase. PK = pyruvate kinase. PC = pyruvate carboxylase...

Anaplerotic reactions refer to C3-carboxylation and C4-decarboxylation around the phosphoenolpyruvate-pyruvate-oxaloacetate node, which interconnect the TCA cycle with glycolysis. These reactions result in direct oxaloacetate formation or depletion. Carboxylation of phosphoenolpyruvate catalyzed by phosphoenolpyruvate carboxylase and that of pyruvate by pyruvate carboxylase contribute to its formation. Accordingly, decarboxylation of oxaloacetate catalyzed by phosphoenolpyruvate carboxykinase and oxaloacetate decarboxylase form phosphoenolpyruvate and... 

The principal substrate for gluconeogenesis is oxaloacetate, which undergoes the reaction catalysed by phosphoenolpyruvate carboxykinase to yield phos-phoenolpyruvate, as shown in Figure 5.31. The onward metabolism of phosphoenolpyruvate to glucose is essentially the reverse of glycolysis shown in Figure 5.10. 

The first "roadblock" to overcome in the synthesis of glucose from pyruvate is the irreversible conversion in glycolysis of pyruvate to phosphoenolpyruvate (PEP) by pyruvate kinase. In gluconeogenesis, pyruvate is first carboxylated by pyruvate carboxylase to oxaloacetate (OAA), which is then converted to PEP by the action of PEP-carboxykinase (Figure 10.3). 

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... 

Figure 6. Variations on phosphoenolpyruvate (PEP) and pyruvate (PYR) metabolism in animals. In mammalian liver, pyruvate kinase (PK) and PEP carboxykinase (PEPCK) function in opposite directions to support glycolysis versus gluconeogenesis. In anoxia-tolerant mollusks, PEP is routed via PK when oxygen is present and via PEPCK in anoxia. Note that PEPCK is adapted for physiological function in opposite directions in the two situations.

Four enzymes are unique to the process of gluconeogenesis and are required to circumvent the unidirectional steps of glycolysis (Fig. 13.1). Two of these are pyruvate carboxylase, which converts pyruvate to oxaloacetate, and PEP carboxykinase, which converts oxaloacetate to phosphoenolpyruvate. These enzymes are required to effectively reverse the action of pyruvate kinase. The other two are fructose-1,6-bisphosphatase and glucose-6-phosphatase, which effectively reverse the actions of PFK and hexokinase or glucokinase. 

The irreversible step of glycolysis catalysed by pyruvate kinase is bypassed in gluconeogenesis by conversion of pyruvate first to oxaloacetate, then conversion of oxaloacetate to phosphoenolpyruvate by phosphoenolpyru-vate carboxykinase. The transfer of the amino group from glutamate to oxaloacetate produces aspartate, catalysed by the enzyme aspartate aminotransferase. Malate dehydrogenase converts oxaloacetate to malate in the malate-aspartate shutde. 

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