Phosphoenolpyruvate carboxylase, pyruvate

The Jirst indirect route in glucose synthesis involves the formation of phosphoenolpyruvate from pyruvate without the intervention of pyruvate kinase. This route is catalyzed by two enzymes. At first, pyruvate is converted into oxaloacetate. This reaction occurs in the mitochondria as the pyruvate molecules enter them, and is catalyzed by pyruvate carboxylase according to the scheme... 

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. 

A problem for gluconeogenesis is that pyruvate carboxylase, which produces oxaloacetate from pyravate, is present in the mitochondria but phosphoenolpyruvate carboxylase, at least in human liver, is present in the cytosol. For reasons given in Chapter 9, oxaloacetate cannot cross the mitochondrial membrane and so a transporter is not present in any cells. Hence, oxaloacetate is converted to phosphoenolpyruvate which is transported across the membrane (Figure 6.25). 

This enzyme [EC 4.1.1.38] (also known as phosphoenolpyruvate carboxytransphosphorylase, phosphopyruvate carboxylase, and phosphoenolpyruvate carboxylase) catalyzes the reaction of phosphoenolpyruvate with orthophosphate and carbon dioxide to produce oxaloacetate and pyrophosphate (or diphosphate). The enzyme also catalyzes the reaction of phosphoenolpyruvate with orthophosphate to produce pyruvate and pyrophosphate. 

Pathway of C02 in Gluconeogenesis In the first bypass step of gluconeogenesis, the conversion of pyruvate to phosphoenolpyruvate (PEP), pyruvate is carboxylated by pyruvate carboxylase to oxaloacetate, which is subsequently decarboxylated to PEP by PEP carboxykinase (Chapter 14). Because the addition of C02 is directly followed by the loss of C02, you might expect that in tracer experiments, the 14C of 14C02 would not be incorporated into PEP, glucose, or any intermediates in gluconeogenesis. 

The Q pathway for the transport of CO2 starts in a mesophyll cell with the condensation of CO2 and phosphoenolpyruvate to form oxaloacetate, in a reaction catalyzed by phosphoenolpyruvate carboxylase. In some species, oxaloacetate is converted into malate by an NADP+-linked malate dehydrogenase. Malate goes into the bundle-sheath cell and is oxidatively decarboxylated within the chloroplasts by an NADP+-linked malate dehydrogenase. The released CO2 enters the Calvin cycle in the usual way by condensing with ribulose 1,5-bisphosphate. Pyruvate formed in this decarboxylation reaction returns to the mesophyll cell. Finally, phosphoenolpyruvate is formed from pyruvate by pyruvate-Pi dikinase. 

Figure 5-25. The conversion of pyruvate to phosphoenolpyruvate (PEP). Follow the diagram by starting with the precursors alanine and lactate. OAA = oxaloacetate FA = fatty acid TG = triacylglycerol PDH = pyruvate dehydrogenase PC = pyruvate carboxylase PEPCK = phosphoenolpyruvate PK = pyruvate kinase PK-P = phos- phorylated pyruvate kinase.

The answer is a. (Murray, pp 190—198. Sci ivei, pp 1521—1552. Sack, pp 121-138. Wilson, pp 287-317.1 In the formation of phosphoenolpyruvate during gluconeogenesis, oxaloacetate is an intermediate. In the first step, catalyzed by pyruvate carboxylase, pyruvate is carboxylated with the utilization of one high-energy ATP phosphate bond ... 

Fig. 6.3. Carbon dioxide-fixing pathway in the C4-plants (prepared mainly on the basis of Hatch et al., 1967). Circled numbers 1, phosphoenolpyruvate carboxylase 2, malate dehydrogenase (NADP ) 3, malate dehydrogenase (NADP ) (OAA decarboxylating) (= malic enzyme) 4, Rubisco 5, pyruvate orthophosphate dikinase. Pi, phosphate PPi, diphosphate...

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). 

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). 

Fig. 31.5. Conversion of pyruvate to phosphoenolpyruvate (PEP). Follow the shaded circled numbers on the diagram, starting with the precursors alanine and lactate. The first step is the conversion of alanine and lactate to pyruvate. Pyruvate then enters the mitochondria and is converted to OAA (circle 2) by pyruvate carboxylase. Pyruvate dehydrogenase has been inactivated by both the NADH and acetyl-CoA generated from fatty acid oxidation, which allows oxaloacetate production for gluconeogenesis. The oxaloacetate formed in the mitochondria is converted to either malate or aspartate to enter the cytoplasm via the malate/aspartate shuttle. Once in the cytoplasm the malate or aspartate is converted back into oxaloacetate (circle 3), and phosphoenolpyruvate carboxykinase will convert it to PEP (circle 4). The white circled numbers are alternate routes for exit of carbon from the mitochondrion using the malate/aspartate shuttle. OAA = oxaloacetate FA = fatty acid TG = triacylglycerol.

STRUCTURE OF THE GENES FOR PHOSPHOENOLPYRUVATE CARBOXYLASE AND PYRUVATE Pi DIKINASE FROM MAIZE. 

Phosphoenolpyruvate carboxylase (PEPC) and pyruvate Pi dikinase (PPDK) play significant roles in the photosynthetic fixation of carbon in and CAM plants. PEPC catalyzes the fixation of atmospheric CO2 to phosphoenolpyruvate (1), and PPDK catalyzes the formation of phosphoenolpyruvate, the substrate for PEPC (2). 

Structure of the Genes for Phosphoenolpyruvate Carboxylase and Pyruvate Pi Dikinase... 

Lin H, San KY, Bennett GN. (2005d). Effect of Sorghum vulgare phosphoenolpyruvate carboxylase and Lactococcus lactis pyruvate carboxylase coexpression on succinate production in mutant strains of Escherichia coli. Appl Microbiol Biotechnol, 67,515-523. 

Cooper TG, Tchen TT, Wood HG and Benedict CR (1968) The carboxylation of phosphoenolpyruvate and pyruvate. I. The active species of CO2 utilized by phosphoenolpyruvate carboxykinase, carboxytransphosphorylase, and pyruvic carboxylase. J Biol Chem 243 3857-3863... 

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