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Query: EC:4.1.1.32 (phosphoenolpyruvate carboxykinase)
 4,204 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ammoniagenesis and gluconeogenesis have been studied in foetal rat kidneys during the five last days of gestation by measuring in vitro NH3 and glucose productions from glutamine and by assaying activities of soluble phosphoenolpyruvate carboxykinase(PEPCK) and glucose 6-phosphatase (G6Pase). These studies were carried out in normal (mean blood pH: 7.30) and acidotic (pH: 7.12) foetuses. In normal foetuses, NH3 production by kidney cortex slices remains constant throughout the studied period of development, at a level 10-fold lower than the maternal one. On day 20 of gestation, a low glucose production (20-fold lower than the maternal one) appears for the first time. This may be related to an increase of PEPCK and G6Pase activities which occurs between day 19 and 20. In 20 days old foetuses, NH4Cl induced acidosis stimulates NH3 production but has no effect on PEPCK activity and glucose production. A response of gluconeogenesis to acidosis is observed one day later (day 21).
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PMID:Development of ammonia and glucose productions from glutamine in foetal rat kidney; effects of metabolic acidosis. 21 16

Experiments were carried out on rats to evaluate the possible regulatory roles of renal glutaminase activity, mitochondrial permeability to glutamine, phosphoenolpyruvate carboxykinase activity and systemic acid-base changes in the control of renal ammonia (NH(3) plus NH(4) (+)) production. Acidosis was induced by drinking NH(4)Cl solution ad libitum. A pronounced metabolic acidosis without respiratory compensation [pH=7.25; HCO(3) (-)=16.9mequiv./litre; pCO(2)=40.7mmHg (5.41kPa)] was evident for the first 2 days, but thereafter acid-base status returned towards normal. This improvement in acid-base status was accompanied by the attainment of maximal rates of ammonia excretion (onset phase) after about 2 days. A steady rate of ammonia excretion was then maintained (plateau phase) until the rats were supplied with tap water in place of the NH(4)Cl solution, whereupon pCO(2) and HCO(3) (-) became elevated [55.4mmHg (7.37kPa) and 35.5mequiv./litre] and renal ammonia excretion returned to control values within 1 day (recovery phase). Renal arteriovenous differences for glutamine always paralleled rates of ammonia excretion. Phosphate-dependent glutaminase and phosphoenolpyruvate carboxykinase activities and the rate of glutamine metabolism (NH(3) production and O(2) consumption) by isolated kidney mitochondria all increased during the onset phase. The increases in glutaminase and in mitochondrial metabolism continued into the plateau phase, whereas the increase in the carboxykinase reached a plateau at the same time as did ammonia excretion. During the recovery phase a rapid decrease in carboxykinase activity accompanied the decrease in ammonia excretion, whereas glutaminase and mitochondrial glutamine metabolism in vitro remained elevated. The metabolism of glutamine by kidney-cortex slices (ammonia, glutamate and glucose production) paralleled the metabolism of glutamine in vivo during recovery, i.e. it returned to control values. The results indicate that the adaptations in mitochondrial glutamine metabolism must be regulated by extra-mitochondrial factors, since glutamine metabolism in vivo and in slices returns to control values during recovery, whereas the mitochondrial metabolism of glutamine remains elevated.
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PMID:Glutamine metabolism in the kidney during induction of, and recovery from, metabolic acidosis in the rat. 70 90

1. Neither alloxan-diabetes nor starvation affected the rate of glucose production in hepatocytes incubated with lactate, pyruvate, propionate or fructose as substrates. In contrast, glucose synthesis with either alanine or glutamine was increased nearly 3- and 12-fold respectively, in comparison with that in fed rabbits. 2. The addition of amino-oxyacetate resulted in about a 50% decrease in glucose formation from lactate in hepatocytes isolated from fed, alloxan-diabetic and starved rats, suggesting that both mitochondrial and cytosolic forms of rabbit phosphoenolpyruvate carboxykinase function actively during gluconeogenesis. 3. Alloxan-diabetes resulted in about 2-3-fold stimulation of urea production from either amino acid studied or NH4Cl as NH3 donor, whereas starvation caused a significant increase in the rate of ureogenesis only in the presence of alanine as the source of NH3. 4. As concluded from changes in the [3-hydroxybutyrate]/[acetoacetate] ratio, in hepatocytes from diabetic animals the mitochondrial redox state was shifted toward oxidation in comparison with that observed in liver cells isolated from fed rabbits.
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PMID:Effect of alloxan-diabetes on gluconeogenesis and ureogenesis in isolated rabbit liver cells. 74 58

Biochemical and metabolic data have led to the conclusion that the enzyme phosphoenolpyruvate carboxykinase (PEPCK; EC 4.1.1.32) contributes to a critical point of divergence in energy conservation pathways between mammals and nematodes. To facilitate the determination of the molecular basis for host vs parasite differences in PEPCK, we have cloned a cDNA encoding this enzyme from a parasitic nematode of ruminants, Haemonchus contortus. H. contortus PEPCK was cloned by functional complementation of a PEPCK-, malic enzyme- strain of Escherichia coli (E1786) using an egg stage H. contortus cDNA library in lambda ZAPII. Selection was for growth on malate as the sole carbon source (malate+ phenotype). We isolated a plasmid, pPEPCK, which reproducibly confers a malate+ phenotype in E1786. The sequence of the 2.0-kb EcoRI insert of pPEPCK predicts a 612-amino acid protein which shows about 74% similarity to Drosophila melanogaster and chicken PEPCK. Extracts of E1786[pPEPCK], but not E1786, contain IDP- or GDP-dependent PEPCK enzyme activity. Sequence analysis revealed that the open reading frame (ORF) in pPEPCK lacked a 5' initiation codon and was probably expressed as an in-frame fusion protein with beta-galactosidase. A strategy combining library screening with PCR analysis of positive clones led to the identification of a clone encoding 6 additional NH2-terminal amino acids, including a Met, which, by comparison with known PEPCK amino acid sequences, is likely to be the translation initiation site.
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PMID:Cloning of a cDNA encoding phosphoenolpyruvate carboxykinase from Haemonchus contortus. 174 Oct 16

Axenic culture amastigote-like forms of Trypanosoma cruzi, grown at 28 degrees C, reach a stationary phase after two generations, and differentiate to epimastigotes, which then resume growth. Axenic culture amastigotes readily ferment glucose to succinate and acetate, and do not excrete NH3; they have high activities of hexokinase and phosphoenolpyruvate carboxykinase, and very low citrate synthase activity; cytochrome o is absent, and cytochrome b-like is present at a very low level. Epimastigotes catabolize glucose and produce succinate and acetate at a considerably lower rate; they exhibit lower levels of hexokinase and carboxykinase, and much higher levels of citrate synthase and cytochromes o and b-like. They catabolize amino acids, as shown by excretion of NH3 to the medium. The results suggest that axenic culture amastigotes have an essentially glycolytic metabolism, and they acquire the ability to oxidize substrates such as amino acids only after differentiation to epimastigotes.
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PMID:Aerobic glucose fermentation by Trypanosoma cruzi axenic culture amastigote-like forms during growth and differentiation to epimastigotes. 332 2

The exchange inert coordination complexes, Cr(H2O)4GDP, Cr(H2O)4GTP, Cr(NH3)4GDP, Cr(NH3)4GTP, Co(NH3)4GDP, and Co(NH3)4GTP have been synthesized and characterized. The lambda and delta coordination isomers of Cr(H2O)4GDP, Cr(NH3)4GDP, and the four Cr(H2O)4GTP isomers have been separated by reverse phase HPLC and characterized by their CD spectra. While the isomers of Co(NH3)4GTP have not been successfully separated, 31P NMR spectroscopy reveals the presence of the lambda and delta forms. The complexes, Cr(H2O)4GDP, Co(NH3)4GDP, Cr(H2O)4GTP, and Co(NH3)4GTP, are linear competitive inhibitors of avian phosphoenolpyruvate carboxykinase. The Ki values of 30 microM, 540 microM, 40 microM, and 12 microM, respectively, were determined for these complexes using Mn-IDP as the nucleotide substrate in the phosphoenolpyruvate carboxylation direction or Mn-ITP as nucleotide substrate for the oxalacetate decarboxylation reaction. The lambda and delta isomers of Cr(H2O)4 GDP show little specificity (a twofold maximum difference in Ki) for the enzyme. The isomeric forms of Cr(H2O)4 GTP demonstrate no observed stereoselectivity of interaction with the enzyme. All of the complexes tested, except for Cr(NH3)4GDP and Co(NH3)4GDP, which have larger Ki values, are good substrate analogs for P-enolpyruvate carboxykinase. When the substrate is Mn-GTP, fixed at 0.2 mM at pH 6.0, enzyme activity is stimulated two- to two and a half-fold by Cr(H2O)4GTP. A Dixon plot reveals that the stimulatory effect is saturated at 0.4 mM Cr(H2O)4GTP. The interaction of the enzyme with Cr(H2O)4GTP appears to produce a "memory" effect which is manifest with guanosine nucleotide substrates, but which is not observed with the alternative substrate Mn-ITP.
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PMID:The preparation and characterization of Cr(III) and Co(III) complexes of GDP and GTP and their interactions with avian phosphoenolpyruvate carboxykinase. 334 64

In primary cultures of rat hepatocytes the glucagon-dependent induction of phosphoenolpyruvate carboxykinase was studied in the presence of putative local hormone and substrate modulators which form clear concentration gradients during liver passage such as adenosine, ketone bodies and ammonia. 1) Adenosine inhibited the induction of phosphoenolpyruvate carboxykinase in a concentration-dependent manner between 50 and 200 microM up to 4 h after glucagon application; AMP had similar, adenine, inosine and guanosine had no effect. Adenosine was almost totally metabolized by the liver cells during the first 4 h of the induction period. The inhibitory action of adenosine was also observed using dibutyryl-cAMP or 8-bromo-cAMP as inducer; it could not be prevented by the adenosine receptor antagonist caffeine nor could it be mimicked by the selective adenosine receptor agonist N6-(phenylisopropyl)adenosine. 2) Acetoacetate suppressed the induction of phosphoenolpyruvate carboxykinase in a concentration-dependent manner between 5 and 20mM during the first 4 h after glucagon addition. beta-Hydroxybutyrate showed no effect. Neither starting with acetoacetate nor with beta-hydroxybutyrate did the cell cultures establish the thermodynamic equilibrium between the two compounds. 3) Ammonia did not affect induction of phosphoenolpyruvate carboxykinase at concentrations up to 2mM. Ammonia was converted to urea within the first 4 h; yet it remained at clearly hyperphysiological concentrations in the medium during that period. It is concluded that the glucagon-dependent induction of phosphoenolpyruvate carboxykinase was modulated by the local hormone adenosine via a mechanism not involving adenylate cyclase and by acetoacetate via an unknown mechanism. The inhibitory action of adenosine may, that of acetoacetate can hardly be physiologically relevant.
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PMID:Modulation of the glucagon-dependent induction of phosphoenolpyruvate carboxykinase by adenosine, but not ketone bodies or ammonia in rat hepatocyte cultures. Possible significance for the zonal heterogeneity of liver parenchyma. 344 1

The metabolism of glycerate and aspartate was investigated in perfused rat kidneys. The major pathway active for aspartate metabolism and NH3 production was found to include transamination, and not the purine nucleotide cycle. Pyruvate cycling was identified as a means by which reducing potential is generated in the cytosol for glucose and lactate production from these substrates. Inhibition of mitochondrial pyruvate transport caused an inhibition of glucose production, accumulation of lactate and pyruvate in the perfusate, and a decrease in the [lactate]/[pyruvate] ratio in kidneys perfused with aspartate. These data indicate a role of mitochondrial pyruvate transport in the provision of cytosolic reducing potential. With either aspartate or glycerate, 3-mercaptopicolinic acid (3-MPA) suppressed glucose synthesis and caused accumulation of malate plus fumarate within the kidney. Glucose production from glycerate was much less sensitive to the presence of 3-MPA than was glucose production from aspartate, illustrating a phosphoenolpyruvate carboxykinase (PEPCK)-independent pathway for the cycling of pyruvate. In aspartate-perfused kidneys, the presence of 3-MPA, at concentrations that completely blocked glucose accumulation in the perfusate, did not affect the rate of NH3 production and had only a minor effect on the rate of aspartate uptake. These data allow for an estimation of the rate of pyruvate formation from aspartate of about 1 mumol/min per kidney under conditions of complete PEPCK inhibition. Thus a PEPCK-independent pathway is operative for amino acid oxidation and pyruvate formation in perfused kidneys. The NADP-linked, but not the NAD-linked, 'malic' enzyme activity of the kidney cortex was found to be sufficient to catalyse this estimated rate of pyruvate formation.
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PMID:The involvement of pyruvate cycling in the metabolism of aspartate and glycerate by the perfused rat kidney. 380 Sep 11

1. The activities of gluconeogenic and glycolytic enzymes and the concentrations of citrate, ammonia, amino acids, glycogen, glucose 6-phosphate, acetyl-CoA, lactate and pyruvate were measured in kidney cortex of normal, diabetic, cortisone-treated and growth hormone-treated rats. 2. In kidney cortex of diabetic, cortisone-treated and growth hormone-treated rats the activities of glucose 6-phosphatase (EC 3.1.3.9), fructose 1,6-diphosphatase (EC 3.1.3.11) and phosphopyruvate carboxylase (EC 4.1.1.32) were increased. 3. The activities of glutamate dehydrogenase (EC 1.4.1.3), alanine aminotransferase (EC 2.6.1.2), aspartate aminotransferase (EC 2.6.1.10) and pyruvate carboxylase (EC 6.4.1.1) were increased in diabetic and cortisone-treated rats. In growth hormone-treated rats the activity of aspartate aminotransferase was depressed but those of the other three enzymes were unchanged. 4. The activity of hexokinase (EC 2.7.1.1) was not altered in any of these conditions. Phosphofructokinase (EC 2.7.1.11) activity was depressed only in growth hormone-treated rats. Pyruvate kinase (EC 2.7.1.40) activity was depressed in cortisone-treated and growth hormone-treated rats but unchanged in diabetic rats. 5. Amino acids, acetyl-CoA and glucose 6-phosphate contents were increased in rat kidneys in all these three conditions. Ammonia content was increased in diabetic and cortisone-treated rats but was markedly diminished in growth hormone-treated rats. 6. The [lactate]/[pyruvate] ratio was elevated in diabetic and cortisone-treated rats but unchanged in growth hormone-treated rats. Citrate content was increased in the kidney cortex of diabetic and growth hormone-treated rats but was unchanged in cortisone-treated rats. The activity of ATP citrate lyase (EC 4.1.3.8) was depressed in diabetic and growth hormone-treated rats but was increased in cortisone-treated rats. 7. Glycogen content was moderately elevated in growth hormone-treated rats and markedly elevated in diabetic rats, whereas no change in glycogen content was observed in cortisone-treated rats. Glycogen synthetase (EC 2.4.1.11) activity was unchanged in all these three conditions. Phosphorylase (EC 2.4.1.1) activity was not affected in cortisone-treated rats but was depressed in diabetic and growth hormone-treated rats.
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PMID:Evaluation of the rate-limiting steps in the pathway of glucose metabolism in kidney cortex of normal, diabetic, cortisone-treated and growth hormone-treated rats. 434 56

The early renal metabolic response was studied in rats made acidotic by oral feeding of ammonium chloride. 2 hr after feeding of ammonium chloride there was already significant acidosis. Urinary ammonia also increased after ammonium chloride ingestion and at 1(1/2) hr was significantly elevated. In vitro gluconeogenesis by renal cortical slices was increased at 2 hr and thereafter increased steadily. Ammonia production by the same slices was also increased at 2 hr, but thereafter fell and at 6 hr had decreased to levels which, although higher than those of the control, were lower than those obtained from the rats acidotic for only 2 hr. There was no correlation between in vitro gluconeogenesis and ammonia production by kidney slices from rats during the first 6 hr of acidosis, but after 48 hr of ammonium chloride feeding, these two processes were significantly correlated. The early increase in renal gluconeogenesis was demonstrable with both glutamine and succinate as substrates. The activity of the enzyme phosphoenolpyruvate carboxykinase was increased after 4-6 hr of acidosis. During this time there was a decrease in renal RNA synthesis as shown by decreased uptake of orotic acid-(5)H into RNA. Metabolic intermediates were also measured in quick-frozen kidneys at varying times after induction of acidosis. There was an immediate rise in aspartate and a fall in alpha-ketoglutarate and malate levels. There was never any difference in pyruvate or lactate levels or lactate:pyruvate ratios between control and acidotic rats. Phosphoenolpyruvate rose significantly after 6 hr of acidosis. All the data indicate that increased gluconeogenesis is an early response to metabolic acidosis and will facilitate ammonia production by utilization of glutamate which inhibits the glutaminase I enzyme. The pattern of change in metabolic intermediates can also be interpreted as showing that there is not only enhanced gluconeogenesis, but also that there may be significant increase of activity of glutaminase II as part of the very early response to metabolic acidosis.
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PMID:Renal metabolic response to acid-base changes. II. The early effects of metabolic acidosis on renal metabolism in the rat. 544 47


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