Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:4.1.1.49 (phosphoenolpyruvate carboxykinase)
4,654 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Oxygen modulates the glucagon-dependent activation of the phosphoenolpyruvate carboxykinase (PCK) gene. The respiratory chain or heme proteins have been proposed to function as O2-sensors. The functions of the respiratory chain are impaired by uncouplers such as 2,4-dinitrophenol (DNP); those of ferro-heme proteins are affected by carbon monoxide (CO), which locks heme in the oxy conformation. Therefore, the effects of different concentrations of CO and DNP on the glucagon-dependent induction of PCK mRNA and PCK activity were investigated at different physiological oxygen tensions in primary rat hepatocyte cultures. The cells were cultured under standard conditions from 4-24 h. After addition of fresh media PCK was induced with 1 nM glucagon. PCK mRNA and PCK activity were elevated after 2h and 3h, respectively, to 100% at 16% O2 (mimicking arterial oxygen tensions) and to about 60% at 8% O2 (mimicking venous oxygen tensions). CO counteracted the reduced induction at lower oxygen tensions: Under 8% O2 + 2% CO PCK mRNA could be elevated again to about 90% and PCK activity to about 80%. CO did not impair the induction by insulin of ornithine decarboxylase (ODC) and the incorporation of 14C-leucine into total protein. CO did not cause lactate dehydrogenase (LDH) to leak from the cells or influence the cell structures at the microscopical level. DNP (50 microM) unspecifically lowered PCK gene expression without affecting its modulation by oxygen. These results are in line with the proposal that a ferro-heme protein rather than the respiratory chain acted as an O2 sensor in the activation of the PCK gene.
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PMID:A ferro-heme protein senses oxygen levels, which modulate the glucagon-dependent activation of the phosphoenolpyruvate carboxykinase gene in rat hepatocyte cultures. 837 14

Rabbit proximal tubule cells in primary culture revert from gluconeogenesis to glycolysis. To determine whether glucose and insulin deprivation of the culture medium could prevent this metabolic conversion without a loss of differentiation, rabbit proximal tubule cells were cultured in hormonally defined medium free of glucose and insulin and compared to rabbit proximal tubule cells cultured in medium supplemented with 17.5 mM glucose and 5 micrograms/ml insulin. In the two culture conditions, RPT cells grew at a similar rate and reached confluency within 4-5 days. Patterns of enzyme activity, including brush-border hydrolases, N-acetyl-beta-D-glucosaminidase and glutathione-S-transferases as a function of culture time were comparable in the two media. During the growth phase in glucose- and insulin-free medium, cells showed higher sodium-dependent glucose uptake. Scanning electron microscopy revealed a high density of microvilli at confluency regardless of the culture conditions. In both the presence and absence of glucose and insulin, the activities of gluconeogenic enzymes, phosphoenolpyruvate carboxykinase and fructose-1,6-bisphosphatase, as well as basal and pyruvate-stimulated glucose production fell markedly as a function of time. By contrast, glucose and insulin deprivation greatly reduced both the lactate production rate and the activities of glycolytic enzymes, pyruvate kinase, hexokinase and lactate dehydrogenase.
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PMID:Effect of glucose and insulin deprivation on differentiation and carbohydrate metabolism of rabbit proximal tubular cells in primary culture. 838 35

We investigated whether the multiple pathophysiological signals generated in a peritonitis septic model alter the mRNA levels of glycolytic and gluconeogenic enzymes, and whether these alterations are associated with glucose dyshomeostasis. Rats were sham-operated in the control group, and peritonitis sepsis was produced by a 1 cm cecal incision in the septic group. At 2, 4, and 6 hr post-surgery, total cellular RNAs were isolated from livers, and Northern blots performed to measure mRNA levels of aldolase B (ADL), lactate dehydrogenase (LDH), pyruvate kinase (PK), phosphoenolpyruvate carboxykinase (PEPCK), and glucokinase (GK). Hepatic PEPCK enzymatic activity was measured by condensing 14CO2 with phosphoenolpyruvate (PEP) to form malate. Serum glucose concentrations were also measured. We found the following: At 2 hr of peritonitis sepsis, serum glucose concentrations, mRNA levels of all enzymes, and PEPCK enzymatic activity increased over control levels. At 4 hr of peritonitis sepsis, serum glucose concentrations and mRNA levels of GK and PK continued to increase; mRNA levels of all other enzymes, as well as PEPCK enzymatic activity decreased to or below control levels. At 6 hr of peritonitis sepsis, serum glucose concentrations, mRNA levels of all enzymes, and PEPCK enzymatic activity decreased to or below control levels. We concluded that sepsis affects mRNA levels of glycolytic and gluconeogenic enzymes at the transcriptional level, and that these alterations are associated with glucose dyshomeostasis.
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PMID:Altered levels of mRNA encoding enzymes of hepatic glucose metabolism in septic rats. 840 44

The presence of phosphoenolpyruvate (PEP) carboxylase (EC 4.1.1.31), an enzyme at the branchpoint of glycolysis and the Krebs cycle was detected in the Filaria Molinema dessetae. This enzyme has not previously been identified in Helminths, which have so far been found to only possess a phosphoenolpyruvate carboxykinase (EC 4.1.1.32). This enzyme had a level of activity comparable to that of pyruvate kinase, and was relatively less active than enzymes such as malate dehydrogenase or lactate dehydrogenase. We propose here a method of purification of M. dessetae PEP-carboxylase. When purified to electrophoretic homogeneity, the enzyme had a molecular weight of 64 kDa. Kinetic studies indicated that the carboxylation reaction had an optimal pH of 5.8. The enzyme was inhibited by cations such as Fe2+, Zn2+, Cd2+, Cu2+ but required the presence of Mg2+ or Mn2+. The enzyme was thermostable. The apparent Km value of 2.38 mmol for phosphoenolpyruvate for the carboxylation reaction was higher than previously reported values. The Km value for KHCO3 was found to be 1.6 mmol. PEP-carboxylase did not catalyse the reverse reaction.
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PMID:Purification and properties of phosphoenolpyruvate carboxylase from Molinema dessetae (Nematoda: Filarioidea). 847 1

Cytoplasmic fractions from species of the Mollicutes genera Entomoplasma, Mesoplasma, Mycoplasma, and Acholeplasma were assayed for NADH oxidase (NADH ox), ATP- and PPi-dependent phosphofructokinase (PFK), ATP- and PPi-dependent deoxyguanosine kinase (dGUOK), thymidine kinase (TK), TMP kinase (TMPK), glucose-6-phosphate dehydrogenase (G6Pde), lactate dehydrogenase (LDH), malate dehydrogenase (MDH), phosphoenolpyruvate carboxylase, hypoxanthine-guanine phosphoribosyl transferase, dUTPase, and uracil-DNA glycosylase (UNG) activities. Membrane fractions were also examined for NADH ox activity. These activities were used as indicators of the presence and relative activities of major Mollicutes metabolic and DNA repair pathways. This was the first study to determine the presence of these enzymes in members of the genera Entomoplasma and Mesoplasma. Using the data obtained, we constructed a preliminary scheme for distinguishing genera of the class Mollicutes on the basis of the results of signature functional enzyme assays. This scheme includes phylogenetic relationships deduced from rRNA analyses, but is more informative with respect to metabolic potential. The criteria used include the presence of PPi-dependent PFK, urease, dUTPase, and dGUOK activities. Entomoplasma ellychniae ELCN-1T (T = type strain), Entomoplasma melaleucae M-1T, Mesoplasma seiffertii F7T, Mesoplasma entomophilum TACT, Mesoplasma florum L1T, Mycoplasma fermentans PG18T, and Acholeplasma multilocale PN525T were similar in most respects. NADH ox activity was localized in the cytoplasm of these organisms. These strains had ATP-dependent PFK, MDH, LDH, ATP- and PPi-dependent dGUOK, and UNG activities, but not dUTPase or G6Pde activities. In contrast, Acholeplasma equifetale C112T, Acholeplasma oculi 19LT, Acholeplasma hippikon C1T, Acholeplasma modicum PG49T, and Acholeplasma morum 72-043T had membrane-localized NADH ox activity, PPi-dependent PFK, G6Pde, and dUTPase activities, and significantly lower MDH and LDH activities and exhibited a faster rate with PPi than with ATP in the dGUOK reaction. All of the members of the Mollicutes tested had hypoxanthine-guanine phosphoribosyl transferase, phosphoenolpyruvate carboxylase, and (except for Mesoplasma entomophilum TAC(T)) UNG activities. All of the Acholeplasma strains except Acholeplasma multilocale PN525T had TK, TMPK, and UNG activities. Mesoplasma entomophilum TAC(T) was distinguished by having no detectable dUTPase, UNG, TK, and TMPK activities, indicating that there is a severe restriction in or an absence of a synthetic route to dTTP. Our data also suggest that A. multilocale PN525T is a member of an unrecognized metabolic subgroup of the genus Acholeplasma or is not an Acholeplasma strain.
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PMID:Comparative metabolism of Mesoplasma, Entomoplasma, Mycoplasma, and Acholeplasma. 886 14

In vivo, bicarbonate can affect proximal tubule intermediary metabolism, including gluconeogenesis, ammoniagenesis and maintenance of the mitochondrial substrate supply. In vitro, rabbit proximal tubule cells (RPTC) in primary culture revert from gluconeogenesis to glycolysis and their mitochondrial metabolism remains lower than in vivo. To determine whether the bicarbonate buffer system could have an effect on these deregulations, RPTC in primary culture grown in the absence of insulin and glucose in the culture medium were developed either with the standard sodium bicarbonate buffer with 5% CO2 or with a Hepes hydrogen ion buffer in the presence of 0.5% CO2. Duration of the bicarbonate-free cultures was increased until at least day 17 after seeding, compared with day 11 in bicarbonate-buffered cultures. As could be expected, succinate dehydrogenase activity remained stable as a function of time in bicarbonate-free cultures while an early marked decrease of this activity occurred from seeding in cultures developed in the presence of bicarbonate buffer. Compared to bicarbonate-buffered cells, higher phosphoenolpyruvate carboxykinase activity concomitant with lower intracellular lactate dehydrogenase activity was observed in cultures developed in the absence of bicarbonate, which is indicative of closer carbohydrate metabolism orientation to the in vivo situation for RPTC. Immunofluorescence staining of RPTC with monoclonal antibodies directed to neutral endopeptidase (NEP), and dipeptidyl-peptidase IV (DPP II) showed similar extensive labelling with DPP and NEP in both culture conditions. Confocal microscopy analysis of NEP subcellular distribution, showed exclusive targetting of NEP to the apical plasma membranes. In both models, cAMP production was stimulated by parathyroid hormone and unaffected by arginine vasopressin. In conclusion, bicarbonate withdrawal from the culture medium (without changing the pH of the medium) allows a marked improvement of mitochondrial capacity and carbohydrate metabolism pattern without any loss of differentiated properties.
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PMID:Effects of the medium HCO3-/CO2 buffer system on differentiation and intermediary metabolism properties of rabbit proximal tubule cells in primary culture. 897 88

The activities of 18 enzymes involved in the intermediary and energy metabolism were measured in certain widely-spread peracarid crustaceans: 3 hypogean (Niphargus virei, Niphargus rhenorhodanensis and Stenasellus virei) and 2 epigean (Gammarus fossarum and Asellus aquaticus) ones. The activities of numerous enzymes were correlated with the known metabolic rates of the 5 species. Such rates are reduced in hypogean organisms: levels of enzymatic activity in subterranean species were 1.2 to 8.6 times lower than in epigean species for the main key regulatory enzymes involved in the Krebs cycle and glycolysis (phosphofructokinase, pyruvate kinase, hexokinase and citrate synthetase). The relative activities of phosphofructokinase, glycogen phosphorylase and hexokinase clearly indicated that glycogen was the main fuel oxidized in both epigean and hypogean organisms. A higher glycogen phosphorylase/hexokinase ratio in hypogean than in epigean crustaceans showed that subterranean species had a greater ability to function anaerobically. The presence of high activities of glutamate-pyruvate transaminase and lactate dehydrogenase in all species (and of malate dehydrogenase and fumarase in hypogean species) was indicative of a coupled fermentation of glycogen and glutamate during anaerobiosis, with lactate and alanine as end-products (as well as succinate in hypogean species). A low fructose-1,6-bisphosphatase/phosphofructokinase ratio, associated with a low level of phosphoenolpyruvate carboxykinase activity, indicated that the glycolytic pathway was active and that gluconeogenic ability was limited in epigean crustaceans. In contrast, in hypogean species, association of a higher ratio and a high level of phosphoenolpyruvate carboxykinase activity suggested a low glycolytic activity and a high gluconeogenic ability.
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PMID:The activities of enzymes associated with the intermediary and energy metabolism in hypogean and epigean crustaceans. 909 Nov 76

A response in heat shock protein 70 (hsp 70) expression in the beta-naphthoflavone (BNF) treated rainbow trout (Oncorhynchus mykiss) corresponded to altered metabolic status of the liver as evidenced by the lower phosphoenolpyruvate carboxykinase (PEPCK), lactate dehydrogenase and 3-hydroxyacylcoA dehydrogenase activities. The BNF-induced increase in hsp70 levels and conjugation enzyme activities (phase I and phase II) were not modified by handling stress. Indeed handling stress did not affect either hsp 70 levels or conjugation enzyme activities in trout liver. The decrease in hepatic PEPCK activity in the BNF group may be responsible for the attenuation of the increase in liver glucose concentration after a 3 min handling stress in this species, suggesting that BNF affects liver gluconeogenic capacity in this species. Handling stress elicited a plasma cortisol and glucose response in both the sham and BNF group, however, the cortisol response with BNF was erratic compared with the sham, implying alterations in the cortisol dynamics post-stress. These results show for the first time that BNF affects cellular metabolic responses to stress and suggests the possibility of using hsp 70 as a biomarker for toxic effects in trout.
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PMID:Handling stress does not affect the expression of hepatic heat shock protein 70 and conjugation enzymes in rainbow trout treated with beta-naphthoflavone. 921 70

Although induction of cytochrome P-450 1A1 (CYP1A1) in the Caco-2 clone TC7 alters glucose utilization and modifies the expression of sucrase-isomaltase (SI) and hexose transporters, nothing is known of the events that control these effects. In this study, we analyzed the effects of beta-naphthoflavone (beta-NF) and hypoxia on these parameters and expression of key enzymes of glucose metabolism. Both beta-NF and hypoxia induce similar changes: 1) induction of CYP1A1 mRNA; 2) increased glucose consumption and lactic acid production and lower glycogen content; 3) downregulation of SI and upregulation of GLUT1 mRNAs; 4) downregulation of fructose-1,6-bisphosphatase and pyruvate kinase mRNAs and upregulation of phosphoenolpyruvate carboxykinase, pyruvate dehydrogenase, lactate dehydrogenase, and phosphofructokinase mRNAs; and 5) upregulation of c-fos and c-jun mRNAs. Although addition of inhibitors of CYP1A1 catalytic activity to beta-NF-treated cells totally inhibits the enzyme activity, it does not modify CYP1A1 mRNA response and associated effects, thus excluding a direct role for the enzyme per se. These results point to a possible physiological implication of the signal-transduction pathway responsible for CYP1A1 induction.
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PMID:Hypoxia and CYP1A1 induction-dependent regulation of proteins involved in glucose utilization in Caco-2 cells. 969 11

The study investigates the influence of different culture conditions on attachment, viability and functional status of rainbow trout (Oncorhynchus mykiss) liver cells in primary culture. Cells were isolated by a two-step collagenase perfusion and incubated in serum-free, chemically defined minimal essential medium (MEM), (a) as a monolayer on uncoated PRIMARIA dishes, (b) as a monolayer on culture dishes coated with calf collagen type 1, and (c) in coculture with the established fish cell lines RTH-149 or RTG-2. Cell attachment was assessed from DNA and protein concentrations per dish, viability was estimated from cellular lactate dehydrogenase release, and the metabolic status was investigated by measuring activities of the phosphoenolpyruvate carboxykinase and biotransformation enzymes as well as the total cytochrome P450 contents. Seeding of hepatocytes on collagen-coated dishes did not alter cell attachment or detachment from the (culture substrate, but had a small, but not significant effect on cell viability and metabolic parameters. Coculture of liver cells and RTG-2 cells reduced hepatocyte detachment from the culture substrate, and it was associated with a significant elevation of 7-ethoxyresorufin-O-deethylase activities in the hepatic cells. Cytochrome P450 contents, however, were not altered. The coculture effect on liver cell physiology clearly depended on the type of cell line, because coculture with RTH-149 cells led to similar, but much weaker effects than obtained in cocultures with RTG-2 cells. Electron microscopical observations revealed the existence of gap junctions and possible exocytosis-like transport between cell lines and hepatocytes. The results point to the potential of coculture systems to improve physiological parameters of trout liver cells in primary culture.
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PMID:Viability and differential function of rainbow trout liver cells in primary culture: coculture with two permanent fish cells. 987 May 25


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