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:6.4.1.1 (pyruvate carboxylase)
1,516 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. Increasing concentrations of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), a mild respiratory-chain inhibitor [Halestrap (1987) Biochim. Biophys. Acta 927, 280-290], caused progressive inhibition of glucose production from lactate + pyruvate by hepatocytes from starved rats incubated in the presence or absence of oleate and gluconeogenic hormones. 2. No significant changes in tissue ATP content were observed, but there were concomitant decreases in ketone-body output and cytochrome c reduction and increases in NADH fluorescence and the ratios of [lactate]/[pyruvate] and [beta-hydroxybutyrate]/[acetoacetate]. 3. The inhibition by DCMU of palmitoylcarnitine oxidation by isolated liver mitochondria was used to calculate a flux control coefficient of the respiratory chain towards gluconeogenesis. In the presence of 1 mM-oleate, the calculated values were 0.61, 0.39 and 0.25 in the absence of hormone and in the presence of glucagon or phenylephrine respectively, consistent with activation of the respiratory chain in situ as previously suggested [Quinlan & Halestrap (1986) Biochem. J. 236, 789-800]. 4. Cytoplasmic oxaloacetate concentrations were shown to decrease under these conditions, implying inhibition of pyruvate carboxylase. 5. Inhibition of gluconeogenesis from fructose and dihydroxyacetone was also observed with DCMU and was accompanied by an increased output of lactate + pyruvate, suggesting that activation of pyruvate kinase was occurring. With the latter substrate, measurements of tissue ADP and ATP contents showed that DCMU caused a small fall in [ATP]/[ADP] ratio. 6. Two inhibitors of fatty acid oxidation, pent-4-enoate and 2-tetradecylglycidate, were shown to abolish and to decrease respectively the effects of hormones, but not valinomycin, on gluconeogenesis from lactate + pyruvate, without changing tissue ATP content. 7. It is concluded that the hormonal increase in mitochondrial matrix volume stimulates fatty acid oxidation and respiratory-chain activity, allowing stimulation of pyruvate carboxylation and thus gluconeogenesis to occur without major changes in [ATP]/[ADP] or [NADH]/[NAD+] ratios. 8. The high flux control coefficient of the respiratory chain towards gluconeogenesis may account for the hypoglycaemic effect of mild respiratory-chain inhibitors.
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PMID:Evidence that the flux control coefficient of the respiratory chain is high during gluconeogenesis from lactate in hepatocytes from starved rats. Implications for the hormonal control of gluconeogenesis and action of hypoglycaemic agents. 342 47

The decarboxylation of oxalacetate by pyruvate carboxylase in the absence of ADP and Pi is stimulated 400-fold by the presence of oxamate, which is an inhibitory analogue of pyruvate. The observation of substrate inhibition when either oxamate or oxalacetate is varied at a fixed concentration of the other indicates that both molecules bind at the same site on the enzyme. The pH profiles for this reaction show no evidence of the involvement of an enzymic acid-base catalyst, suggesting that the proton and CO2 units may be exchanged directly between the reactants (although CO2 sequestered in the active site may be an intermediate in the process). The pH profiles of the full reverse reaction of pyruvate carboxylase in which oxalacetate decarboxylation is coupled to ATP formation and where Pi is the variable substrate do, however, indicate that such an acid-base catalyst is involved in the other partial reaction of the enzyme in proton transfer to and from biotin. The enzyme also displays two oxamate-independent oxalacetate decarboxylating activities, one of which is biotin-dependent and the other is independent of biotin.
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PMID:Decarboxylation of oxalacetate by pyruvate carboxylase. 381 78

We have previously shown (Comp. Biochem. Physiol. 77B: 35-39, 1984) that a rapid postnatal increase in hepatic mitochondrial adenine nucleotide content activates pyruvate carboxylation and gluconeogenesis in the newborn rabbit. This study investigated factors limiting flux through the gluconeogenic pathway and examined the physiological stimuli responsible for the activation phenomenon. There is a 2.3-fold increase in total mitochondrial adenine nucleotides, along with a threefold increase in the matrix ATP/ADP ratio, by 2 h after birth, resulting overall in a sixfold increase in the amount of ATP/mg mitochondrial protein. Analysis of gluconeogenic intermediates, measured in freeze-clamped livers between birth and 4 h postnatal, suggests that pyruvate carboxylase controls gluconeogenic flux during this period. Newborn rabbits reared in an hypoxic environment (5% O2) exhibited decreased mitochondrial adenine nucleotide content, decreased rates of pyruvate carboxylation, and depressed blood glucose levels compared with littermates reared in room air or 95% O2. Manipulation of the insulin-to-glucagon ratio in vivo by injecting insulin at birth significantly delayed postnatal increases in the mitochondrial adenine nucleotide content and the rate of pyruvate carboxylation. Conversely, glucagon injection produced a supranormal increase in both mitochondrial adenine nucleotide content and pyruvate carboxylation. In addition, insulin injection prevented, whereas glucagon enhanced, the normal postnatal increase in tissue ATP/ADP. These results suggest that tissue oxygenation and a decreased insulin-to-glucagon ratio promote the rapid influx of adenine nucleotides from the liver cytosol into the mitochondrial matrix, thereby activating pyruvate carboxylation and gluconeogenesis during the presuckling period.
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PMID:Regulation of hepatic gluconeogenesis in newborn rabbit: controlling factors in presuckling period. 390 80

1. The kinetics of rat liver pyruvate carboxylase were examined and the effect of various agents as activators or inhibitors determined. 2. Essentially similar results were obtained in comparisons of kinetics determined by a radioactivity method involving extracts of acetone-dried powders from whole livers and with a spectrophotometric assay using partially purified enzyme from the mitochondrial fraction. Activity per g of liver from fed or starved rats assayed under optimum substrate and activator conditions was 3 or 6 mumol of oxaloacetate formed/min at 30 degrees C, respectively. 3. The enzyme exhibited cold-lability and lost activity on standing, even in 1.5m-sucrose. 4. The K(m) towards pyruvate was about 0.33mm and towards bicarbonate 4.2mm. K(m) towards MgATP(2-) was 0.14mm. Mg(2+) ions activated the enzyme, in addition to their role in MgATP(2-) formation. From calculations of likely concentrations of free Mg(2+) in the assay medium a K(a) towards Mg(2+) of about 0.25mm was deduced. Mn(2+) also activated the enzyme as well as Mg(2+), but at much lower concentrations. It appeared to be inhibitory when concentrations of free Mn(2+) as low as 0.1mm were present. 5. Excess of ATP is inhibitory, and this appears at least in part independent of the trapping of Mg(2+). 6. Both Co(2+) and Zn(2+) were inhibitory; 2mol of the latter appeared to be bound even in the presence of excess of Mg(2+) and the inhibition was time-dependent. 7. Ca(2+) inhibited by competition with Mg(2+) (K(i) about 0.38mm). The inhibition due to Ca(2+) was less pronounced when activation was with Mn(2+). Inhibition by Ca(2+) and ATP appeared to be additive. 8. Hill plots suggested that no interactions occurred between ATP-binding sites. Although similar plots for total Mg(2+) gave n=3.6, no conclusions could be drawn due to the chelation of the cation with other components of the assay. Similar difficulties arose in assessing the values for Ca(2+). 9. The enzyme was inactive in the absence of acetyl-CoA and showed a sigmoidal response in its presence. K(a) was about 0.1mm with possibly up to four binding sites. Malonyl-CoA was a competitive inhibitor, with K(i) 0.01mm. 10. There was no apparent inhibition by glucose, glucose 6-phosphate, fructose 6-phosphate, fructose 1,6-diphosphate, acetoacetate, beta-hydroxybutyrate, malate, aspartate, pyruvate, palmitoylcarnitine, octanoate, glutathione, butacaine, triethyltin or potassium chloride under the conditions used. Inhibition was found with citrate (possibly by chelation) and adenosine, and also by phosphoenolpyruvate, AMP, ADP and cyclic AMP, K(i) towards the last four being 0.55, 0.76, 0.25 and 1.4mm respectively.
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PMID:Some aspects of the kinetics of rat liver pyruvate carboxylase. 432 33

1. The activities of pyruvate carboxylase, phosphoenolpyruvate carboxylase and fructose diphosphatase in crude homogenates of vertebrate and invertebrate muscles are reported. 2. Pyruvate carboxylase activity was present in all insect flight muscles that were investigated: in homogenates of bumble-bee flight muscle the activity was inhibited by ADP and activated by acetyl-CoA, and it was distributed mainly in the mitochondrial fraction. This is the first demonstration of pyruvate carboxylase activity in muscle. However, the activity appears to be restricted to insect flight muscle, since it was not found in other invertebrate or vertebrate muscles. 3. Since the three enzymes were never found together in the same muscle, it is concluded that these enzymes cannot provide a pathway for the synthesis of glycogen from lactate or pyruvate in muscle. Other roles for these enzymes in muscle are suggested. In particular, pyruvate carboxylase may be present in insect flight muscle for the provision of oxaloacetate to support the large increase in activity of the tricarboxylic acid cycle which occurs when an insect takes flight.
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PMID:The activities of pyruvate carboxylase, phosphoenolpyruvate carboxylase and fructose diphosphatase in muscles from vertebrates and invertebrates. 435 25

1. With freshly isolated blowfly mitochondria 38% of the intramitochondrial adenine nucleotide was present as AMP. 2. On incubation with oxidizable substrates the AMP and ADP concentrations fell and that of ATP rose; with pyruvate together with proline the ATP concentration reached its maximum value at 6min; with glycerol phosphate the phosphorylation of endogenous nucleotide was more rapid. 3. Addition of the uncoupling agent carbonyl cyanide phenylhydrazone caused a rapid fall of ATP and a parallel rise in ADP, then ADP was converted into AMP. 4. This was in contrast with rat liver mitochondria endogenous AMP concentrations, which were always lower than those of blowfly mitochondria and changed little under different metabolic conditions. 5. Evidence is presented that adenylate kinase (EC 2.7.4.3) has a dual distribution in blowfly mitochondria, a part being located in the matrix space and a part in the space between the outer and inner mitochondrial membranes, as in liver and other mitochondria. 6. The possible regulatory role of changing AMP concentrations in the mitochondrial matrix was investigated. Partially purified pyruvate carboxylase (EC 6.4.1.1) and citrate synthase (EC 4.1.3.7) were inhibited 30% by 2mm-AMP, whereas pyruvate dehydrogenase (EC 1.2.4.1) was unaffected. 7. AMP activated the NAD(+)-linked isocitrate dehydrogenase (EC 1.1.1.41) activity of blowfly mitochondria in the absence of ADP, but in the presence of ADP, AMP caused inhibition. 8. It is suggested that AMP may exert a controlling effect on the oxidative activity of blowfly mitochondria.
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PMID:Changes in intramitochondrial adenine nucleotides in blowfly flight-muscle mitochondria. 437 97

1. Pyruvate carboxylase from baker's yeast is inhibited by ADP, AMP and adenosine at pH8.0 in the presence of magnesium chloride concentrations equal to or higher than the ATP concentration. The adenine moiety is essential for the inhibitory effect. 2. In the absence of acetyl-CoA (an allosteric activator) ADP, AMP and adenosine are competitive inhibitors with respect to ATP. In the presence of acetyl-CoA, besides the effect with respect to ATP, AMP competes with acetyl-CoA, whereas ADP and adenosine are non-competitive inhibitors with respect to the activator. 3. Pyruvate carboxylase is inhibited by NADH. The inhibition is competitive with respect to acetyl-CoA and specific with respect to NADH, since NAD(+), NADP(+) and NADPH do not affect the enzyme activity. In the absence of acetyl-CoA, NAD(+), NADH, NADP(+) and NADPH do not inhibit pyruvate carboxylase. 4. Pyruvate carboxylase is inhibited by ADP, AMP and NADH at pH6.5, in the presence of 12mm-Mg(2+), 0.75mm-Mn(2+) and 0.5mm-ATP, medium conditions similar to those existing inside the yeast cell. The ADP and NADH effects are consistent with a regulation of enzyme activity by the intracellular [ATP]/[ADP] ratio and secondarily by NADH concentration. These mechanisms would supplement the already known control of yeast pyruvate carboxylase by acetyl-CoA and l-aspartate. Inhibition by AMP is less marked and its physiological role is perhaps limited.
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PMID:Effects of adenosine phosphates and nicotinamide nucleotides on pyruvate carboxylase from baker's yeast. 439 Mar 78

1. The reaction pathway for the carboxylation of pyruvate, catalysed by pig liver pyruvate carboxylase, was studied in the presence of saturating concentrations of K(+) and acetyl-CoA. 2. Free Mg(2+) binds to the enzyme in an equilibrium fashion and remains bound during all further catalytic cycles. MgATP(2-) binds next, followed by HCO(3) (-) and then pyruvate. Oxaloacetate is released before the random release, at equilibrium, of P(i) and MgADP(-). 3. This reaction pathway is compared with the double displacement (Ping Pong) mechanisms that have previously been described for pyruvate carboxylases from other sources. The reaction pathway proposed for the pig liver enzyme is superior in that it shows no kinetic inconsistencies and satisfactorily explains the low rate of the ATP[unk][(32)P]P(i) equilibrium exchange reaction. 4. Values are presented for the stability constants of the magnesium complexes of ATP, ADP, acetyl-CoA, P(i), pyruvate and oxaloacetate.
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PMID:Pig liver pyruvate carboxylase. The reaction pathway for the carboxylation of pyruvate. 444 12

1. The fixation of CO(2) by pyruvate carboxylase in isolated rat brain mitochondria was investigated. 2. In the presence of pyruvate, ATP, inorganic phosphate and magnesium, rat brain mitochondria fixed H(14)CO(3) (-) into tricarboxylic acid-cycle intermediates at a rate of about 250nmol/30min per mg of protein. 3. Citrate and malate were the main radioactive products with citrate containing most of the radioactivity fixed. The observed rates of H(14)CO(3) (-) fixation and citrate formation correlated with the measured activities of pyruvate carboxylase and citrate synthase in the mitochondria. 4. The carboxylation of pyruvate by the mitochondria had an apparent K(m) for pyruvate of about 0.5mm. 5. Pyruvate carboxylation was inhibited by ADP and dinitrophenol. 6. Malate, succinate, fumarate and oxaloacetate inhibited the carboxylation of pyruvate whereas glutamate stimulated it. 7. The results suggest that the metabolism of pyruvate via pyruvate carboxylase in brain mitochondria is regulated, in part, by the intramitochondrial concentrations of pyruvate, oxaloacetate and the ATP:ADP ratio.
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PMID:Regulation of pyruvate metabolism via pyruvate carboxylase in rat brain mitochondria. 472 35

1. Metabolism of pyruvate and malate by isolated fat-cell mitochondria incubated in the presence of ADP and phosphate has been studied by measuring rates of pyruvate uptake, malate utilization or production, citrate production and oxygen consumption. From these measurements calculations of the flow rates through pyruvate carboxylase, pyruvate dehydrogenase and citrate cycle have been made under various conditions. 2. In the presence of bicarbonate, pyruvate was largely converted into citrate and malate and only about 10% was oxidized by the citrate cycle; citrate and malate outputs were linear after lag periods of 6-9min and 3min respectively, and no other end products of pyruvate metabolism were detected. On the further addition of malate or hydroxymalonate, the lag in the rate of citrate output was less marked but no net malate disappearance was detected. If, however, bicarbonate was omitted then net malate uptake was observed. Addition of butyl malonate was found to greatly inhibit the metabolism of pyruvate to citrate and malate in the presence of bicarbonate. 3. These results are in agreement with earlier conclusions that in adipose tissue acetyl units for fatty acid synthesis are transferred to the cytoplasm as citrate and that this transfer requires malate presumably for counter transport. They also support the view that oxaloacetate for citrate synthesis is preferentially formed from pyruvate through pyruvate carboxylase rather than malate through malate dehydrogenase and that the mitochondrial metabolism of citrate in fat-cells is restricted. The possible consequences of these conclusions are discussed. 4. Studies on the effects of additions of adenine nucleotides to pyruvate metabolism by isolated fat-cell mitochondria are consistent with inhibition of pyruvate carboxylase in the presence of ADP and pyruvate dehydrogenase in the presence of ATP.
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PMID:Metabolism of pyruvate and malate by isolated fat-cell mitochondria. 515 97


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