Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.2 (PDK1)
 2,238 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The proportion of active (dephosphorylated) pyruvate dehydrogenase in perfused rat heart was decreased by alloxan-diabetes or by perfusion with media containing acetate, n-octanoate or palmitate. The total activity of the dehydrogenase was unchanged. 2. Pyruvate (5 or 25mM) or dichloroacetate (1mM) increased the proportion of active (dephosphorylated) pyruvate dehydrogenase in perfused rat heart, presumably by inhibiting the pyruvate dehydrogenase kinase reaction. Alloxan-diabetes markedly decreased the proportion of active dehydrogenase in hearts perfused with pyruvate or dichloroacetate. 3. The total activity of pyruvate dehydrogenase in mitochondria prepared from rat heart was unchanged by diabetes. Incubation of mitochondria with 2-oxo-glutarate plus malate increased ATP and NADH concentrations and decreased the proportion of active pyruvate dehydrogenase. The decrease in active dehydrogenase was somewhat greater in mitochondria prepared from hearts of diabetic rats than in those from hearts of non-diabetic rats. Pyruvate (0.1-10 mM) or dichloroacetate (4-50 muM) increased the proportion of active dehydrogenase in isolated mitochondria presumably by inhibition of the pyruvate dehydrogenase kinase reaction. They were much less effective in mitochondria from the hearts of diabetic rats than in those of non-diabetic rats. 4. The matrix water space was increased in preparations of mitochondria from hearts of diabetic rats. Dichloroacetate was concentrated in the matrix water of mitochondria of non-diabetic rats (approx. 16-fold at 10 muM); mitochondria from hearts of diabetic rats concentrated dichloroacetate less effectively. 5. The pyruvate dehydrogenase phosphate phosphatase activity of rat hearts and of rat heart mitochondria (approx. 1-2 munit/unit of pyruvate dehydrogenase) was not affected by diabetes. 6. The rate of oxidation of [1-14C]pyruvate by rat heart mitochondria (6.85 nmol/min per mg of protein with 50 muM-pyruvate) was approx. 46% of the Vmax. value of extracted pyruvate dehydrogenase (active form). Palmitoyl-L-carnitine, which increased the ratio of [acetyl-CoA]/[CoA] 16-fold, inhibited oxidation of pyruvate by about 90% without changing the proportion of active pyruvate dehydrogenase.
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PMID:Regulation of pyruvate dehydrogenase in rat heart. Mechanism of regulation of proportions of dephosphorylated and phosphorylated enzyme by oxidation of fatty acids and ketone bodies and of effects of diabetes: role of coenzyme A, acetyl-coenzyme A and reduced and oxidized nicotinamide-adenine dinucleotide. 18 Sep 74

1. The conversion of inactive (phosphorylated) pyruvate dehydrogenase complex into active (dephosphorylated) complex by pyruvate dehydrogenase phosphate phosphatase is inhibited in heart mitochondria prepared from alloxan-diabetic or 48h-starved rats, in mitochondria prepared from acetate-perfused rat hearts and in mitochondria prepared from normal rat hearts incubated with respiratory substrates for 6 min (as compared with 1 min). 2. This conclusion is based on experiments with isolated intact mitochondria in which the pyruvate dehydrogenase kinase reaction was inhibited by pyruvate or ATP depletion (by using oligomycin and carbonyl cyanide m-chlorophenylhydrazone), and in experiments in which the rate of conversion of inactive complex into active complex by the phosphatase was measured in extracts of mitochondria. The inhibition of the phosphatase reaction was seen with constant concentrations of Ca2+ and Mg2+ (activators of the phosphatase). The phosphatase reaction in these mitochondrial extracts was not inhibited when an excess of exogenous pig heart pyruvate dehydrogenase phosphate was used as substrate. It is concluded that this inhibition is due to some factor(s) associated with the substrate (pyruvate dehydrogenase phosphate complex) and not to inhibition of the phosphatase as such. 3. This conclusion was verified by isolating pyruvate dehydrogenase phosphate complex, free of phosphatase, from hearts of control and diabetic rats an from heart mitochondria incubed for 1min (control) or 6min with respiratory substrates. The rates of re-activation of the inactive complexes were then measured with preparations of ox heart or rat heart phosphatase. The rates were lower (relative to controls) with inactive complex from hearts of diabetic rats or from heart mitochondria incubated for 6min with respiratory substrates. 4. The incorporation of 32Pi into inactive complex took 6min to complete in rat heart mitocondria. The extent of incorporation was consistent with three or four sites of phosphorylation in rat heart pyruvate dehydrogenase complex. 5. It is suggested that phosphorylation of sites additional to an inactivating site may inhibit the conversion of inactive complex into active complex by the phosphatase in heart mitochondria from alloxan-diabetic or 48h-starved rats or in mitochondria incubated for 6min with respiratory substrates.
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PMID:Conversion of inactive (phosphorylated) pyruvate dehydrogenase complex into active complex by the phosphate reaction in heart mitochondria is inhibited by alloxan-diabetes or starvation in the rat. 21 16

1. The ;initial activity' of the pyruvate dehydrogenase enzyme complex in whole tissue or mitochondrial extracts of lactating rat mammary glands was greatly decreased by 24 or 48h starvation of the rats. Injection of insulin and glucose into starved rats 60min before removal of the glands abolished this difference in ;initial activities'. 2. The ;total activity' of the enzyme complex in such extracts was revealed by incubation in the presence of free Mg(2+) and Ca(2+) ions (more than 10 and 0.1mm respectively) and a crude preparation of pig heart pyruvate dehydrogenase phosphatase. Starvation did not alter this ;total activity'. It is assumed that the decline in ;initial activity' of the enzyme complex derived from the glands of starved animals was due to increased phosphorylation of its alpha-subunit by intrinsic pyruvate dehydrogenase kinase. 3. Starvation led to an increase in intrinsic pyruvate dehydrogenase kinase activity in both whole tissue and mitochondrial extracts. Injection of insulin into starved animals 30min before removal of the lactating mammary glands abolished the increase in pyruvate dehydrogenase kinase activity in whole-tissue extracts. 4. Pyruvate (1mm) prevented ATP-induced inactivation of the enzyme complex in mitochondrial extracts from glands of fed animals. In similar extracts from starved animals pyruvate was ineffective. 5. Starvation led to a decline in activity of pyruvate dehydrogenase phosphatase in mitochondrial extracts, but not in whole-tissue extracts. 6. These changes in activity of the intrinsic kinase and phosphatase of the pyruvate dehydrogenase complex of lactating rat mammary gland are not explicable by current theories of regulation of the complex.
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PMID:The mode of regulation of pyruvate dehydrogenase of lactating rat mammary gland. Effects of starvation and insulin. 21 55

1. The interconversion of pyruvate dehydrogenase between its inactive phosphorylated and active dephosphorylated forms was studied in skeletal muscle. 2. Exercise, induced by electrical stimulation of the sciatic nerve (5/s), increased the measured activity of (active) pyruvate dehydrogenase threefold in intact anaesthetized rated within 2 min. No further increase was seen after 15 min of stimulation. 3. In the perfused rat hindquarter, (active) pyruvate dehydrogenase activity was decreased by 50% in muscle of starved and diabetic rats. Exercise produced a twofold increase in its activity in all groups; however, the relative differences between fed, starved and diabetic groups persisted. 4. Perfusion of muslce with acetoacetate (2 mM) decreased (active) pyruvate dehydrogenase activity by 50% at rest but not during exercise. 5. Whole-tissue concentrations of pyruvate and citrate, inhibitors of (active) pyruvate dehydrogenase kinase and (inactive) pyruvate dehydrogenase phosphate phosphatase respectively, were not altered by excerise. A decrease in the ATP/ADP ratio was observed, but did not appear to be sufficient to account for the increase in (active) pyruvate dehydrogenase activity. 6. The results suggest that interconversion of the phosphorylated and dephosphorylated forms of pyruvate dehydrogenase plays a major role in the regulation of pyruvate oxidation by eomparison of enzyme activity with measurements of lactate oxidation in the perfused hindquarter [see the preceding paper, Berger et al. (1976)] suggest that pyruvate oxidation is also modulated by the concentrations of substrates, cofactors and inhibitors of (active) pyruvate dehydrogenase activity.
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PMID:Glucose metabolism in perfused skeletal muscle. Pyruvate dehydrogenase activity in starvation, diabetes and exercise. 82 12

Full activation of rat liver pyruvate dehydrogenase in vitro by ADP was prevented by palmitoyl-CoA at a concentration sufficiently low to preclude substrate effects secondary to its oxidation by mitochondria. Activation of pyruvate dehydrogenase by ADP in livers of fat-fed rats was less than in the control animal. The results are consistent with the experiments demonstrating an inhibition of adenine nucleotide translocase and on increased intramitochondrial ATP/ADP ratio by palmitoyl-CoA which could account for the effect on pyruvate dehydrogenase. Inactivation of brain pyruvate dehydrogenase by ATP was also diminished by palmitoyl-CoA indicating that the effect was at the level of the adenine nucleotides rather than at either the pyruvate dehydrogenase kinase or phosphatase enzymes.
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PMID:Interrelationship in the regulation of pyruvate dehydrogenase and adenine-nucleotide translocase by palmitoyl-CoA in isolated mitochondria. 86 23

The effect of severe insulin-induced hypoglycemia on the activity of the pyruvate dehydrogenase enzyme complex (PDHC) was investigated in homogenates of frozen rat cerebral cortex during burst suppression EEG, after 10, 30, and 60 min of isoelectric EEG, and after 30 and 180 min and 24 h of recovery following 30 min of hypoglycemic coma. Changes in PDHC activity were correlated to levels of labile organic phosphates and glycolytic metabolites. In cortex from control animals, the rate of [1-14C]pyruvate decarboxylation was 7.1 +/- 1.3 U/mg of protein, or 35% of the total PDHC activity. The activity was unchanged during burst suppression EEG whereas the active fraction increased to 81-87% during hypoglycemic coma. Thirty minutes after glucose-induced recovery, the PDHC activity had decreased by 33% compared to control levels, and remained significantly depressed after 3 h of recovery. This decrease in activity was not due to a decrease in the total PDHC activity. At 24 h of recovery, PDHC activity had returned to control levels. We conclude that the activation of PDHC during hypoglycemic coma is probably the result of an increased PDH phosphatase activity following depolarization and calcium influx, and allosteric inhibition of PDH kinase due to increased ADP/ATP ratio. The depression of PDHC activity following hypoglycemic coma is probably due to an increased phosphorylation of the enzyme, as a consequence of an imbalance between PDH phosphatase and kinase activities. Since some reduction of the ATP/ADP ratio persisted and since the lactate/pyruvate ratio had normalized by 3 h of recovery, the depression of PDHC most likely reflects a decrease in PDH phosphatase activity, probably due to a decrease in intramitochondrial Ca2+.
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PMID:Changes in pyruvate dehydrogenase complex activity during and following severe insulin-induced hypoglycemia. 198 96

The effect of thiamine triphosphate (ThTP) and thiamine diphosphate (ThDP) on the activity of rat liver pyruvate dehydrogenase complex regulatory enzymes (kinase and phosphatase) was studied in experiments with isolated enzyme preparations. It is shown that ThDP caused a pronounced activation of pyruvate dehydrogenase phosphatase (Ka is equal to 65.0 nM). ThTP inhibits phosphatase competitively against the substrate--the phosphorylated pyruvate dehydrogenase complex. The both thiamine phosphates inhibit the pyruvate dehydrogenase kinase activity almost similarly in concentrations exceeding 10 microM. The physiological significance of the antagonistic action of ThDP and ThTP on the pyruvate dehydrogenase phosphatase activity is discussed.
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PMID:[Effect of thiamine phosphates on the activity of regulatory enzymes of the pyruvate dehydrogenase complex]. 282 86

The putative mediator of intracellular insulin action has been assayed quantitatively by its ability to increase the activity of solubilized pyruvate dehydrogenase (PDH) phosphatase. Conversion of soluble beef heart PDH b to PDH a by PDH phosphatase increased when incubation was carried out in the presence of a crude insulin mediator fraction generated from insulin-treated adipose tissue or liver plasma membranes. Increased PDH phosphatase activity was proportional to the concentration of added insulin mediator. Mediator generation was rapid, with a half-time of approximately 45 sec and was insulin dose dependent. Half-maximal mediator activity was produced at 0.3 nM added insulin, with maximal activity being generated at approximately 3 nM insulin. Mediator activity was significantly decreased at 7 nM insulin, but was increased 4-fold after ethanol extraction. Mediator behaved as an activator of PDH phosphatase, apparently by abolishing the inhibitory effects of ATP on phosphatase activity, but had no effect on PDH kinase activity. The assay of insulin mediator activity described here can be carried out under standardized conditions, in contrast to previously described methods using particulate mitochondrial preparations.
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PMID:Assay of insulin mediator activity with soluble pyruvate dehydrogenase phosphatase. 298 64

The regulatory properties of the Ca2+-sensitive intramitochondrial enzymes (pyruvate dehydrogenase phosphate phosphatase, NAD+-isocitrate dehydrogenase and 2-oxoglutarate dehydrogenase) in extracts of rat liver mitochondria appeared to be essentially similar to those described previously for other mammalian tissues. In particular, the enzymes were activated severalfold by Ca2+, with half-maximal effects at about 1 microM-Ca2+ (K0.5 value). In intact rat liver mitochondria incubated in a KCl-based medium containing 2-oxoglutarate and malate, the amount of active, non-phosphorylated, pyruvate dehydrogenase could be increased severalfold by increasing extramitochondrial [Ca2+], provided that some degree of inhibition of pyruvate dehydrogenase kinase (e.g. by pyruvate) was achieved. The rates of 14CO2 production from 2-oxo-[1-14C]glutarate at non-saturating, but not at saturating, concentrations of 2-oxoglutarate by the liver mitochondria (incubated without ADP) were similarly enhanced by increasing extramitochondrial [Ca2+]. The rates and extents of NAD(P)H formation in the liver mitochondria induced by non-saturating concentrations of 2-oxoglutarate, glutamate, threo-DS-isocitrate or citrate were also increased in a similar manner by Ca2+ under several different incubation conditions, including an apparent 'State 3.5' respiration condition. Ca2+ had no effect on NAD(P)H formation induced by beta-hydroxybutyrate or malate. In intact, fully coupled, rat liver mitochondria incubated with 10 mM-NaCl and 1 mM-MgCl2, the apparent K0.5 values for extramitochondrial Ca2+ were about 0.5 microM, and the effective concentrations were within the expected physiological range, 0.05-5 microM. In the absence of Na+, Mg2+ or both, the K0.5 values were about 400, 200 and 100 nM respectively. These effects of increasing extramitochondrial [Ca2+] were all inhibited by Ruthenium Red. When extramitochondrial [Ca2+] was increased above the effective ranges for the enzymes, a time-dependent deterioration of mitochondrial function and ATP content was observed. The implications of these results on the role of the Ca2+-transport system of the liver mitochondrial inner membrane are discussed.
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PMID:Characterization of the effects of Ca2+ on the intramitochondrial Ca2+-sensitive enzymes from rat liver and within intact rat liver mitochondria. 300 Mar 55

The specificities of pyruvate dehydrogenase kinase and pyruvate dehydrogenase phosphatase were probed using synthetic peptides corresponding to the sequence around phosphorylation sites 1 and 2 on pyruvate dehydrogenase [Tyr-His-Gly-His-Ser(P1)-Met-Ser-Asp-Pro-Gly-Val-Ser(P2)-Tyr-Arg]. The dephosphotetradecapeptide containing aspartic acid at position 8 was a better substrate for the kinase than was the tetradecapeptide containing asparagine at position 8. The apparent Km and V values for the two peptides were 0.43 and 6.1 mM and 2.7 and 2.4 nmol of 32P incorporated/min/mg, respectively. Methylation of the aspartic acid residue also increased the apparent Km of the tetradecapeptide about 14-fold. These results indicate that an acidic residue on the carboxyl-terminal side of phosphorylation site 1 is an important specificity determinant for the kinase. Phosphate was incorporated only into site 1 of the synthetic peptide by the kinase. The phosphatase exhibited an apparent Km of 0.28 mM and a V of 2.3 mumol of 32P released/min/mg for the phosphorylated tetradecapeptide containing aspartic acid. Methylation of the aspartic acid residue had no significant effect on dephosphorylation. The octapeptide and phosphooctapeptide produced by cleavage of the aspartyl-prolyl bond by formic acid were poorer substrates for the kinase and phosphatase than were the tetradecapeptide and phosphotetradecapeptide, respectively. Modification of the amino terminal by acetylation or lysine addition had only a slight effect on the kinase and phosphatase activities.
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PMID:Synthetic peptide substrates for mammalian pyruvate dehydrogenase kinase and pyruvate dehydrogenase phosphatase. 300 77


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