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)

Purified preparations of rat heart pyruvate dehydrogenase kinase have two polypeptides with molecular weights of 48,000 (p48) and 45,000 (p45). Recently, we reported the primary structure of p48 (Popov, K. M., Kedishvili, N. Y., Zhao, Y., Shimomura, Y., Crabb, D. W., and Harris, R. A. (1993) J. Biol. Chem. 268, 26602-26606) and presented evidence that (i) it exhibits kinase activity for pyruvate dehydrogenase and (ii) it belongs to a family of mitochondrial protein kinases unique from other eukaryotic protein kinases. Here, we report the molecular cloning and deduced amino acid sequence of p45. The protein sequence of p45 has 70% identity to the protein sequence of p48. Minor differences exist throughout the protein sequences with the greatest difference occurring at the amino termini. Recombinant p45 protein, expressed in Escherichia coli and purified to homogeneity, catalyzed the phosphorylation and inactivation of kinase-depleted pyruvate dehydrogenase complex, indicating that p45 and p48 correspond to different isoforms of pyruvate dehydrogenase kinase. Northern blot analysis revealed a single hybridizing species of 2.5 kilobases. The highest level of p45 message expression was found in heart and skeletal muscle and the lowest in spleen and lung. Liver, kidney, brain, and testis express intermediate amounts of p45 mRNA. In contrast, p48 mRNA is predominantly expressed in heart, with other tissues expressing only a modest amount of this message. Tissue-specific expression of isoforms of pyruvate dehydrogenase kinase may indicate the existence of tissue-specific mechanisms for the regulation of pyruvate dehydrogenase activity.
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PMID:Molecular cloning of the p45 subunit of pyruvate dehydrogenase kinase. 796 63

Antibodies were raised in rabbits to free rat liver pyruvate dehydrogenase (PDH) kinase alpha-chain and shown to react with PDH kinase alpha-chain in rat heart and liver PDH complexes, in purified pig heart PDH complex and in bovine kidney dihydrolipoamide acetyltransferase-protein X-PDH kinase subcomplex. E.l.i.s.a for PDHE1 (pyruvate dehydrogenase) and PDH kinase have been developed and applied to assays of these proteins in extracts of rat liver and rat heart mitochondria; the measured immunoreactivities for PDHE1 (heart > liver) and for PDH kinase alpha-chain (liver > heart) paralleled known differences in PDH complex and PDH kinase activities respectively. The results of e.l.i.s.a of PDH kinase alpha-chain in extracts of rat liver mitochondria showed that the effects of starvation to increase PDH kinase activity in vivo, and the effects of dibutyryl cyclic AMP or palmitate to increase PDH kinase activity in hepatocytes cultured in vitro, are due largely (> 90%) to an increase in the specific activity of PDH kinase. The effect, in cultured hepatocytes, of dibutyryl cyclic AMP to increase PDH kinase activity was blocked by cycloheximide; the effect of palmitate was blocked by an inhibitor of carnitine palmitoyltransferase I (Etomoxir), but not by cycloheximide.
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PMID:Role of protein synthesis and of fatty acid metabolism in the longer-term regulation of pyruvate dehydrogenase kinase. 801 Sep 47

The present study investigated the effects of chronic food restriction (achieved by limiting access to food to 2 h daily for up to 8 weeks) on the activity of the active form of pyruvate dehydrogenase (PDHa) in liver. Accelerated and exaggerated activation of hepatic PDH in response to a meal, previously demonstrated to occur within 10 days of food restriction, was demonstrated to persist for 4 and 8 weeks of food restriction, despite a food intake of only 50-60% of controls. Activation of hepatic PDH during feeding in rats subjected to food restriction for 4 weeks was dependent on continued food intake. As a consequence, hepatic PDHa activities in food-restricted rats were suppressed relative to controls for 19 h of the 24 h daily cycle. Curve-fitting by second-order polynomial regression analysis demonstrated a significant positive correlation between hepatic PDHa activity and lipogenic rate over the range of PDHa activities observed during the 2 h feeding period. Increased lipogenesis during feeding in food-restricted rats was not at the expense of hepatic glycogen synthesis or deposition; measurement of concurrent rates of glycogenesis and lipogenesis revealed simultaneous flux through both pathways, but specific activation of lipogenesis. The accelerated re-activation of hepatic PDH observed within 1 h of feeding in rats subjected to 4 weeks of food restriction was facilitated by a failure of the 22 h interprandial fasting period to induce a stable increase in hepatic PDH kinase activity. The present study indicates differential regulation of hepatic PDH kinase activity during periods of food withdrawal between food-restricted rats and starved/re-fed control rats. Such regulation occupies a critical role in determining the rate of activation of hepatic PDH during feeding. In turn, increased activity of hepatic PDHa during feeding in food-restricted rats bears a close positive relationship with hepatic lipogenic rate.
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PMID:The regulation of hepatic carbon flux by pyruvate dehydrogenase and pyruvate dehydrogenase kinase during long-term food restriction. 825 Aug 46

The review examines the mechanisms regulating the activities of the two key enzymes determining rates of glucose and fatty acid oxidation, i.e., the pyruvate dehydrogenase (PDH) complex and the carnitine palmitoyltransferase (CPT) system. The review also evaluates the regulatory importance of gene expression in the control of tissue fuel selection within the context of substrate competition between glucose and fatty acids. It identifies a strong indirect input of nutrient-gene interactions in the control of pyruvate oxidation through the regulated provision of pyruvate as a substrate for PDH and as an inhibitor of PDH kinase. Nutrient-gene interactions are also identified in relation to the regulation of CPT I activity by malonyl-CoA (inhibitor) and by the provision of long-chain acyl-CoA (substrate/activator), the latter via the hydrolysis of plasma or tissue triacylglycerol (by lipoprotein lipase and hormone-sensitive lipase, respectively). We discuss how such regulation is reinforced by long-term modulation of PDH kinase-specific activity and CPT I maximal activity. We also explore the role of mechanisms operating at the levels of the PDH complex and the CPT system that act to promote and accelerate a switch in fuel utilization once a committed change in nutrient supply has been established. In particular, we discuss the regulatory influences exerted by altered sensitivities of PDH kinase to inhibition by pyruvate and CPT I to inhibition by malonyl-CoA, respectively.
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PMID:Interactive regulation of the pyruvate dehydrogenase complex and the carnitine palmitoyltransferase system. 829 90

Mammalian pyruvate dehydrogenase kinase binds to the lipoyl domain region of the core structure forming dihydrolipoyl acetyltransferase (E2) subunits. The bound kinase has a greatly enhanced rate in phosphorylating E2-bound pyruvate dehydrogenase (E1) tetramers versus the rate at which resolved kinase phosphorylates dissociated E1. This E2-activated kinase function was completely prevented by selective alkylation of reduced lipoyl groups while kinase and E1 binding to the E2 core were retained. Selective removal of lipoyl cofactors from intact E2 by treatment with Enterococcus faecalis lipoamidase decreased kinase activity by 4-fold and caused selective release of a major portion of the kinase from E2 in a sucrose-step gradient procedure. Selective and reversible modification of the lipoyl groups of E2 subunits also allowed the kinase to be dissociated under mildly chaotropic conditions. Thus, the lipoyl prosthetic group on one of the two lipoyl domains of E2 subunits is critically important for maintaining E2-activated kinase function and contributes to binding of the kinase to E2. Since removal of the lipoyl group weakened kinase binding to E2 more than modifying lipoyl thiols, it is suggested that the hydrophobic inner portion of the lipoyl conjugate (i.e., lysine carbons and C1 to C5 of the lipoic acid) is important in the binding of the kinase.
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PMID:Critical role of a lipoyl cofactor of the dihydrolipoyl acetyltransferase in the binding and enhanced function of the pyruvate dehydrogenase kinase. 843 47

Despite significant increases in circulating concentrations of lipid fuels (triacylglycerol, non-esterified fatty acids (NEFA) and ketone bodies) in late-pregnant rats sampled in the fed (absorptive) state, cardiac and skeletal muscle active pyruvate dehydrogenase (PDHa) activities remained comparable with those observed in fed, age-matched virgin controls. Cardiac PDHa activity was suppressed in response to acute (6 h) starvation in late-pregnant (as well as virgin) rats: this inactivation was opposed by inhibition of mitochondrial long-chain FA oxidation. Starvation (6 h) also led to PDH inactivation in skeletal muscles of late-pregnant, but not virgin, rats. Starvation for 24 h led to further suppression of cardiac PDHa activity and was associated with significant increases in PDH kinase activities in both virgin and late-pregnant rats. Late pregnancy did not itself influence cardiac PDH kinase activity.
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PMID:Control of muscle pyruvate oxidation during late pregnancy. 847 40

The pyruvate dehydrogenase (PDH) complex undergoes reversible phosphorylation catalyzed by a PDH kinase (inactivating) and a PDH phosphatase (activating). In skeletal muscle, a decreased proportion of active PDH (PDHa) complex limits glucose oxidation in insulin-deficient states. The time-course for reactivation of the PDH complex by insulin in skeletal muscle of diabetic rats is important to understanding the potential mode of the action of insulin in regulating glucose metabolism. A single injection of insulin (1 U/kg) completely reversed the effects of alloxan-diabetes on PDHa activity within 1 hour. The normalization of the effects of diabetes on PDHa activity by insulin was maintained for a minimum of 6 hours. The increase in PDHa activity occurred before an insulin-induced decrease in plasma free fatty acids levels, demonstrating a dissociation between the antilipolytic effects of insulin and its ability to activate the PDH complex. PDH kinase activity was not normalized to control values following a single injection of insulin. Therefore, acute (1 to 6 hours) insulin-mediated activation of the PDH complex does not result from a decrease in PDH kinase activity. However, longer-term insulin therapy (1 U/kg body weight; twice daily) restored both PDHa and PDH kinase activities. The results are consistent with the hypothesis that activation of the PDH complex immediately following insulin administration is not mediated by a decreased PDH kinase activity. However, with daily insulin therapy in diabetes, activation of the PDH complex results from decreased PDH kinase activity.
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PMID:Insulin-induced activation of pyruvate dehydrogenase complex in skeletal muscle of diabetic rats. 849 17

The provision of a high-fat diet (47% of energy as fat) for 28 days led to a significant increase in hepatic pyruvate dehydrogenase kinase activity, together with significant suppression of hepatic pyruvate dehydrogenase (active form). An enhanced hepatic pyruvate dehydrogenase kinase activity continued to be observed at 6 h after the withdrawal of the high-fat diet. Significant suppression of hepatic pyruvate dehydrogenase kinase activity was observed in post-absorptive, high-fat-fed rats after a 2.5 h euglycaemic-hyperinsulinaemic clamp, such that differences in pyruvate dehydrogenase kinase activities between control and high-fat-fed rats were no longer evident. Starvation for 24 h in rats previously maintained on standard diet also evoked a substantial increase in hepatic pyruvate dehydrogenase kinase activity. This latter response was only partially reversed by 2.5 h of euglycaemic hyperinsulinaemia. Suppression of pyruvate dehydrogenase kinase activity by 2.5 h euglycaemic hyperinsulinaemia in high-fat-fed rats was associated with a substantial increase in hepatic pyruvate dehydrogenase activity (active form) whereas no significant increase in hepatic pyruvate dehydrogenase activity (active form) was observed after 2.5 h euglycaemic hyperinsulinaemia in 24 h-starved rats. The results are consistent with the proposition that hepatic pyruvate dehydrogenase kinase responds directly to an increase in lipid oxidation which is facilitated by insulin deficiency or an impaired action of insulin.
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PMID:Regulation of hepatic pyruvate dehydrogenase kinase by insulin and dietary manipulation in vivo. Studies with the euglycaemic-hyperinsulinaemic clamp. 867 48

Ranolazine has shown anti-anginal efficacy in humans and cardiac anti-ischaemic activity in models, but without affecting haemodynamics or baseline contraction. In isolated normoxic rat hearts, Langendorff-perfused for 30 min with 11 mM glucose, 3% albumin, and 0.4 mM or 0.8 mM palmitate, 20 microM ranolazine significantly increased active, dephosphorylated, pyruvate dehydrogenase (PDHa), but not with no palmitate or 1.2 mM palmitate. Dichloroactetate (DCA, 1 mM), a PDHa kinase inhibitor, significantly increased PDHa in hearts perfused with 0, 0.4 or 0.8 mM but not 1.2 mM palmitate. PDHa was significantly increased with 1.2 mM palmitate by DCA plus ranolazine, and additive effects were also seen at 0.8 mM palmitate. Activation of PDH by ranolazine and promotion of glucose oxidation offers a plausible means by which the drug may be anti-ischaemic nonhaemodynamically. Extensive studies with extracted enzymes and isolated rat heart mitochondria failed to demonstrate any effects of ranolazine on PDH kinase or phosphatase, or on PDH catalytic activity, whereas effects of other known effectors (such as DCA) were readily demonstrable, suggesting that ranolazine activates PDH indirectly. Further analyses of the hearts revealed that ranolazine reduced acetyl CoA content under all conditions where fatty acid was present, and +/- DCA which itself had little effect. In the absence of fatty acid, ranolazine and/or DCA raised acetyl CoA. In perfusions where octanoate (+/- albumin) replaced palmitate, ranolazine still decreased acetyl CoA, but not when acetate replaced palmitate. In octanoate-perfused hearts, the contents of the C4, C6 and C8 CoA esters were all increased by ranolazine. This is consistent with ranolazine causing an inhibition of fatty acid beta-oxidation leading to decreased acetyl CoA and activation of PDH.
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PMID:Ranolazine increases active pyruvate dehydrogenase in perfused normoxic rat hearts: evidence for an indirect mechanism. 872 66

Fasting inhibits glucose-induced insulin secretion. We investigated the role of a glucose fatty acid cycle for such inhibition and its molecular basis in pancreatic islets from 48-h fasted rats. The fasting-impaired insulin response to 27 mM glucose was restored by 41% with a carnitine palmitoyltransferase I inhibitor, etomoxir. Etomoxir also restored (by 50%) impaired glucose oxidation in islets from fasted rats and increased the ratio of oxidation to glycolytic flux from 33 to 43%. Fasting decreased total pyruvate dehydrogenase (PDH) activity (active, unphosphorylated plus inactive, phosphorylated form) by 29%, as well as the percentage of active form (54 +/- 5 vs. 79 +/- 2% in fed rats, P < 0.001). Fasting increased islet PDH kinase activity as follows: PDH-bound activity by 36% and free (not PDH bound) PDH kinase by 70%. Fasting failed to affect PDH kinase content when assayed by an enzyme-linked immunoabsorbent assay with antibodies raised against 45 kDa PDH kinase alpha-chain. We conclude that fasting impairs B cell function to a major extent through the operation of a glucose fatty acid cycle and that decreased PDH activity resulting from increased specific activity of PDH kinase constitutes an important molecular mechanism.
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PMID:Fasting and decreased B cell sensitivity: important role for fatty acid-induced inhibition of PDH activity. 876 83


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