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:2.7.11.11 (AMPK)
12,425 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Glucagon stimulates gluconeogenesis in part by decreasing the rate of phosphoenolpyruvate disposal by pyruvate kinase. Glucagon, via cyclic AMP (cAMP) and the cAMP-dependent protein kinase, enhances phosphorylation of pyruvate kinase, phosphofructokinase, and fructose-1,6-bisphosphatase. Phosphorylation of pyruvate kinase results in enzyme inhibition and decreased recycling of phosphoenolpyruvate to pyruvate and enhanced glucose synthesis. Although phosphorylation of 6-phosphofructo 1-kinase and fructose-1,6-bisphosphatase is catalyzed in vitro by the cAMP-dependent protein kinase, the role of phosphorylation in regulating the activity of and flux through these enzymes in intact cells is uncertain. Glucagon regulation of these two enzyme activities is brought about primarily by changes in the level of a novel sugar diphosphate, fructose 2,6-bisphosphate. This compound is an activator of phosphofructokinase and an inhibitor of fructose-1,6-bisphosphatase; it also potentiates the effect of AMP on both enzymes. Glucagon addition to isolated liver systems results in a greater than 90% decrease in the level of this compound. This effect explains in large part the effect of glucagon to enhance flux through fructose-1,6-bisphosphatase and to suppress flux through phosphofructokinase. The discovery of fructose 2,6-bisphosphate has greatly furthered our understanding of regulation at the fructose 6-phosphate/fructose 1,6-bisphosphate substrate cycle.
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PMID:Regulation by glucagon of hepatic pyruvate kinase, 6-phosphofructo 1-kinase, and fructose-1,6-bisphosphatase. 628 62

The inhibition of hepatocyte 6-phosphofructo-1-kinase by glucagon was suppressed by insulin when the enzyme was measured in crude extracts. However, no effect of either hormone was observed after the removal of allosteric effectors from the enzyme, suggesting that the alterations in activity may be due to changes in the level of fructose 2,6-bisphosphate, a potent allosteric activator of the enzyme. Insulin opposed the action of both glucagon and exogenous cyclic AMP to lower fructose 2,6-bisphosphate levels. The concentration of glucagon and of cyclic AMP that gave a half-maximal decrease in fructose 2,6-bisphosphate levels was increased in the presence of 10 nM insulin from 0.03 to 0.09 nM and from 12 to 36 microM, respectively. Insulin also counteracted the effect of maximal concentrations of epinephrine on fructose 2,6-bisphosphate levels. In the presence of 0.02 nM glucagon or 10 microM epinephrine, 10 nM insulin enhanced 6-phosphofructo-2-kinase and decreased fructose 2,6-bisphosphatase activity in (NH4)2SO4-treated hepatocyte extracts. The bifunctional enzyme 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase was shown to be a substrate for the cAMP-dependent protein kinase but not for phosphorylase kinase. It was concluded that insulin opposed the action of glucagon and epinephrine by affecting the phosphorylation state of 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase. Fructose 2,6-bisphosphate levels were decreased in liver cells from diabetic rats. Addition of 30 mM glucose elevated fructose 2,6-bisphosphate levels in cells from fed and 24-h-starved rats but not in cells from diabetic rats. This was probably due to decreases in both 6-phosphofructo-2-kinase and glucokinase activity in the diabetic state. These results show that insulin has both short and long term effects on fructose 2,6-bisphosphate metabolism in liver.
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PMID:The action of insulin on hepatic fructose 2,6-bisphosphate metabolism. 629 99

Bovine cardiac glycogen-free glycogen synthase I reacts with oxidized glutathione at low temperature to partially inactivate the enzyme. Evidence is presented that a mixed disulfide between glutathione and the enzyme is formed in this reaction. A short incubation of the GSSG-treated enzyme with dithiothreitol restores full enzyme activity. The reaction with GSSG is pH dependent and the product is quite stable at neutral pH. Oxidation of one sulfhydryl group in glycogen synthase is associated with a loss of 60-70% of the enzyme activity. Further modification of protein sulfhydryls has less effect on the enzyme activity. Other low molecular weight disulfides also inactivate glycogen synthase and treatment with [35S]cystine to produce a 40% loss of enzyme activity gave rise to a single major radioactive peptide after cyanogen bromide digestion. Thus the GSSG-mediated inactivation of glycogen synthase apparently occurs through a single reactive sulfhydryl group that forms a mixed disulfide with low molecular weight disulfide molecules. Uridine 5'-diphosphate glucose and glycogen prevent the inactivation of glycogen-free glycogen synthase with GSSG, and glucose 6-phosphate retards the rate of inactivation. Reduction and reactivation of the GSSG-oxidized glycogen synthase is not affected by glycogen and it occurs readily at neutral pH with dithiothreitol, mercaptoethanol, or cysteamine. Oxidation of the reactive sulfhydryl group with GSSG has no effect on the rate of glycogen synthase phosphorylation by the catalytic subunit of cAMP-dependent protein kinase.
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PMID:Specific mixed disulfide formation with purified bovine cardiac glycogen synthase I and glutathione. 629 40

Forskolin (40 microM) stimulated adenylate cyclase activities of bovine thyroid plasma membranes without the addition of guanine nucleotides. GDP had little effect on the forskolin-stimulated adenylate cyclase activity while Gpp[NH]p (0.1-1.0 microM) decreased it. In the presence of TSH (10 mU/0.11), Gpp[NH]p no longer caused inhibition. Forskolin did not affect phosphodiesterase activities of thyroid homogenates. Forskolin (10 microM) rapidly increased cAMP levels in bovine thyroid slices both in the absence and presence of a phosphodiesterase inhibitor. The effect of TSH (50 mU/ml) on cAMP levels was additive or greater than additive to that of forskolin. An initial 2-h incubation of slices with forskolin did not decrease their subsequent cAMP responses to either forskolin and/or TSH while similar treatment of slices with TSH induced desensitization of the cAMP response to TSH, but not to forskolin. Forskolin (10 microM) as well as TSH (50 mU/ml) activated cAMP-dependent protein kinase of slices in the absence of a phosphodiesterase inhibitor. Although forskolin activated the adenylate cyclase cAMP system, it did not stimulate iodide organification or glucose oxidation, effects which have been attributed to cAMP. In fact, forskolin inhibited these parameters and 32P incorporation into phospholipids as well as their stimulation by TSH. These results indicate that an increase in cAMP levels and cAMP-dependent protein kinase activity in thyroid slices may not necessarily reproduce the effects of TSH on the thyroid.
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PMID:Effects of forskolin on adenylate cyclase, cyclic AMP, protein kinase and intermediary metabolism of the thyroid gland. 629 78

In hepatocytes 32P-incorporation into rat liver phosphofructokinase is stimulated by glucose as well as by glucagon, the effects of both stimuli being prevented by L-alanine [Eur. J. Biochem. (1982) 122, 175]. The phosphopeptides of the enzyme derived from limited proteolysis by subtilisin and from exhaustive tryptic digestion were analyzed either by one-dimensional mapping on sodium dodecyl sulphate-polyacrylamide slab gels and by fingerprint mapping, respectively. It is shown that in vivo stimulation of 32P-incorporation by glucose or by glucose plus glucagon results in identical phosphopeptide maps, and that these maps were identical with those obtained from phosphofructokinase phosphorylated in vitro with catalytic subunit of cAMP-dependent protein kinase. It is concluded that in the intact liver cell phosphofructokinase is phosphorylated by cAMP-dependent protein kinase but that the state of phosphorylation is modified by metabolite control.
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PMID:Metabolite-controlled phosphorylation of hepatic phosphofructokinase proceeds by cAMP-dependent protein kinase. 629 95

The diastereomeric forms of adenosine cyclic 3',5'-phosphorothioate, Rp cAMPS and Sp cAMPS, were studied in isolated hepatocytes from fed rats for their ability to interact with the intracellular cAMP-dependent protein kinase and to affect the phosphorylase kinase-phosphorylase glycogenolytic cascade. Incubation of the cells with increasing concentrations of Sp cAMPS produced a concentration-dependent activation of cAMP-dependent protein kinase with a concomitant increase in the glycogenolytic rate. Half-maximal and maximal velocities of glycogenolysis were reached at 8 X 10(-7) and 1 X 10(-5) M Sp cAMPS, respectively. Incubation of the cells with 10(-9) to 10(-4) M Rp cAMPS had no effect on basal glucose production or on cAMP-dependent protein kinase activity. Incubation of the cells simultaneously with 3 X 10(-6) M Sp cAMPS and increasing concentrations of Rp cAMPS produced half-maximal inhibition of glycogenolysis at 1 X 10(-5) M Rp cAMPS and maximal inhibition at 1 X 10(-4) M. The concentrations of Sp cAMPS required for half-maximal and maximal activation of glycogenolysis were increased 10-fold when 1 X 10(-5) M Rp cAMPS was present. These data imply that Sp cAMPS is a cAMP-agonist while Rp cAMPS is a cAMP-antagonist.
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PMID:Inhibition of glycogenolysis in isolated rat hepatocytes by the Rp diastereomer of adenosine cyclic 3',5'-phosphorothioate. 631 39

A phosphoprotein phosphatase has been purified from rat liver cytosol. The purification involved chromatography on DEAE-cellulose. Sephacryl S-200, fast protein liquid chromatography (FPLC) and sucrose density gradient centrifugation. It resulted in an almost homogeneous enzyme with a relative molecular mass, Mr, of 90 000 by gel filtration and sucrose gradient centrifugation and Mr = 44 500 by sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS-PAGE). Therefore it seems to be a dimeric enzyme. This protein phosphatase (termed PFK-phosphatase) is completely dependent on Mg2+, which can be replaced partly by Mn2+. It can be eluted from DEAE-cellulose with 120 mM NaCl, is not affected by Ca2+, 100 microM trifluoperazine or the heat-stable inhibitor-2. Inhibition occurs with phosphate, ammonium sulfate and fluoride. PFK-phosphatase dephosphorylates preferentially the alpha subunit of phosphorylase kinase (alpha/beta dephosphorylation ratio 5-10). Phosphorylase a, mixed histone and casein do not serve as substrates. The enzyme dephosphorylates effectively the key enzymes of glucose metabolism 6-phosphofructo-1-kinase, fructose 1,6-bisphosphatase, pyruvate kinase and 6-phosphofructo-2-kinase. Using this protein phosphatase and the catalytic subunit of cAMP-dependent protein kinase, a complete phosphorylation, dephosphorylation and rephosphorylation cycle was possible with 6-phosphofructo-1-kinase as substrate.
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PMID:Purification and characterization of a protein phosphatase from rat liver acting on key enzymes of glucose metabolism. 632 87

The ability of the Rp diastereomer of adenosine cyclic 3',5'-phosphorothioate (Rp cAMPS) to inhibit glucagon-induced glycogenolysis was studied in hepatocytes isolated from fed rats. Preincubation of the cells for 20 min with progressively higher concentrations of Rp cAMPS followed by a 1 X 10(-9) M glucagon challenge resulted in a 50% inhibition of glucose production over a 30-min period at 2-3 X 10(-6) M Rp cAMPS. A maximal inhibition of 50-74% was achieved, the actual value depending upon the length of preincubation with Rp cAMPS. The inhibitory effect did not increase when the concentration of Rp cAMPS was increased from 3 X 10(-6) to 3 X 10(-4) M. Addition of 1 X 10(-5) M Rp cAMPS to the cells followed by 10(-11) to 10(-6) M glucagon shifted the glucagon concentration required for half-maximal glucose production measured at 10 min to 6-fold higher glucagon concentrations and the concentration of glucagon required for apparent maximal glucose production measured at 10 min to greater than 10-fold higher glucagon concentrations. The cAMP-dependent protein kinase activation curve was similarly shifted to higher concentrations of glucagon. These data show that Rp cAMPS acts as a cAMP antagonist capable of opposing the glucagon-induced activation of cAMP-dependent protein kinase and the concomitant activation of the glycogenolytic cascade.
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PMID:Inhibition of glucagon-induced glycogenolysis in isolated rat hepatocytes by the Rp diastereomer of adenosine cyclic 3',5'-phosphorothioate. 633 Jan 2

The effects of adenylate cyclase inhibition on the transport of glucose and fructose and their incorporation into glycogen were investigated in order to assess the extent to which lowered cAMP levels can take part in the various components of glycogen synthesis regulation in isolated rat epididymal adipocytes. The dose-response characteristics of (R)-N-(2-phenylisopropyl)adenosine (PIA), a potent and specific adenylate cyclase inhibitor, on glycogen synthesis were compared with those effectively inhibiting lipolysis, a measure of functional cAMP levels. PIA had no effect on basal glucose or fructose transport but stimulated glucose and fructose incorporation into glycogen. Their respective incorporation was 10 and 69% of that achieved in the presence of insulin. These effects of PIA were shown to be in part the result of increased glycogen synthase I activity. PIA was 20% as effective as insulin in this action. Thus, were insulin to lower cAMP levels and/or inhibit cAMP-dependent protein kinase, this action would be irrelevant to glucose transport but would contribute to the stimulation of glycogen metabolism. However, an additional mechanism(s) involving neither increased glucose transport nor lowered cAMP levels is required to account for the full action of insulin. Fat cells in the absence of medium glucose and in the presence of 10(-7) M PIA and adenosine deaminase constitute a system functionally depleted of cAMP where this mechanism can be studied in isolation.
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PMID:Glycogen synthesis stimulation by adenylate cyclase inhibition in rat epididymal adipocytes. 634 22

Phosphorylation of rat liver glycogen synthase by rabbit skeletal muscle phosphorylase kinase results in the incorporation of approximately 0.8-1.2 mol of PO4/subunit. Analyses of the tryptic peptides by isoelectric focusing and thin layer chromatography reveal the presence of two major 32P-labeled peptides. Similar results were obtained when the synthase was phosphorylated by rat liver phosphorylase kinase. This extent of phosphorylation does not result in a significant change in the synthase activity ratio. In contrast, rabbit muscle glycogen synthase is readily inactivated by rabbit muscle phosphorylase kinase; this inactivation is further augmented by the addition of rabbit muscle cAMP-dependent protein kinase or cAMP-independent synthase (casein) kinase-1. Addition of cAMP-dependent protein kinase after initial phosphorylation of liver synthase with phosphorylase kinase, however, does not result in an inactivation or additional phosphorylation. The lack of additive phosphorylation under this condition appears to result from the phosphorylation of a common site by these two kinases. Partial inactivation of liver synthase can be achieved by sequential phosphorylation with phosphorylase kinase followed by synthase (casein) kinase-1. Under this assay condition, the phosphate incorporation into the synthase is additively increased and the synthase activity ratio (-glucose-6-P/+glucose-6-P) is reduced from 0.95 to 0.6. Nevertheless, if the order of the addition of these two kinases is reversed, neither additive phosphorylation nor inactivation of the synthase is observed. Prior phosphorylation of the synthase by phosphorylase kinase transforms the synthase such that it becomes a better substrate for synthase (casein) kinase-1 as evidenced by a 2- to 4-fold increase in the rate of phosphorylation. This increased rate of phosphorylation of the synthase appears to result from the rapid phosphorylation of a site neighboring that previously phosphorylated by phosphorylase kinase.
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PMID:Phosphorylation of rat liver glycogen synthase by phosphorylase kinase. 642 35


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