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)

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 which has an apparent molecular weight of 240,000 was partially purified (500-fold) from the glycogen-protein complex of rabbit skeletal muscle. The enzyme exhibited broad substrate specificity as it dephosphorylated phosphorylase, phosphohistones, glycogen synthase, phosphorylase kinase, regulatory subunit of cAMP-dependent protein kinase, and phosphatase inhibitor 1. The phosphatase showed high specificity towards dephosphorylation of the beta-subunit of phosphorylase kinase and site 2 of glycogen synthase. With the latter substrate, the presence of phosphate in sites 1a and 1b decreased the apparent Vmax, perhaps by inhibiting the dephosphorylation of site 2. The phosphorylated form of inhibitor 1 did not significantly inhibit this high-molecular-weight phosphatase. However, an inhibitor 1-sensitive phosphatase activity could be derived from this preparation by limited trypsinization. Furthermore, greater than 70% of the phosphatase activity in skeletal muscle extracts and in the glycogen-protein complex was insensitive to inhibitor 1. Limited trypsinization of each fraction obtained from the phosphatase purification increased the total activity (1.5- to 2-fold) and converted the enzyme into a form which was inhibited by inhibitor 1. The results suggest that inhibitor 1-sensitive phosphatase may be a proteolyzed enzyme.
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PMID:Rabbit muscle glycogen-bound phosphoprotein phosphatases: substrate specificities and effects of inhibitor-1. 631 11

A hypersensitivity of glycogen phosphorylase activation by epinephrine and glucagon has been demonstrated in isolated perfused working and non-working hearts from diabetic rats. Accumulation of tissue cAMP and activation of cAMP-dependent protein kinase in response to epinephrine and glucagon were no greater and usually less in hearts of diabetic than of normal rats. Insulin deficiency was not associated with greater changes in epinephrine-induced activation of glycogen phosphorylase kinase than that observed in normal hearts. Perfusion of hearts with subphysiological concentrations of calcium (0.83 mM) partially reversed the diabetes-related hypersensitivity of phosphorylase activation by epinephrine. The phosphorylase activation hypersensitivity to epinephrine was completely reversed by adrenalectomizing diabetic rats 5 days before heart perfusion, an effect potentially caused by steroid-induced changes in cardiac calcium metabolism. These data are consistent with the hypothesis that phosphorylase activation by phosphorylase kinase is allosterically increased in the diabetic due to a diabetes-related increase in free intracellular calcium concentrations.
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PMID:Phosphorylase activation hypersensitivity in hearts of diabetic rats. 632 Jun 71

Yeast phosphorylase is phosphorylated and activated by a cyclic AMP-independent protein kinase (called phosphorylase kinase) and a cyclic AMP-dependent protein kinase. Only in the presence of both kinases is phosphorylase fully activated and phosphorylated. No evidence was found for the presence of two phosphorylation sites as an identical phosphopeptide pattern of phosphorylase is obtained after phosphorylation by either one or both kinases. The kinases probably phosphorylate identical sites but recognize different subunits of phosphorylase. Phosphorylase kinase phosphorylates the high-Mr subunit while cAMP-dependent protein kinase phosphorylates the low-Mr subunit.
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PMID:Regulation of yeast phosphorylase by phosphorylase kinase and cAMP-dependent protein kinase. 635 52

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

Initial autophosphorylation of nonactivated rabbit skeletal muscle phosphorylase kinase at pH 8.0 caused an increase in enzymatic activity that closely paralleled phosphorylation of the beta subunit. Peptide maps revealed that the first phosphate incorporated into the beta subunit during autophosphorylation was on the same tryptic peptide previously isolated from phosphorylase kinase that had been phosphorylated by cAMP-dependent protein kinase (Cohen P., Watson, D.C., and Dixon, G.H. (1975) Eur. J. Biochem. 51, 79-92). When preincubated with phosphorylase kinase for one min, Ca2+ and Mg2+ synergistically stimulated subsequent autophosphorylation at pH 6.8. After this treatment phosphorylation of both the alpha and beta subunits became linear, and the first site phosphorylated on the beta subunit at pH 6.8 corresponded to the first site phosphorylated at pH 8.0. Removal of the lag as a consequence of the synergistic action of the metal ions allowed determination of a Km for MgATP of approximately 20 microM during initial autophosphorylation at either pH 6.8 or 8.2. With phosphorylase b as the substrate the Km values for MgATP under identical conditions were determined to be approximately 30 and 60 microM at pH 6.8 and 8.2, respectively. Initial rates of autophosphorylation over a 30-fold range of phosphorylase kinase concentrations suggest that incorporation of the first 1 to 2 mol of phosphate per alpha beta gamma delta tetramer occurs through an intramolecular mechanism.
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PMID:Characterization of initial autophosphorylation events in rabbit skeletal muscle phosphorylase kinase. 660 53

Polylysine greatly stimulated the autophosphorylation of phosphorylase kinase from rabbit skeletal muscle. When fully autophosphorylated, about 14 mol of phosphate per tetramer (alpha beta gamma delta) were incorporated in the presence of polylysine, which was twice as much as those observed without polylysine. In contrast to this stimulatory effect of polylysine on the autophosphorylation, polylysine strongly inhibited the conversion reaction of phosphorylase b to a. The inhibition is competitive with a Ki of 2.3 micrograms/ml. No effects of polylysine were observed on the activities of phosphorylase and cAMP-dependent protein kinase.
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PMID:Two diverse effects of poly(L-lysine) on rabbit skeletal muscle phosphorylase kinase: stimulation of autophosphorylation and inhibition of its activity. 669 31

A rapid reverse-phase high performance liquid chromatography (HPLC) method is presented for isolating the alpha, beta, gamma, and delta subunits of rabbit muscle phosphorylase kinase. The HPLC separation allows micropreparative purification of all the subunits with 66-88% recoveries. Relative molecular weights of the subunits as determined by sodium dodecyl sulfate gel electrophoresis in 4, 5, 7, and 10% acrylamide are alpha 132,000, alpha' 127,000, beta 113,000 and gamma 43,000. Amino acid compositions are reported for the HPLC purified subunits. alpha contains about 2 mol of endogenous phosphate/mol of protein and beta, gamma, and delta each contain about 1 mol of phosphate/mol of protein. Despite the identity of delta and calmodulin, essentially no protein-bound phosphate was found associated with bovine brain calmodulin. Holophosphorylase kinase contains about 20 mol of endogenous phosphate/mol of protein. The first NH2-terminal sequence analyses of the alpha, beta, and gamma subunits were determined by Tarr manual Edman degradation. Within the NH2-terminal 23 residues of gamma ( TRDAALPGSHSTHGFYENYESKE . . . ) there are six identities and one conservative interchange with the catalytic subunit of bovine cAMP-dependent protein kinase. The first 17 residues of the NH2-terminal sequence of alpha ( MRSRSNSGVRLDSYARL . . . ) exhibit six identities and one conservative interchange with the transforming protein from the Rous sarcoma virus (Schmidt- Rupin strain) provided a single gap is inserted in the src gene product. Further structural information is required to evaluate the significance of these sequence similarities. The beta subunit has a blocked NH2 terminus.
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PMID:High performance liquid chromatography purification and structural characterization of the subunits of rabbit muscle phosphorylase kinase. 672 54

A rapid method for the purification of rat liver phosphorylase kinase 30,000-fold over homogenate values is described. The method allows the isolation of a near homogeneous preparation of phosphorylase kinase initially associated with the glycogen pellet to be accomplished within 24 h. The enzyme has Mr (apparent) = 1.3 million by gel filtration and is composed of subunits similar in size to those of skeletal muscle phosphorylase kinase. The enzyme is phosphorylated by the cAMP-dependent protein kinase: phosphate is incorporated into two of the subunits (Mr = 140,000 and Mr = 116,000) and is closely paralleled by activation of the enzyme. The enzyme is partially inhibited by ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid and is stimulated by 10(-8)-10(-6) M Ca2+. The pH optimum of the nonactivated enzyme is 7.0. Activation by cAMP-dependent protein kinase does not appear to alter the Ca2+ sensitivity of the enzyme. However, it results in a large increase in activity at pH 7 through 8, but not at pH below 6.5. Purified rat liver phosphorylase kinase thus shows many similarities to purified skeletal muscle phosphorylase kinase, but differs in respect to its incomplete inhibition by ethylene glycol bis(beta-amino-ethyl ether)-N,N,N',N'-tetraacetic acid and to the effects of phosphorylation by cAMP-dependent protein kinase on its pH activity profile and Ca2+ sensitivity.
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PMID:Purification of rat liver phosphorylase kinase. 680 57

Separation of phosphorylated sarcoplasmic reticulum (SR) fragments by polyacrylamide gel disc electrophoresis in the presence of Na-DS revealed that the radioactivity is distributed in protein zones with molecular weights of 95,000 and 6000-8000. The phosphorylation of the protein with m. w. of 95,000 is Ca2+-dependent. The tryptic hydrolysis of the phosphorylated SR fragments from fast skeletal muscles results in a loss of radioactivity by 60-70%; phospholipase C from Clostridium welchii reduces the labelled phosphate content by 40-50%. The cAMP-dependent protein kinase inhibitor decreases the phosphorylation of both substrates. The substrate of phosphorylation with m. w. of 6000-8000 is not stained with Amidoschwartz 10B or Coumassie brilliant blue. Extraction by an acidified chlorophorm--methanol mixture results in a proteolipid with specific radioactivity exceeding that of the original preparation of phosphorylated SR membranes 3-4-fold. Thin-layer chromatography on Silufol plates and Silicagel KSK showed that the proteolipid is not chromatographically homogeneous after 2-fold precipitation by diethyl ether and is localized in a band with Rf varying from 0.6 to 0.8. The fluorescence spectrum of the proteolipid in a chlorophorm--methanol--HCl solution is represented by an assymmetrical structure-free band with a maximum at 350 nm. A possible role of phosphorylase b and proteolipid in manifestation of the functional activity of the SR fragments is discussed.
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PMID:[Characterization of endogenous phosphorylation substrates of sarcoplasmic reticulum fragments from fast skeletal muscles of the rabbit]. 711 8


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