Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.11 (AMPK)
 12,425 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Modifications in characteristics and activities of beta-adrenergic receptors and certain parameters of the cyclic nucleotide systems were observed in the hypertrophied heart of the rat chronically treated with T4. These include: 1) an increased number of beta-adrenergic receptors without a change in their affinity, as determined by binding of (-)-[3H]dihydroalprenolol to the membrane; 2) increased sensitivity and magnitude of stimulation of adenylate cyclase in homogenates by isoproterenol, without a change in the basal or NaF-stimulated (total) enzyme activity; 3) decreased formation of cAMP and decreased activation of cAMP-dependent protein kinase in the minced heart stimulated by isoproterenol, probably due to decreased myocardial ATP concentration; 4) decreased activity of cAMP phosphodiesterase in the particulate fraction; 5) decreased activity of cGMP-dependent protein kinase in both the soluble and particulate fractions, accompanied by decreased activity of cAMP-dependent protein kinase in the particulate fraction; 6) decreased activity of the stimulatory modulator of cGMP-dependent protein kinase and, conversely, increased activity of the inhibitory modulator of cAMP-dependent protein kinase; and 7) increased sensitivity accompanied by decreased maximum tension development of the ventricular strip to contract in response to isoproterenol. These alterations largely disappeared upon regression of the hyperthyroid state. It is suggested that the above changes, many of which were the opposite of those reported earlier for the desensitized and hypertrophied rat heart caused by isoproterenol, may in part consitute the molecular basis for the reputed catecholamine supersensitivity of the heart in the hyperthyroid state.
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PMID:Thyroxine-induced changes in characteristics and activities of beta-adrenergic receptors and adenosine 3',5'-monophosphate and guanosine 3',5'-monophosphate systems in the heart may be related to reputed catecholamine supersensitivity in hyperthyroidism. 624 45

cGMP-dependent protein kinase from bovine lung is labile to specific proteolysis. Limited digestion with chymotrypsin produces a 65,000-dalton monomer and a 16,000-dalton dimer from a 150,000-dalton dimeric enzyme. The larger proteolytic fragment represents the COOH-terminal portion of the enzyme and contains the catalytic site along with the cGMP binding site. The smaller fragment representing the NH2-terminal portion of the enzyme contains the autophosphorylation site and the interchain disulfide bond(s). A model defining the functional domains of cGMP-dependent protein kinase is presented and comparisons with cAMP-dependent protein kinase regulatory subunit are discussed.
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PMID:Structural analysis of cGMP-dependent protein kinase using limited proteolysis. 624 42

The regulatory subunit of the type I cAMP-dependent protein kinase (RI) can be separated into multiple forms on isoelectric focusing gels. RI from bovine skeletal muscle gives rise to bands at pI = 5.57 and 5.45. Phosphate determinations indicate that the more acidic band contains protein-bound phosphate. RI from rat skeletal muscle can be separated into three bands of pI = 5.57, 5.45, and 5.35. The two acidic forms comigrate with labeled RI isolated from rat soleus muscles that were incubated with [32P]orthophosphate. RI from bovine muscle is isolated mainly in the unphosphorylated state while that from rat muscle is primarily phosphorylated. At lest 4 mol of phosphate can be incorporated into each RI dimer following extensive phosphorylation by cGMP-dependent protein kinase in vitro. Two phosphopeptides are observed on polyacrylamide gels following partial proteolysis of in vitro phosphorylated rat RI. One of these peptides is also observed following proteolysis of rat RI phosphorylated in intact soleus muscles.
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PMID:Studies on the phosphorylation of the type I cAMP-dependent protein kinase. 625 Oct 86

The regional and cellular distribution of guanosine 3',5'-cyclic monophosphate (cGMP)-dependent protein kinase (ATP:protein phosphotransferase,EC 2.7.1.37) in mammalian brain was examined by use of the photoaffinity label 8-azidoinosine 3',5'-cyclic monophosphate. Of the regions examined, cerebellum had by far the highest concentration of this enzyme. The cellular localization of cGMP-dependent protein kinase within the cerebellum was determined by examination of mutant mice missing specific types of cerebellar neurons. Mutant mice lacking Purkinje cells had greatly reduced amounts of cGMP-dependent protein kinase, whereas the loss of another cell type, granule cells, did not reduce cGMP-dependent protein kinase levels. By using the same strains of mutant mice, a 23,000-dalton soluble cerebellar substrate for cGMP-dependent protein kinase was also shown to be enriched in Purkinje cells. In contrast, the concentration of type I 3',5'-cyclic AMP-dependent protein kinase in the cerebellum was unaffected by the absence of Purkinje cells and only slightly reduced by the absence of granule cells. The enrichment in Purkinje cells of the cGMP-dependent protein kinase and its substrate suggests an important role for cGMP and cGMP-dependent protein phosphorylation in the function of this type of neuronal cell.
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PMID:Localization of cyclic GMP-dependent protein kinase and substrate in mammalian cerebellum. 625 89

Homogenous regulatory subunit from rabbit skeletal muscle cAMP-dependent protein kinase (isozyme I) was partially hydrolyzed with low (1 g/1300 g) or high (1 g/6 g) concentrations of trypsin. After treatment with low trypsin two main peptides (Mr = 35,000 and 12,000) were produced. The cAMP-binding activity (2 mol cAMP/mol of subunit monomer) was recovered in the monomeric Mr = 35,000 peptide. The ability of either fragment to inhibit catalytic subunit activity was lost. Treatment of the regulatory subunit with a high concentration of trypsin yielded three main fragments (Mr = 32,000, 16,000, and 6,000) which could be resolved by Sephadex G-75 and purified further on DEAE-cellulose columns. One of the peptides (Mr = 32,000) bound 2 mol cAMP/mol fragment. The Mr = 16,000 fragment was very labile and bound cAMP with an undetermined stoichiometry. Cyclic AMP dissociation curves for the native regulatory subunit and its Mr = 32,000 component were similar and suggested the presence of two nonidentical binding sites in each monomer. Using the same procedure, the Mr = 16,000 fragment or homogenous cGMP-dependent protein kinase appeared to contain a single type of binding site. Purified Mr = 32,000 fragment was readily converted to the Mr = 16,000 fragment using high trypsin as assessed by protein bands on SDS-disc gels or by following transfer of radioactivity from Mr = 32,000 peptide covalently labeled with 8-N3-[32P] cAMP to radiolabeled Mr = 16,000 fragment. The smallest regulatory subunit fragment (Mr = 6,000) did not bind cAMP, but was dimeric and could be part of the dimerization domain in the native protein. A model is presented to explain the possible structural-functional relationships of the regulatory subunit.
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PMID:Studies of functional domains of the regulatory subunit from cAMP-dependent protein kinase isozyme I. 625 20

Interaction of cGMP-dependent protein kinase with histones H2A, H2B, H3, and H4, or poly(L-arginine) resulted in changes in enzyme conformation such that inactivation of cGMP binding and activation of basal catalytic activity (assayed without cGMP) occurred. Total kinase activity as determined by phosphorylation of exogenous substrates subsequently decreased, but autophosphorylation of the enzyme was enhanced. The reaction was specific for nucleosome core histones and poly(L-arginine); H1, troponin, and poly(L-lysine) had no effect. Inactivation of cyclic nucleotide binding sites followed pseudo-first order kinetics and, at various histone concentrations, exhibited saturation kinetics at low ionic strength (2 mM potassium phosphate, pH 6.8), but non-saturation kinetics at higher ionic strength (37.5 mM potassium phosphate, pH 6.8, 12.5 mM MgCl2). Saturation kinetics was observed with poly(L-arginine) at both low and high ionic strength. Kinetic parameters measured under saturation conditions were determined for each core histone and poly(L-arginine). Core histones and poly(L-arginine) were noncompetitive inhibitors of cGMP binding; core histones and poly(L-arginine) interacted competitively at an enzyme site designated as the poly(L-arginine) binding site. Regulatory subunits of cAMP-dependent protein kinase contain a similar poly(L-arginine) binding site. Modulator proteins bind to poly(L-arginine) or arginyl residues in histone to prevent interaction with the poly(L-arginine) binding site on the enzymes. Through this mechanism, modulator proteins maintain cyclic nucleotide dependency and full enzyme activity.
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PMID:Regulation of cyclic nucleotide-dependent protein kinase activity by histones and poly(L-arginine). 625 84

A protein that exhibits greater substrate specificity for cGMP-dependent protein kinase than for cAMP-dependent protein kinase has been purified 8,000-fold from cytosol of rabbit cerebellum to apparent homogeneity as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The protein, termed G-substrate, is a monomer of 23,000 daltons. It is heterogeneous on isoelectric focusing, exhibiting three isoelectric forms over the pH range of 5.2-5.6 cGMP-dependent protein kinase catalyzes the incorporation of 2 mol of phosphate/mol of G-substrate, both into threonine residues. The protein has a high content of aspartate, glutamate, and proline. The hydrodynamic properties, heat stability, and acid solubility of this protein are consistent with an unfolded, nonglobular structure. G-substrate is localized primarily in the cytosol of cerebellum, although low concentrations of a phosphorylated protein with a similar molecular weight are detected in other brain regions.
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PMID:A specific substrate from rabbit cerebellum for guanosine 3':5'-monophosphate-dependent protein kinase. I. Purification and characterization. 625 70

Kinetic studies on the activity of purified cGMP-dependent protein kinase and catalytic subunit of cAMP-dependent protein kinase have been carried out using a protein termed G-substrate (see preceding paper) as the phosphate acceptor. Each enzyme catalyzed the phosphorylation of 2.0-2.1 mol of 32P/mol of G-substrate, with phosphorylation occurring primarily at threonine residues. When phosphorylation was carried out in the simultaneous presence of the two enzymes, the stoichiometry increased only slightly, to a value of 2.4, suggesting that both enzymes phosphorylated the same two sites. Initial rate studies on the phosphorylation of G-substrate by cGMP-dependent protein kinase yielded a Km of 0.21 microM and a Vmax of 2.2 mumol/min/mg. Similar studies with the cAMP-dependent protein kinase yielded a Km of 5.8 microM and a Vmax of 2.3 mumol/min/mg. cGMP-dependent protein kinase thus exhibited a high degree of specificity towards this substrate which was apparently based on selective substrate binding rather than catalytic efficacy. The activity of cGMP-dependent protein kinase towards G-substrate was maximal at pH 7.5-8.0 and a Mg2+ concentration of 1-3 mM. Activity declined sharply at high ionic strength (greater than 20 mM KCl).
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PMID:A specific substrate from rabbit cerebellum for guanosine 3':5'-monophosphate-dependent protein kinase. II. Kinetic studies on its phosphorylation by guanosine 3':5'-monophosphate-dependent and adenosine 3':5'-monophosphate-dependent protein kinases. 625 71

The regulatory subunit of the type I cAMP-dependent protein kinase (RI), isolated from bovine or rat skeletal muscle, can be phosphorylated both in vitro (Geahlen, R. L., and Krebs, E. G. (1980) J. Biol. Chem. 255, 1164-1169) and in vivo (Geahlen, R. L., and Krebs, E. G. (1980) J. Biol. Chem. 255, 9375-9379). The effects of each modification on the ability of RI to associate with the catalytic subunit (C) and with cAMP are compared. The phosphorylation of bovine RI in vitro by cGMP-dependent protein kinase results in a loss of inhibitory activity toward C and in the loss of one of two cAMP binding sites per RI monomer. Inhibitory activity can be regained upon removal of the phosphate with potato acid phosphatase. Similar effects are not observed for the subunits phosphorylated in vivo. A comparison of unmodified bovine RI with RI modified in vivo reveals no differences in their interactions with either C or cAMP. Dephosphorylation of purified rat RI also does not affect its association with C or subsequent activation by cAMP. Dephosphorylated rat RI is not a substrate for C but can be slowly phosphorylated by cGMP-dependent protein kinase. The phosphorylation of RI in isolated rat soleus muscles is not inhibited by cycloheximide, ruling out the possibility that only nascent polypeptide chains are substrates for phosphorylation in vivo.
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PMID:Effect of phosphorylation on the regulatory subunit of the type I cAMP-dependent protein kinase. 626 Aug 3

The rat heart contains a large amount of two protein kinase inhibitors. The type I inhibitor blocks the activity of cAMP-dependent protein kinase while the type II inhibitor blocks cGMP-dependent protein kinase, cAMP-dependent protein kinase and cyclic nucleotide-independent protein kinase. Isoproterenol produced a dose-dependent decrease of the type I inhibitor activity and an increase of cAMP content in rat heart. The decrease of the type I inhibitor activity is mediated through adrenergic beta receptor because propranolol and practolol but not phentolamine prevented the effects of isoproterenol. Moreover, prior treatment with aminophylline markedly enhanced isoproterenol-induced increase of cAMP content and decrease of the type I inhibitor activity. Isoproterenol did not change the activity of the type II inhibitor. These results are compatible with the hypothesis that the neurotransmitter generated cAMP modulates phosphorylation in the heart by changing the relationship between cAMP-dependent protein kinase and the type I inhibitor activity.
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PMID:Modulation of the activity of endogenous protein kinase inhibitors in rat heart by the beta adrenergic receptor. 626 7


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