Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Adenosine 3',5'-monophosphate (cAMP) dependent protein kinase (EC 2.7.1.37) catalyzes the phosphorylation of serine and threonine residues of a number of proteins according to the following chemical equation: ATP + protein leads to phosphoprotein + ADP. The DEAE-cellulose peak II holoenzyme from bovine brain, which is composed of regulatory and catalytic subunits, is resistant to ethoxyformic anhydride inactivation. After adding cAMP, the protein kinase becomes susceptible to ethoxyformic anhydride inhibition. Ethoxyformic anhydride (2mM) inhibits the enzyme 50% (5 min, pH 6.5, 30 degrees) in the presence of 10 muM cAMP, but less than 5% in its absence. The substrate, Mg2+-ATP, protects against inactivation suggesting that inhibition is associated with modification of the active site. Addition of regulatory subunit or Mg2+-ATP to the isolated catalytic subunit also prevents ethoxyformic anhydride inactivation. These results suggest that the regulatory subunit shields the active site of the catalytic subunit thereby inhibiting it. In contrast to the bovine brain or muscle DEAE-cellulose peak II holoenzyme, the bovine muscle peak I holoenzyme is susceptible to ethoxyformic anhydride inactivation in the absence of cAMP.
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PMID:Bovine brain adenosine 3',5'-monophosphate dependent protein kinase. Mechanism of regulatory subunit inhibition of the catalytic subunit. 24 Apr 4

Purified cyclic adenosine 3':5'-monophosphate (cAMP)-dependent protein kinase of bovine cardiac muscles catalyzes the incorporation of 2 mol of 32P from [gamma-32P]ATP to seryl residues in its cAMP-binding protein. The reaction appears to be catalyzed by the protein kinase itself rather than by a protein kinase kinase and is enhanced by cAMP and by the addition of polyarginine. Phosphorylation of the purified enzyme facilitates its dissociation by cAMP (Erlichman, J., Rosenfeld, R., and Rosen, O.M. (1974) J. Biol. Chem. 249, 5000-5003) but does not affect cAMP binding. At equilibrium, 2 mol of cAMP are bound to both the phospho- and dephospho-enzymes. Phosphorylation of protein kinase is reversible. Upon addition of ADP and Mg2+, phosphate is transferred from the protein to ADP, and ATP is formed. The reverse reaction is optimal at pH 5.5 unlike the forward reaction which has a broad, more alkaline pH activity optimum. It is activated by polyarginine and dependent upon the addition of cAMP to a much greater degree than the forward reaction. The data suggest that the catalytic subunit of protein kinase catalyzes the forward and reverse reactions but do not exclude the possibility that the holoenzyme may also be active. Autophosphorylation by protein kinase and dephosphorylation by phosphrprotein phosphatases of by reverals of the autophosphorylation reaction may regulate the sensitivity of certain protein kinases to activation by cAMP in vivo.
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PMID:Reversible autophosphorylation of a cyclic 3':5'-AMP-dependent protein kinase from bovine cardiac muscle. 24 Aug 40

The role of Ca2+ ions in alpha-adrenergic activation of hepatic phosphorylase was studied using isolated rat liver parenchymal cells. The activation of glucose release and phosphorylase by the alpha-adrenergic agonist phenylephrine was impaired in cells in which calcium was depleted by ethylene glycol bis(beta-aminoethyl ether)N,N'-tetraacetic acid (EGTA) treatment and restored by calcium addition, whereas the effects of a glycogenolytically equivalent concentration of glucagon on these processes were unaffected. EGTA treatment also reduced basal glucose release and phosphorylase alpha activity, but did not alter the level of cAMP or the protein kinase activity ratio (-cAMP/+cAMP) or impair viability as determined by trypan blue exclusion, ATP levels, or gluconeogenic rates. The effect of EGTA on basal phosphorylase and glucose output was also rapidly reversed by Ca2+, but not by other ions. Phenylephrine potentiated the ability of low concentrations of calcium to reactivate phosphorylase in EGTA-treated cells. The divalent cation inophore A23187 rapidly increased phosphorylase alpha and glucose output without altering the cAMP level, the protein kinase activity ratio, and the levels of ATP, ADP, or AMP, The effects of the ionophore were abolished in EGTA-treated cells and restored by calcium addition. Phenylephrine rapidly stimulated 45Ca uptake and exchange in hepatocytes, but did not affect the cell content of 45Ca at late time points. A glycogenolytically equivalent concentration of glucagon did not affect these processes, whereas higher concentrations were as effective as phenylephrine. The effect of phenylephrine on 45Ca uptake was blocked by the alpha-adrenergic antagonist phenoxybenzamine, was unaffected by the beta blocker propranolol, and was not mimicked by isoproterenol. The following conclusions are drawn: (a) alpha-adrenergic activation of phosphorylase and glucose release in hepatocytes is more dependent on calcium than is glucagon activation of these processes; (b) variations in liver cell calcium can regulate phosphorylase alpha levels and glycogenolysis; (c) calcium fluxes across the plasma membrane are stimulated more by phenylephrine than by a glycogenolytically equivalent concentration of glucagon. It is proposed that alpha-adrenergic agonists activate phosphorylase by increasing the cytosolic concentration of Ca2+ ions, thus stimulating phosphorylase kinase.
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PMID:Studies on alpha-adrenergic activation of hepatic glucose output. Studies on role of calcium in alpha-adrenergic activation of phosphorylase. 32 50

A physiologically and biochemically realistic model of the regulation of pyruvate dehydrogenase complex (PDH) was constructed for the perfused rat heart. It includes conversion between inactive (phospho) and active (dephospho) forms by a specific protein kinase (PDHK) and phosphoprotein phosphatase (PDHP). The activity of the tightly bound PDHK is influenced by synergistic activation/inhibition by acetyl CoA/CoASH and NADH/NAD. PDHK in this simulation was more sensitive to the fraction of ADP that was Mg2+-chelated than to the ATP-to-ADP ratio. Ca2+ stimulates binding of Mg2+-dependent PDHP to the complex; the bound enzyme was considered to be the active species. The fraction of PDH in the active form, rather than substrate and inhibitor levels, determines PDH activity under these conditions. This fraction depends on the present value and recent history of the difference between PDHK and PDHP activities. Both of these are active continuously and continuously control PDH.
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PMID:Computer simulation of metabolism in pyruvate-perfused rat heart. III. Pyruvate dehydrogenase. 47 88

One ribosomal protein kinase activity and 3 soluble protein kinase activities have been identified in plasma cell tumors by DEAE-cellulose chromatography. We have shown phosphorylation in vivo and in vitro of a protein fraction from the ribosomal KCl wash which we have termed 'PPx fraction'. Phosphorylation of this protein fraction has been obtained in vitro with the ribosome-associated protein kinase. We have determined for the ribosomal protein kinase the following characteristics. 1. It is an Mg2+-dependent enzyme that transfers the gamma-phosphate from ATP into phosphoseryl and phosphothreonyl residues of the substrate. 2. It has a wide substrate specificity. Like the soluble protein kinases it catalyses the phosphorylation of several proteins like histone, phosvitin, casein and ribosomal proteins but it differs from the main soluble kinases (I, II) by the fact that it catalyses specifically the phosphorylation at least of one of the ribosomal KCl wash proteins. On dodecylsulfate-polyacrylamide gels this protein has a molecular weight of approximately 90000 and it is released from ribosomes under conditions commonly employed for extraction of initiation factors. 3. The ribosome-associated protein kinase is not stimulated by the addition of cyclic adenosine 3':5'-monophosphate. 4. KCl has no effect, NaCl has a weak effect on the phosphorylation, Mn2+ and Ca2+ are inhibitors. 5. ADP has been found to be a competitive inhibitor. 6. The maximum velocity of the ribosomal protein-kinase-catalysed reaction is 0.65 nmol of 32P incorporated in the KCl wash protein per min and per mg protein. 7. The apparent Km for the ribosomal KCl wash protein as substrate is 0.71 mg/ml and the Km for ATP is 94 muM. 8. The molecular weight of the ribosomal protein kinase, estimated by electrophoresis in polyacrylamide-dodecylsulfate gels, is 60000 and corresponds probably to a catalytic subunit.
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PMID:A plasmocytoma ribosome-associated protein kinase which phosphorylates a specific protein of the ribosomal KCl wash. 124 81

Most platelet agonists activate and elevate the cytosolic free calcium concentration in human platelets through receptor-dependent mechanisms that are antagonized by cAMP- and cGMP-elevating agents. Nitrovasodilators such as nitroprusside and endothelium-derived relaxing factor are potent cGMP-elevating platelet inhibitors. In the present study, the role of cGMP and cGMP-dependent protein kinase in nitrovasodilator inhibition of ADP- and thrombin-evoked calcium elevation and activation of human platelets was investigated. Preincubation of platelets with 8-(4-chlorophenylthio)guanosine 3',5'-cyclic monophosphate (8-pCPT-cGMP; a membrane-permeant selective activator of the cGMP-dependent protein kinase that does not significantly affect cGMP-regulated phosphodiesterases) inhibited the thrombin-induced phosphorylation mediated by myosin light chain kinase and protein kinase C. Nitrovasodilator-induced protein phosphorylation in human platelets was distinct from that induced by cAMP-elevating prostaglandins and could be mimicked by 8-pCPT-cGMP. Preincubation of human platelets with nitrovasodilators or 8-pCPT-cGMP inhibited the ADP- and thrombin-evoked calcium elevation in the presence and absence of external calcium. Nitrovasodilators and 8-pCPT-cGMP also inhibited the agonist-induced Mn2+ influx, but stopped-flow experiments indicated that the ADP receptor-operated cation channel was not significantly inhibited. These results suggest that in human platelets nitrovasodilators inhibit the agonist-induced calcium mobilization from intracellular stores and the secondary store-related calcium influx but not the ADP receptor-operated cation channel. The results also suggest that these nitrovasodilator effects are mediated by cGMP and the cGMP-dependent protein kinase.
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PMID:Role of cGMP and cGMP-dependent protein kinase in nitrovasodilator inhibition of agonist-evoked calcium elevation in human platelets. 131 May 37

Murine melanoma cells treated with the melanocyte-stimulating hormone (MSH) family of peptides undergo differentiation characterized by enhanced melanogenesis and altered morphology. These effects are mediated via the adenylate cyclase-cAMP pathway leading to activation of protein kinase A (PKA). We have discovered that inhibition of a post-translational modification of chromatin proteins, viz. poly(ADP-ribosylation), also induces melanogenesis and differentiation in these cells. A range of competitive inhibitors (benzamide and its derivatives) of the nuclear enzyme poly(ADP-ribose) polymerase (PADPRP; EC 2.4.2.30) was utilized, and their ability to induce melanogenesis reflected their potency as PADPRP inhibitors. These compounds induced melanogenesis at low doses (20 microM-2 mM) which did not affect cell growth or viability. Induction of melanogenesis was not attributable to inhibition of cyclic nucleotide phosphodiesterase by these compounds. MSH treatment caused a transient rise in cAMP levels (up to 200-fold by 5 min and returning to near basal levels by 5 h). It also stimulated PKA activity up to 5-fold, and the temporal kinetics of this activation mirrored the changes in cAMP levels. In comparison, the PADPRP inhibitors had no effect on either of these processes. These data constitute a novel demonstration of a cAMP-independent mechanism for the induction of melanoma cell differentiation, including melanogenesis.
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PMID:Murine melanoma cell differentiation and melanogenesis induced by poly(ADP-ribose) polymerase inhibitors. 132 52

In contrast to other systems in which angiotensin-II (AII) inhibits adenylyl cyclase, in fetal skin fibroblasts the peptide stimulates cAMP accumulation. The mechanism of this novel effect was studied by analysis of the actions of AII and other regulators on the adenylyl cyclase system in cultured cells. In the presence of phosphodiesterase inhibitors, AII, isoproterenol (ISO), choleratoxin (CTx), and forskolin (Fk) stimulated cAMP accumulation by 2.0 +/- 0.26-, 26 +/- 0.9-, 75 +/- 5.6-, and 88 +/- 3.3-fold, respectively. AII potentiated the stimulatory effect of ISO and CTx by 1.5 +/- 0.1- and 1.25 +/- 0.03-fold, respectively, but had no effect on that of Fk. Preincubation of the cells with PTx did not prevent the stimulatory effect of AII on basal and ISO- and CTx-stimulated cAMP, indicating that the effect of AII was not due to interaction with Gi. Unexpectedly, pretreatment of the cells with PTx for 18 h inhibited cAMP production stimulated by ISO and Fk. Similar inhibition by PTx was observed in adult rat skin fibroblasts, but not in adult human fibroblasts, in which pretreatment with PTx resulted in potentiation of Fk-stimulated cAMP production. ADP ribosylation studies showed that the optical density of the band corresponding to Gs was less than 20% that of Gi and Go in rat fetal cells, suggesting that excess release of the beta-gamma-subunit is responsible for the inhibition of cAMP production by PTx. However, immunoblot analysis of G-proteins showed that the content of Gs alpha was similar to that of Gi alpha and Go alpha in rat and human, fetal and adult cells. In contrast to the effect in intact cells, AII had no effect on basal or stimulated adenylyl cyclase activity in cell homogenates, suggesting that the stimulatory effect observed in intact cells is indirect. The stimulatory action of AII on cAMP production was not blocked by indomethacin, indicating that the effect is not mediated by prostaglandin formation. The stimulation of cAMP by AII was mimicked by 10-min incubation with phorbol 12-myristate 13-acetate (PMA), and prevented after cellular protein kinase-C (PKC) depletion by 4- or 6-h preincubation with PMA. However, the stimulation was not prevented by the PKC inhibitors staurosporine and H7 or 24-h preincubation with PMA, suggesting that the effect is not mediated by a traditional PKC-dependent mechanism.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Studies on the mechanism of the novel stimulatory effect of angiotensin-II on adenylate cyclase in rat fetal skin fibroblasts. 133 May

Bovine adrenal medullary chromaffin cells maintained in tissue culture accumulated [3H]-noradrenaline by a high affinity, Na(+)-dependent, desipramine-sensitive process. The accumulation was linear with time (1-90 min) and had an apparent Km of 0.52 +/- 0.24 mumol/l and Vmax of 1.70 +/- 0.48 pmol/(10(5) cells.15 min). Pretreatment of the cells with the ADP-ribosylating agent pertussis toxin resulted in a reduction in the Vmax [0.81 +/- 0.39 pmol/(10(5)cells.15 min)] but no significant change in the apparent affinity (Km = 0.42 +/- 0.07 mumol/l). This inhibition of [3H]noradrenaline accumulation was distinct from that produced by the vesicular transport inhibitor reserpine. Pertussis toxin inhibition probably did not arise through an indirect action on the Na(+)-gradient because while, as expected, Na+,K(+)-ATPase inhibition reduced [3H]noradrenaline accumulation, pertussis toxin pretreatment always caused a further significant reduction even in the presence of maximally effective concentrations of ouabain. Stimulation of the cAMP-protein kinase A system by forskolin or 8-bromocyclic AMP also caused a reduction in [3H] noradrenaline accumulation but again pertussis toxin pretreatment always resulted in a further reduction. Thus, the data provide evidence for a pertussis toxin-sensitive element in the catecholamine accumulation process and are consistent with an action at a site directly associated with the transporter itself rather than with an indirect action via secondary processes.
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PMID:Pertussis toxin inhibits noradrenaline accumulation by bovine adrenal medullary chromaffin cells. 133 72

Exposure of adrenal Y-1 cells to C2 toxin results in an increase in steroid release that is accompanied by a rounding of the cell. The actions of C2 toxin mimic those of adrenocorticotropin and cholera toxin except that there is no increase in intracellular cyclic AMP content. In the present study we provide evidence that C2 toxin increases steroid output from Y-1 cells through an alteration in the microfilament network of the cell. C2 toxin significantly increased steroid output after 3 hr of exposure. This effect was accompanied by a significant increase in the transport of [3H]cholesterol to the mitochondrial fraction, independent of cholesterol uptake by the cell. The toxin was unable to increase steroid output from cells prerounded in suspension culture. The protease inhibitors benzamidine and phenylmethylsulfonyl fluoride did not attenuate the ability of C2 toxin to alter the morphology of Y-1 cells. A 3-hr exposure to C2 toxin resulted in the ADP-ribosylation of 50 to 60% of the total actin pool. Fluorescein isothiocyanate-labeled phalloidin visualization of the cytoskeleton of toxin-treated cells confirmed that the toxin caused a decrease in the stress fiber network. C2 toxin treatment of a protein kinase A mutant Y-1 cell (Kin 8) resulted in morphological changes and an increase in steroid output that was not different from that observed for wild type Y-1 cells. The data suggest that C2 toxin increases steroid output from adrenal Y-1 cells by a cyclic AMP-independent mechanism that involves the microfilament network of the cell.
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PMID:Botulinum C2 toxin and steroid production in adrenal Y-1 cells: the role of microfilaments in the toxin-induced increase in steroid release. 137 Nov 60


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