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:3.5.4.4 (adenosine deaminase)
5,136 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The mechanism of inhibition of neutrophil phagocytic functions by cAMP-elevating agents has not yet been clarified. In the present work, the effects of adenylate cyclase agonists on protein phosphorylation in the formylmethionyl-leucyl-phenylalanine (fMLP)-stimulated human neutrophils were studied. Before stimulation, 32Pi-labelled cells were incubated with adenosine deaminase to remove the endogenously produced adenosine, an adenylate cyclase agonist itself. A protein of about 52,000 molecular weight was rapidly and transiently phosphorylated when neutrophils were stimulated with fMLP in the presence of isoproterenol, prostaglandin E1, histamine or 2-chloroadenosine. This phosphorylation was blocked by the antagonists of the receptors for the above-listed agents. No phosphorylation of the 52,000 molecular weight protein could be observed if either fMLP or the cAMP-elevating agent were applied alone. A calcium ionophore A23187 and dibutyryl-cAMP could replace fMLP and a cAMP-elevating agent, respectively. Phosphorylation of the 52,000 molecular weight protein was also demonstrated in cell lysates in the presence of cAMP, and in membrane preparations in the presence of the catalytic subunit of cAMP-dependent protein kinase. These data suggest that phosphorylation of the 52,000 molecular weight protein in intact cells is dependent on the cross-talk between the fMLP- and the cAMP-signalling pathways, and may thus be involved in the cAMP-regulatory mechanism.
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PMID:Cross-talk between cAMP and formylmet-leu-phe in human neutrophils: phosphorylation of a 52,000 molecular weight protein. 132 1

With the use of -cAMP/+cAMP activity ratios of cAMP-dependent protein kinase (A-kinase) in fat cell extracts as an index of cellular cAMP concentrations, it is apparent from both the current literature and from data presented in this paper that classical cell isolation procedures yield cells whose behavior is unpredictable from day to day. Herein, procedures are described for isolating adipocytes, preparing cytosolic extracts, and assaying A-kinase that result in kinase activity ratios in isolated cells equal to those in the fat pad from which cells are derived, approximately 0.05. An important modification in the procedure is the inclusion of 200 nM exogenous Ado in all cell manipulation media, and the data indicate that variable removal of contaminating endogenous Ado accounts for unpredictable results with standard cell isolation techniques. A further benefit of Ado inclusion is greatly reduced cell lysis. Acute removal of Ado with adenosine deaminase results in rapid elevation of A-kinase activity ratios and lipolysis which, in fasted animals, equals that achieved with lipolytic hormones. Cells from fed animals exhibit poor predictability in behavior. Moreover, A-kinase activity ratios exhibit seasonal tendencies in response to Ado removal, with cells isolated in spring being more activated than cells isolated later in the year. The information and procedures in this paper form the basis for succeeding papers on the regulation of adipocyte metabolism by hormones.
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PMID:cAMP-dependent protein kinase and lipolysis in rat adipocytes. I. Cell preparation, manipulation, and predictability in behavior. 241 13

This paper examines the modulation of insulin-stimulated glucose transport activity in rat adipose cells by ligands for receptors (R) that mediate stimulation (Rs; lipolytic) or inhibition (Ri; antilipolytic) of adenylate cyclase. The changes in glucose transport activity and cAMP, as assessed by 3-O-methylglucose uptake and (-/+) cAMP-dependent protein kinase (A-kinase) activity ratios, respectively, were monitored under conditions that maintain steady-state A-kinase activity ratios (Honnor, R. C., Dhillon, G. S., and Londos, C. (1985) J. Biol. Chem. 260, 15122-15129). Removal of endogenous adenosine with adenosine deaminase decreased insulin-stimulated glucose transport activity by approximately 30%, which was prevented or restored with Ri agonists such as phenylisopropyladenosine, nicotinic acid, and prostaglandin E1. These changes in transport activity were not accompanied by changes in A-kinase activity ratios, indicating that Ri-mediated effects on transport are independent of cAMP changes. Addition of an Rs ligand, isoproterenol, in the presence of adenosine increased kinase activity but did not change glucose transport activity. Conversely, upon removal of adenosine, addition of Rs ligands such as isoproterenol, adrenocorticotropic hormone, or glucagon strongly inhibited transport (approximately 50%) and stimulated kinase activity. However, subsequent addition of phenylisopropyladenosine nearly restored transport activity without alteration of A-kinase activity. These data and additional kinetic experiments suggest that Rs-mediated glucose transport modulations are also independent of cAMP. The interchangeability of ligands for both Rs and Ri receptors in modulating transport activity suggests that these cAMP-independent effects are mediated by the stimulatory (Ns) and inhibitory (Ni) guanyl nucleotide-binding regulatory proteins of adenylate cyclase. All Rs-and Ri-induced changes in transport activity occurred without a change in glucose transporter distribution, as assessed by D-glucose-inhibitable cytochalasin B binding, suggesting that Rs and Ri ligands modulate the intrinsic activity of the glucose transporter present in the plasma membrane.
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PMID:Regulation of insulin-stimulated glucose transport in the isolated rat adipocyte. cAMP-independent effects of lipolytic and antilipolytic agents. 302 4

The steady-state relationship between the activation state of cAMP-dependent protein kinase (A-kinase) and lipolysis has been defined quantitatively. A-kinase activation was assessed by measuring the ( +/- cAMP) activity ratio in adipocyte extracts, and lipolysis was determined by measuring glycerol release from cells. Both processes were stimulated either by incubating cells in a ligand-free environment achieved with adenosine deaminase or by addition of lipolytic hormones. A response spectrum was obtained with a variety of adenylate cyclase stimulators and inhibitors, both receptor- and nonreceptor-mediated. Regardless of the ligands used to manipulate adipocyte activity, lipolysis varied from nil to maximal as the A-kinase activity ratio varied from approximately 0.05 to 0.3-0.35. These data provide a quantitative description of the steady-state relationship between A-kinase activity and lipolysis and indicate that the various lipolytic and antilipolytic agents tested act on the lipolytic process exclusively by altering adenylate cyclase activity and, thus, cellular cAMP concentrations. The data reveal also that transient "peaking" of cAMP, as measured by A-kinase activity ratios, is not an inherent feature of adipocyte metabolism. Moreover, the concentration requirements for lipolytic hormone action are critically dependent on the ambient concentration of antilipolytic agents, and t concentration requirements for antilipolytic agents are dependent on the extent to which cells are stimulated. The data in this paper provide the basis for assessing the relationship between A-kinase activity ratio and lipolysis in the presence of insulin (Londos, C., Honnor, R. C., and Dhillon, G. S. (1985) J. Biol. Chem. 260, 15139-15145).
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PMID:cAMP-dependent protein kinase and lipolysis in rat adipocytes. II. Definition of steady-state relationship with lipolytic and antilipolytic modulators. 387 23

The relationship between cAMP-dependent protein kinase (A-kinase) activity ratios and lipolysis in the presence of insulin was compared to the standard relationship between these two parameters established with a variety of adenylate cyclase modulators (Honnor, R. C., Dhillon, G., and Londos, C. (1985) J. Biol. Chem. 260, 15130-15138). Three phases of insulin action were observed. First, when tested in control cells exhibiting A-kinase activity ratios up to approximately 0.25, insulin inhibition of lipolysis could be accounted for by the decrease in A-kinase activity. Second, in cells exhibiting A-kinase activity ratios greater than 0.3, the decrease in kinase activity by insulin did not account for the decrease in lipolysis. Finally, as the A-kinase activity ratio approached 0.6 the insulin effect on lipolysis was lost. The data suggest that protein phosphatase activation accounts for the cAMP-independent insulin action. Moreover, the insulin effect not accounted for by a decrease in A-kinase activity appears to be elicited only upon elevation of A-kinase activity. The method by which cells were stimulated determined the IC50 for insulin inhibition of: 1) A-kinase activity ratios, 2) lipolysis explained by the decrease in A-kinase activity ratios, and 3) lipolysis not explained by a decrease in A-kinase activity ratios. For all three parameters, cells stimulated by lipolytic hormones were approximately 5 times more sensitive to insulin than cells stimulated by incubation in a ligand-free environment achieved with adenosine deaminase; insulin IC50 values were approximately 120 and 600 pM, respectively. Such data establish a link between insulin actions in modifying cAMP concentrations and in modifying events apparently independent of changes in cAMP. It is proposed that the receptors and regulatory components associated with adipocyte adenylate cyclase are associated also with components of the insulin response system separate from cyclase.
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PMID:cAMP-dependent protein kinase and lipolysis in rat adipocytes. III. Multiple modes of insulin regulation of lipolysis and regulation of insulin responses by adenylate cyclase regulators. 390 91

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

Purified striatal synaptosomes were continuously superfused with L,3,5[3H]tyrosine in order to estimate the synthesis ([3H]water) and release of newly formed [3H]dopamine. In the presence of magnesium, L-glutamate, D,L-alpha-amino-3-hydroxy-5-methyl-4-isoxazole-4-propionate (AMPA) and kainate, but not N-methyl-D-aspartate (NMDA) and 1-aminocyclopentane-1S,3R-dicarboxylate (t-ACPD), stimulated the release of [3H]dopamine, in a dose-dependent manner. When magnesium was omitted or in the presence of AMPA, NMDA also increased the release of [3H]dopamine. The effects of AMPA and kainate were competitively inhibited by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) or 6,7-dinitro-quinoxaline-2,3-dione (DNQX), whereas those of NMDA were reduced by 2-amino-5-phosphonovalerate (APV) or (+)-5-methyl-10,11-dihydro-5-H-dibenzo(a,d)cyclo-hepten-5,10-imine maleate (MK801). The stimulation of [3H]dopamine release by a high concentration of glutamate resulted from the concomitant activation of AMPA and NMDA receptors since this effect was potentiated by glycine and reduced by 2-amino-5-phosphonovalerate or MK801. This reduction was almost complete in the combined presence of DNQX and MK801. Surprisingly, glutamate and NMDA (in the absence of magnesium) reduced the efflux of [3H]water. The reduction of [3H]dopamine synthesis was blocked by 2-amino-5-phosphonovalerate indicating the involvement of NMDA receptors. Neither AMPA nor kainate affected dopamine synthesis. The inhibition of [3H]dopamine synthesis resulting from the stimulation of NMDA receptors was prevented when synaptosomes were continuously superfused with adenosine deaminase and quinpirole, a combined treatment known to markedly reduce the phosphorylation of tyrosine hydroxylase by cAMP-dependent protein kinase. The opposite effects of a high concentration of glutamate on [3H]dopamine synthesis and release were mimicked by ionomycin. As a working hypothesis, it is proposed that the NMDA-triggered calcium influx could lead to a reduction of tyrosine hydroxylase phosphorylation, possibly through an activation of calcineurin.
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PMID:Presynaptic control of dopamine synthesis and release by excitatory amino acids in rat striatal synaptosomes. 799 95

Acetylcholine acting via muscarinic cholinoceptors decreased phosphorylation of phospholamban and troponin I without reducing adenosine 3',5'-cyclic monophosphate (cAMP) levels or cAMP-dependent protein kinase activity ratio in the presence of 10-100 nM isoproterenol in guinea pig ventricular myocytes. The effect of acetylcholine was more pronounced when adenosine deaminase (5 U/ml) was present and incubation period was short (10 s). Okadaic acid, an inhibitor of protein phosphatase activity, blocked the acetylcholine-mediated inhibition of isoproterenol-stimulated phosphorylation of phospholamban. It is suggested that acetylcholine reduces protein phosphorylation by a cAMP-independent mechanism in guinea pig ventricular myocytes.
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PMID:M2-specific muscarinic cholinergic receptor-mediated inhibition of cardiac regulatory protein phosphorylation. 816 Aug 16

The effects of theophylline upon human alveolar macrophage function were assessed and compared with its action upon macrophage cyclic nucleotide phosphodiesterase (PDE) activity and cyclic adenosine monophosphate (cAMP) levels. In the concentration range of 10 mumol/liter to 1 mmol/liter, theophylline caused a concentration-dependent inhibition of opsonized zymosan-stimulated hydrogen peroxide (H2O2) generation and PDE-catalyzed cAMP hydrolysis and increased the cellular cAMP content. Macrophage H2O2 generation was also inhibited by forskolin, an activator of adenylyl cyclase, but whereas theophylline (1 mmol/liter) and forskolin (1 mumol/liter) exhibited a synergic elevation of macrophage cAMP, there was no synergy between the two agents in the inhibition of respiratory burst. The inhibition of H2O2 generation by theophylline was reversed by the competitive inhibitor of cAMP-dependent protein kinase, (Rp)8-bromoadenosine cyclic 3':5'-monophosphorothioate (Rp-8-Br-cAMPS; 100 mumol/liter), indicating that the functional effect of theophylline was mediated through the elevation of cAMP. The inhibition of H2O2 generation by theophylline was not affected by adenosine deaminase (0.1 U/ml), indicating that the inhibition did not involve adenosine antagonism. It is concluded that theophylline exerts a direct inhibitory action upon human alveolar macrophage function through the elevation of cAMP levels as a result of PDE inhibition, and that this effect is observed at concentrations of theophylline that may be achieved in serum during therapy.
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PMID:Theophylline suppresses human alveolar macrophage respiratory burst through phosphodiesterase inhibition. 817 21

The 8-chloro analogue of the regulatory molecule, cyclic AMP (cAMP), modulates the intracellular concentrations of cAMP-dependent protein kinases (PKA) and inhibits both in vitro and in vivo growth of several neoplastic cell types. Because 8-chloro-cyclic AMP (8-Cl-cAMP) can be converted to 8-chloroadenosine (8-Cl-adenosine) by serum enzymes contained in cell growth media, we tested whether 8-Cl-cAMP effects were mediated by its adenosine metabolite in normal and neoplastic cell lines of mouse lung epithelial origin. 8-Cl-adenosine, directly added to cells or derived from exogenously applied 8-Cl-cAMP, specifically decreased the intracellular concentration of the type I isozyme of cAMP-dependent protein kinase (PKA I). 8-Cl-adenosine and 8-Cl-cAMP were equipotent at inhibiting cell growth, and elicited similar changes in the proportion of cells in the G1, S, and G2-M phases of the cell cycle. The presence of adenosine deaminase, which converts 8-Cl-adenosine to 8-chloroinosine, completely prevented growth inhibition by 8-Cl-cAMP and the concomitant diminution of PKA I. 8-Cl-cAMP had no discernible effect on cells when its conversion into 8-Cl-adenosine was prevented by 3-isobutyl-1-methyl-xanthene, an inhibitor of phosphodiesterase. 6-(p-Nitrobenzyl)-thioinosine, an inhibitor of adenosine uptake, protected cells from cytostasis, indicating that 8-Cl-adenosine acts intracellularly. 8-Cl-adenosine greatly decreased RI (regulatory subunit of PKA I) and PKA catalytic (C) subunit protein concentrations without affecting RII (regulatory subunit of the PKA type II isozyme) or intracellular cAMP levels. Northern blot analysis of PKA subunit mRNAs following treatment of each cell line with 8-Cl-adenosine demonstrated decreased C alpha mRNA expression, increased RII alpha mRNA, and no change in RI alpha mRNA abundance. Our results indicate that 8-Cl-adenosine inhibits lung cell growth and induces PKA I down-regulation via a cAMP-independent mechanism.
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PMID:8-Chloroadenosine mediates 8-chloro-cyclic AMP-induced down-regulation of cyclic AMP-dependent protein kinase in normal and neoplastic mouse lung epithelial cells by a cyclic AMP-independent mechanism. 838 Feb 55


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