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 importance of intact adenosine deaminase (ADA) activity in the generation of superoxide anion by xanthine oxidase has been disputed in studies using human neutrophils or mouse macrophages. The latter demonstrated a positive correlation between ADA activity and superoxide production during phagocytosis. The immunodeficiency in inherited ADA deficiency was related to a defect in this process. Since there is considerable interspecies variation in the tissue distribution of xanthine oxidase, the metabolism of [8-14C]deoxyadenosine (dAR), the toxic metabolite which accumulates in inherited ADA deficiency, was investigated in human peritoneal macrophages. Evaluation of the distribution of radiolabel in both cell and medium demonstrated that human macrophages with intact ADA metabolize dAR under physiological conditions to deoxyinosine and hypoxanthine exclusively. The hypoxanthine is further metabolized within the cell to ATP and GTP, via IMP. No xanthine or uric acid could be detected, confirming that in human macrophages xanthine oxidase activity is insignificant, as it is in most other human cells and tissues, except liver and intestinal mucosa. Thus production of superoxide radicals in such cells via this route would be impossible, and consequently unaffected either by ADA deficiency or the xanthine oxidase inhibitor allopurinol.
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PMID:Superoxide radicals, immunodeficiency and xanthine oxidase activity: man is not a mouse! 298 25

Adenine nucleotides displace the binding of the selective adenosine A-1 receptor ligand [3H]cyclopentyladenosine (CPA) to rat brain membranes in a concentration-dependent manner, with the rank order of activity being ATP greater than ADP greater than AMP. Binding was also displaced by GTP, ITP, adenylylimidodiphosphate (AppNHp), 2-methylthioATP, and the beta-gamma-methylene isostere of ATP, but was unaffected by the alpha-beta-methylene isosteres of ADP and ATP, and UTP. At ATP concentrations greater than 100 microM, the inhibitory effects on CPA binding were reversed, until at 2 mM ATP, specific binding of CPA was identical to that seen in controls. Concentrations of ATP greater than 10 mM totally inhibited specific binding. Inclusion of the catabolic enzyme adenosine deaminase in the incubation medium abolished the inhibitory effects of ATP, indicating that these were due to adenosine formation, presumably due to ectonucleotidase activity. The inhibitory effects were also attenuated by the alpha-beta-methylene isostere of ATP, an ectonucleotidase inhibitor. Adenosine deaminase, alpha-beta-methylene ATP (100 microM), and beta-gamma-methylene ATP (100 microM) had no effect on the "stimulatory" phase of binding, although GTP (100 microM) slightly attenuated it. Comparison of the binding of [3H]CPA in the absence and presence of 2 mM ATP by saturation analysis showed that the KD and apparent Bmax values were identical. Examination of the pharmacology of the control and "ATP-dependent" CPA binding sites showed slight changes in binding of adenosine agonists and antagonists. The responses observed with high concentrations of ATP were not observed with GTP, AppNHp, the chelating agents EDTA and EGTA, or inorganic phosphate. The divalent cations Mg2+ and Ca2+ at 10 mM attenuated the stimulatory actions of high (2 mM) concentrations of ATP, whereas EGTA and EDTA (10 mM) enhanced the "stimulatory" actions of ATP. EDTA (10 mM) abolished the inhibitory effects of ATP, indicating a specific dependence on Mg2+ for the inhibitory response. The effects of ATP on [3H]CPA binding were reversible for antagonists but not agonists. The mechanism by which ATP reverses its own inhibitory action on adenosine A-1 radioligand binding is unclear, and from the observed actions of the divalent cations and chelating agents probably does not involve a phosphorylation-dependent process.
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PMID:Effects of purine nucleotides on the binding of [3H]cyclopentyladenosine to adenosine A-1 receptors in rat brain membranes. 308 5

Although many of the new cardiotonic agents are known to increase cAMP and to inhibit with variable potency a low Km cAMP phosphodiesterase, there is still debate as to the mechanism(s) by which these agents act. In a rat adipocyte membrane model we demonstrate that only approximately 50% of the effect of the new cardiotonic agent sulmazole on cAMP accumulation can be attributed to phosphodiesterase inhibition and that the remaining production of cAMP involves stimulation of adenylate cyclase activity. Two distinct pathways for stimulation of adenylate cyclase are herein reported. Sulmazole, UD-CG 212 CL, enoximone, piroximone, amrinone, and milrinone are all shown to be competitive antagonists of inhibitory A1 adenosine receptors, with EC50 values of 11-909 microM. Elimination of the effects of endogenous adenosine with adenosine deaminase reveals a third distinct mechanism for activation of adenylate cyclase. This mechanism appears to involve Gi, the inhibitory guanine nucleotide-regulatory protein, in that sulmazole attenuates the capacity of GTP to inhibit adenylate cyclase activity, and covalent modification of Gi by pertussis toxin treatment abolishes the capacity of sulmazole to mediate stimulation. Thus, functional blockade of Gi activity is the likely mode of action. Restoration of sulmazole's stimulatory effect on adenylate cyclase activity in pertussis toxin-treated membranes can be accomplished by reconstituting purified preparations of either Gi or mixtures of Gi/Go into treated adipocyte membranes. Of note, this stimulatory effect is completely reversed by inhibitory receptor agonists. Thus, the new cardiotonic agent sulmazole mediates increases in cAMP accumulation by mechanisms other than phosphodiesterase inhibition, including A1 adenosine receptor antagonism and inhibition of Gi function.
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PMID:The new cardiotonic agent sulmazole is an A1 adenosine receptor antagonist and functionally blocks the inhibitory regulator, Gi. 312 27

XAC, a high affinity antagonist of the A1 adenosine receptor, enhances adenylate cyclase activity by 1.3-2 fold with an EC50 of approximately 47 nM in adipocyte membranes pretreated with adenosine deaminase to eliminate adenosine and in the presence of total phosphodiesterase inhibition by 100 microM papaverine. This effect of XAC is observed only at concentrations of GTP sufficient to activate Gi (approximately 5 x 10(-6) M GTP) and is not evident in the absence or presence of lower GTP concentrations. ADP ribosylation of Gi by pertussis toxin treatment also abolishes this stimulatory action of XAC. Furthermore, in the presence of GTP activation of inhibitory prostaglandin E1 receptors diminishes the stimulatory effect of XAC on adenylate cyclase. In addition, XAC interferes with GTP-mediated inhibition of forskolin-stimulated adenylate cyclase activity in a noncompetitive manner. Finally, XAC is only a weak inhibitor of the low Km cyclic AMP phosphodiesterase, producing approximately 40% inhibition of phosphodiesterase activity at a concentration of 100 microM. These data suggest that XAC increases adenylate cyclase activity in absence of endogenous adenosine by inhibiting tonic Gi activity in a reversible manner.
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PMID:A novel site of action of a high affinity A1 adenosine receptor antagonist. 313 23

The mechanisms responsible for the large increases of intracellular ATP levels seen after isolated rabbit proximal tubules are treated with exogenous adenine nucleotides were studied. Exogenous ATP was rapidly degraded via adenosine as far as hypoxanthine. Degradation of AMP to adenosine was substantially inhibited by beta-glycerol phosphate. In studies of the ability of individual exogenous purines to increase intracellular ATP levels, single large doses of adenosine were less effective than equimolar doses of exogenous ATP but were substantially more effective than exogenous inosine or hypoxanthine. Exogenous guanine derived compounds increased only cell GTP. Incremental delivery of smaller doses of adenosine to maintain medium levels greater than 5 microM or inhibition of adenosine deaminase with erythro-9-[3-(2-hydroxynonyl)]adenine or 2'-deoxycoformicin enhanced the nucleoside's effectiveness. However, the initial increase of cell ATP was still greater after treatment with exogenous ATP than after adenosine and, in the presence of adenosine deaminase inhibition, larger increases of cell ATP were produced by 50 microM adenosine than by 250 microM adenosine. These observations are most consistent with substrate inhibition of adenosine kinase by adenosine. Furthermore, the adenosine kinase inhibitor, 5-iodotubercidin, prevented the increases of cell ATP resulting from exogenous adenosine or exogenous ATP. These studies demonstrate how the differential uptake and utilization characteristics of nucleosides and bases can fully account for the increases of intracellular nucleotides produced in isolated tubules by exogenous purines.
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PMID:Modulation of cell nucleotide levels of isolated kidney tubules. 334 10

This paper compares erythrocyte nucleotide levels in patients with eight different inherited purine or pyrimidine enzyme defects identified amongst a variety of patients referred predominantly for investigation of severe neurological abnormalities, or immunodeficiency syndromes. Characteristic nucleotide patterns were identified only in the six disorders (four involving purine and two pyrimidine metabolism) where there was clinical evidence of cellular toxicity. They were frequently related to the accumulation of abnormal metabolites in body fluids. These erythrocyte studies have demonstrated the following. 1. ATP depletion is not an invariable feature of adenosine deaminase (ADA) deficiency, but the accumulation of the deoxyribonucleotides dATP, or dGTP, is diagnostic of ADA, or purine nucleoside phosphorylase (PNP) deficiency, respectively. The early accumulation of dATP in foetal blood is a valuable aid to prenatal diagnosis of ADA deficiency. 2. GTP depletion appears to reflect the degree of CNS involvement in hypoxanthine-guanine phosphoribosyltransferase and PNP deficiency, as well as PP-ribose-P synthetase superactivity. Other diagnostic changes involving increased pyrimidine sugars and increased or decreased NAD levels, or ZTP in Lesch Nyhan erythrocytes, show no consistent correlation with the clinical manifestations. 3. These altered nucleotide levels afford a novel means for carrier detection of the X-linked defect associated with aberrant PP-ribose-P synthetase activity, where no other test is yet available. Measurement of erythrocyte nucleotide levels thus provides a simple and rapid aid to diagnosis and may sometimes be essential for determining prognosis, carrier detection, or monitoring therapy. These characteristic 'fingerprints' may give some insight into the mechanism by which the abnormal gene product produces disease. Such grossly altered nucleotide levels could also result in loss of erythrocyte flexibility, increased destruction and hence the anaemia, or other clinical manifestations, observed in some disorders.
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PMID:Altered erythrocyte nucleotide patterns are characteristic of inherited disorders of purine or pyrimidine metabolism. 337 Aug 20

Neplanocin A and aristeromycin are carbocyclic adenosine analogs that differ only in that neplanocin A contains a double bond in the carbocyclic ring, whereas this ring in aristeromycin is saturated. We have compared the metabolism and some of the metabolic effects of neplanocin A and synthetic (+/-)-aristeromycin (C-Ado) in murine leukemia L1210 cells in culture. C-Ado, as shown earlier, was not only converted to its own phosphates but also was metabolized to phosphates of carbocyclic guanosine. Both rapidly proliferating and slowly proliferating or resting cells phosphorylated C-Ado, but C-Ado was not converted to phosphates of carbocyclic guanosine in detectable amounts in cells whose growth had reached a plateau. When the metabolism of neplanocin and C-Ado was examined in the same experiment, both analogs were converted to the triphosphate analogs of ATP; no conversion of neplanocin A to the corresponding carbocyclic analogs of guanine nucleotides was detected, whereas C-Ado was converted to the carbocyclic analog of GTP in amounts that approximated the GTP pool. This difference in metabolism was associated with a marked difference in effects of the two analogs on the utilization of hypoxanthine and guanine which was inhibited by C-Ado but not by neplanocin. The failure of neplanocin A to be converted to analogs of guanine nucleotides apparently is the result of poor capacity of its monophosphate to serve as a substrate for AMP deaminase; the Vmax for deamination of neplanocin-5'-monophosphate by this enzyme was only 5% of that for C-Ado monophosphate. In contrast, neplanocin A was a better substrate than C-Ado for adenosine deaminase.
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PMID:Differences in the metabolism and metabolic effects of the carbocyclic adenosine analogs, neplanocin A and aristeromycin. 370 57

Three general questions regarding nucleosides and lymphocytes are discussed: (a) Why are so many measurements being made of adenosine deaminase activity, what do the results mean, and why is there still disagreement about some of the conclusions; (b) what do we understand about nucleosides and lymphocyte death; and (c) to what extent do we really understand nucleoside and nucleotide metabolism in lymphocytes? Experimental studies show that treatment of mice with deoxycoformycin, to produce accumulation of deoxyadenosine, leads to rapid thymus involution, elevated dATP concentrations in thymus and liver, and inhibition of adenosylhomocysteine hydrolase in these tissues. Deoxyguanosine inhibits the growth of mouse lymphoma L5178Y cells, and this toxicity is prevented by deoxycytidine plus adenine. In cells treated with deoxyguanosine, concentrations of both GTP and dGTP are elevated, and this is not affected by deoxycytidine. Adenine, however, reduces GTP concentrations to normal, and prevents most of the elevation in dGTP concentrations. Contrary to previous belief, it has been demonstrated that lymphocytes and nucleated bone marrow cells will synthesize purine nucleotides de novo if incubated in an appropriate medium; carbon dioxide is particularly important for this process.
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PMID:Regulation of purine metabolism in lymphocytes. 387 99

Inherited adenosine deaminase (ADA) deficiency is associated with a lymphospecific cytotoxicity affecting both dividing and non-dividing cells. The metabolic basis for this was investigated using different cell types and the potentially toxic metabolite 2'-deoxyadenosine (dAR) in short-term experiments under physiological conditions simulating ADA deficiency (1 mM Pi 8.7 microM dAR). In the uncultured cells, [8-14C] dAR alone was metabolized almost completely only by thymocytes and tonsil-derived B-lymphocytes. The greater percentage of counts (greater than 75%) were in the medium (deoxyinosine, hypoxanthine). Cellular counts were predominantly in adenine nucleotides, and to a lesser extent guanine nucleotides. Interestingly, both thymocytes and tonsil-derived B-lymphocytes, and a partially ADA deficient B lymphoblast line, accumulated detectable amounts of dATP even in the absence of ADA inhibition. Peripheral blood lymphocytes (PBMs) did not, and showed little dAR metabolism. In experiments simulating ADA deficiency varying amounts of 2'-deoxycoformycin (2'dCF) were needed to completely inhibit ADA (20-60 microM), with thymocytes requiring the highest amount. ADA inhibited thymocytes and tonsillar B-lymphocytes accumulated very high dATP levels, which were sustained to an equal extent by both over a 60-min period; PBMs accumulated the lowest values. Results in cultured cells reflected findings in previous studies. Some counts were also found in ATP by a route excluding ADA or PNP. These results again question the hypothesis that B-cells are more resistant than T-cells to the toxic effects of dAR because of an inability to accumulate and sustain elevated dATP levels and underline the lack of comparability between enzyme activity in intact as distinct from lysed cells. They cast doubt on the validity of cultured cells as a model for ADA deficiency and suggest the observed toxicity in some instances might result from altered ATP or GTP pools through inadequate ADA inhibition. They indicate that combined immunodeficiency in ADA deficiency could relate to an equal sensitivity of B-cells and T-cell precursors to the toxic effects of dATP accumulation.
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PMID:Human B lymphocytes and thymocytes but not peripheral blood mononuclear cells accumulate high dATP levels in conditions simulating ADA deficiency. 387 35

The action of endogeneous adenosine on isolated hamster brown adipocytes was examined. Adenosine production from brown adipocytes was measured after labeling of the intracellular nucleotide pool with [3H]adenine. Accumulation of [3H]adenosine in the incubation medium was maximum after 5 min of incubation and was still present after 20 min. When adenosine accumulation was prevented by addition of adenosine deaminase, the stimulatory effects of isoproterenol on oxygen uptake, lipolysis, and adenosine 3',5'-cyclic monophosphate (cAMP) generation were enhanced. However, basal rates of lipolysis and oxygen consumption and levels of cAMP were not affected on addition of adenosine deaminase. A similar potentiation of isoproterenol responses was produced by the adenosine receptor antagonist, 3-isobutyl-1-methylxanthine, present at a concentration (10 microM) which did not change basal levels of respiration or lipolysis. Addition of the adenosine analogue 2-chloroadenosine antagonized isoproterenol-stimulated respiration and lipolysis and prevented potentiation of isoproterenol responses with 3-isobutyl-1-methylxanthine. To localize the site of adenosine action, activity of adenylate cyclase in membrane preparations from brown adipocytes was measured. Isoproterenol-stimulated adenylate cyclase activity was partially inhibited by 2-chloroadenosine in a GTP-dependent manner. Addition of Na+ enhanced the inhibitory effect of 2-chloroadenosine, and 3-isobutyl-1-methylxanthine blocked it. The calculated 50% effective dose for 2-chloroadenosine inhibition was between 10 and 15 nM. These data suggest that adenosine produced by brown adipocytes is an endogenous regulator of respiration in these cells acting at the level of the adenylate cyclase enzyme.
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PMID:Role of adenosine as an endogenous regulator of respiration in hamster brown adipocytes. 619 83


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