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 activity of myocardial adenosine kinase (E.N. 2.7.1.20) in a number of species was assayed. Rat heart contained the highest specific activity. From this source adenosine kinase was purified in a simple way 80-fold, until it was free of adenosine deaminase activity. A molecular weight of about 39 000 was measured. NSC 113939 (1), NSC 113940 and 8-azaadenosine inhibited myocardial adenosine kinase. Dipyridamole stimulated the enzyme at high adenosine levels, and inhibited at low substrate concentrations. A number of divalent cations could (partially) substitute for Mg2+. The optimal concentration of MgCl2 or MnCl2 was about 0.5 mM; concentrations exceeding 1 mM inhibited severely. An apparent Km for ATP of 0.1 mM was measured, whereas an apparent Km for adenosine of 0.5 muM was was found. The latter increased to 3.3 muM, when dipyridamole was added. Replacement of ATP by GTB or ITP increased the activity, and UTP and CTP were inferior as a phosphate donor.
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PMID:Partial purification and properties of rat-heart adenosine kinase. 7 32

The effect of increasing doses of GTP on agonist and antagonist binding to adenosine A1-receptors in different regions of rat brain was studied by autoradiography. A high concentration of GTP (100 microM) practically eliminated the binding of the agonist [3H]N6-cyclohexyladenosine in all regions. However, there were regional differences in the effects of low concentrations of GTP (0.1-10 microM). In some regions, for example the hippocampus, all concentrations of GTP decreased [3H]N6-cyclohexyladenosine binding, by decreasing the Bmax. In other structures, e.g. the superior colliculus, there was a biphasic response to GTP. Concentrations of 0.1-3 microM increased agonist binding, apparently due to a decrease in KD, whereas higher concentrations also decreased binding in these regions. The effects of GTP were mimicked by the stable GTP analogue guanosine-5'-O-(3-thiotriphosphate). GTP (0.5-100 microM) increased the binding of the antagonist [3H]8-cyclopentyl-1,3-dipropylxanthine in all regions, but most markedly in those where GTP had a biphasic effect on agonist binding. Decreasing the levels of endogenous adenosine by increasing the concentration of adenosine deaminase and adding the 5'-nucleotidase inhibitor alpha-beta-methylene adenosine-5'-diphosphate gave an increase in [3H]8-cyclopentyl-1,3-dipropylxanthine binding and diminished the response to GTP. In sections treated with adenosine deaminase and alpha-beta-methylene adenosine-5'-diphosphate, GTP steadily decreased [3H]N6-cyclohexyladenosine binding in all regions. Thus, the GTP-induced increase in both agonist and antagonist binding may be due to a displacement of endogenous adenosine. In the presence of 1 mM EDTA, GTP had a monophasic effect on the binding of [3H]N6-cyclohexyladenosine in all regions. In the presence of 2 mM MgCl2 a biphasic response to GTP was seen in all regions. In EDTA washed sections, the effect of MgCl2 on [3H]N6-cyclohexyladenosine binding was more pronounced in the superior colliculus, where we had observed a biphasic response to GTP. The results suggest that there are regional differences in the effects of GTP on adenosine A1-receptor binding in rat brain, that reflect regional differences in the magnesium-dependent binding of endogenous adenosine, which is bound to the receptor by tight binding, is very difficult to remove, and easily interferes with radioligand binding in in vitro experiments. There may be regional differences in the sensitivity of A1-receptor-G-protein complexes to magnesium, that reflect a heterogeneity of the G-proteins to which the A1-receptors are coupled.
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PMID:Regional differences in the effect of guanine nucleotides on agonist and antagonist binding to adenosine A1-receptors in rat brain, as revealed by autoradiography. 235 51

In the present study it is reported that [3H]NECA binds in a specific and saturable manner to membranes from the cerebral cortex of the rat. Scatchard analysis revealed two binding sites. The high affinity binding site (Kd 10.66 +/- 5 nM, Bmax 0.305 +/- 0.05 pmol/mg prot) was characterized by the following features: maximum binding at 25 degrees C, sensitivity to pretreatment with NEM and regulation by Gpp[NH]p, enhancing of binding in the presence of 1.0 mM MgCl2 and 1.5 mM CaCl2; the rank order of potency of several analogues of adenosine in competing with [3H]NECA for binding, was CHA greater than L-PIA greater than NECA greater than CADO. The low affinity binding site (Kd261.8 +/- 50 nM, Bmax 4.19 +/- 0.33 pmol/mg prot) showed maximum binding at 0 degrees C, insensitivity to pretreatment with NEM up to 1 mM and to regulation by Gpp[NH]p, and inhibition of binding in the presence of MgCl2 and CaCl2. The low affinity site was also present in membranes not pretreated with adenosine deaminase and, even in this condition, an IC50 of 188.5 +/- 36 nM for NECA and an IC50 of 4.35 +/- 0.20 microM for adenosine were found. It is concluded that the high affinity binding site is similar to the A1 adenosine receptors. The low affinity binding site is not classifiable among the A-type adenosine receptors, although it shows peculiar features shared both with the human platelet A2 receptor and the adenosine receptor formerly studied with [3H]adenosine in membranes from the brain of the rat; these results could reflect heterogeneity of adenosine receptors in central nervous system.
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PMID:5'-N-ethylcarboxamido[8-3H]adenosine binds to two different adenosine receptors in membranes from the cerebral cortex of the rat. 335 69

Examination of the binding characteristics of the adenosine agonist radioligands [3H]N6-cyclohexyladenosine [( 3H]CHA), [3H]cyclopentyladenosine [( 3H]CPA), and [3H]5'-N-ethylcarboxamido adenosine [( 3H]NECA) to membranes prepared from PC12 cells showed that the A-1-selective ligands (CHA and CPA) had minimal binding, which was not amenable to analysis using curve-fitting programs. However, [3H]NECA, a nonselective A-1/A-2 agonist, gave reproducible binding, which was enhanced by removal of endogenous adenosine, using the catabolic enzyme adenosine deaminase. This binding was of high affinity (KD = 4.7 nM) with limited capacity (263 fmol/mg of protein). Specific binding of [3H]NECA was unaffected by the presence of either CPA (50 nM) or MgCl2 (10 mM) but was sensitive to guanylylimidodiphosphate (100 microM), a finding suggesting involvement of an N-protein mechanism in the coupling of the adenosine receptor labeled by [3H]NECA to other components of the receptor complex. Binding of [3H]NECA to PC12 cell membranes was stereo-selective, with the R isomer of N6-phenylisopropyladenosine (PIA) being approximately 12 times more active than S-PIA. The A-1-selective agonist CPA was a weak inhibitor of [3H]NECA binding (Ki = 251 nM). The rank order of activity of adenosine agonists in displacing specific [3H]NECA binding was NECA greater than or equal to 2-chloroadenosine greater than CHA greater than or equal to 5'-N-methylcarboxamido adenosine greater than or equal to R-PIA greater than CPA greater than S-PIA. Binding was also displaced by the marine adenosine agonist 1-methylisoguanosine and by a series of xanthine antagonists with the activity order being 1,3-dipropyl-8-(2-amino-4-chloro)phenylxanthine greater than 8-phenyltheophylline greater than 8-p-sulfophenyltheophylline.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Characterization of adenosine receptors in the PC12 pheochromocytoma cell line using radioligand binding: evidence for A-2 selectivity. 379 18

The wide range of values reported for activity of adenylate cyclase (AC) in human skeletal muscle prompted re-evaluation of conditions used for homogenization and assay. Adenylate cyclase activity in the same normal muscle differed with different techniques of homogenization. In pH 7.5 isotonic Tris buffer, basal and catecholamine-activated activities declined rapidly in homogenates kept at 4 degrees C. Loss of basal activity was prevented by addition of a chelator of divalent cations. Loss of response to isoproterenol was prevented by addition of guanylnucleotides. Enzyme activity was maximal at 37 degrees C and pH 7.6. Enzyme activity was lower when theophylline was used to prevent degradation of labelled 3',5' cyclic adenosine monophosphate (cyclic AMP) than when unlabelled cyclic AMP was used to this purpose. Basal activity increased with increased MgCl2 concentration up to 50 mmol/l, but isoproterenol-activated activity was maximal at 4 mmol/l MgCl2. AC was inhibited by exogenous adenosine, but addition of adenosine deaminase to the assay mixture did not increase AC activity. Based upon these observations, standardized procedures of homogenization and assay were devised and used to measure AC activity in muscles of boys with Duchenne muscular dystrophy: basal and isoproterenol-stimulated activities were abnormally low.
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PMID:Assay of adenylate cyclase in homogenates of control and Duchenne human skeletal muscle. 722 46

Cell-free extracts of nitrate-grown Aspergillus terricola catalyze the hydrolytic deamination of adenosine to inosine at maximum rate at pH 6.5 and 50 degrees C. Incubation of the extracts at 60 degrees C for 30 minutes caused about 66.7% loss in activity. Results indicated the involvement of SH groups in the catalytic site of adenosine deaminase. Frequent freezing and thawing of the enzyme preparation for three days (3 times) resulted in about 47% loss in activity. The enzyme is also inhibited by EDTA indicating that adenosine deaminase is a metaloenzyme. MgCl2 and CoSO4 had a remarkable activating effect, whereas MnCl2 showed a slight inhibitory effect on enzyme activity. The apparent Km value was calculated for adenosine and found to be 6.66 x 10(-3) M, which indicates the greater affinity of adenosine deaminase for adenosine.
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PMID:Properties of adenosine deaminase in extracts of Asperigillus terricola. 774 Sep 80

The drug (2-amino-4,5-dimethyl-3-thienyl)-[3(trifluoromethyl)-phenyl]methanone (PD 81,723) has been shown to enhance allosterically A1 adenosine receptor binding in brain membranes. The objective of this study was to determine the specificity and selectivity (A1 versus A2) of PD 81,723 as an enhancer of the negative dromotropic effect of exogenous adenosine in guinea pig isolated and in situ hearts. In isolated hearts, PD 81,723 alone produced only a small stimulus to His bundle (S-H) interval prolongation of 1.5 to 4 msec, which was completely reversed by the A1 adenosine receptor antagonist 8-cyclopentyltheophylline and adenosine deaminase. PD 81,723 (5 microM) significantly decreased the EC50 value of adenosine for prolongation of the S-H interval from 6.7 +/- 0.6 to 4.4 +/- 0.5 microM. The potentiation of the negative dromotropic effect of adenosine by PD 81,723 was dose dependent, i.e., 5 and 10 microM PD 81,723 enhanced the maximal S-H interval prolongation caused by 3 microM adenosine by 207% and 609%, respectively. In contrast, the same concentration of PD 81,723 had no effect on either the S-H interval prolongation caused by carbachol or MgCl2 or the coronary vasodilatory effect of adenosine. In in situ hearts, PD (2 mumol/kg i.v.) alone caused only a small but not significant negative dromotropic effect, increasing the atrium to His interval from 58 +/- 2 to 61 +/- 1 msec. However, the same dose of PD 81,723 caused a significant leftward and upward shift of the adenosine dose-response curve for inducing atrium to His bundle interval prolongation and increased the degree of atrioventricular block caused by adenosine.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Selective potentiation by an A1 adenosine receptor enhancer of the negative dromotropic action of adenosine in the guinea pig heart. 835 94

Cell-free extracts of nitrate-grown Penicillium politans NRC-510 could catalyze the hydrolytic deamination of adenosine to inosine maximally at pH 6.0 and 45 degrees C. However the same extracts could not catalyze the N-glycosidic bond cleavage of adenosine at pH 4.0, 6.0 and 8.0. Incubation of the extracts at 55 degrees C for 30 minutes caused about 31% loss in activity whereas incubation of the extracts at 60 degrees C for 15 minutes caused a complete loss of enzyme activity. Results indicated the absence of the involvement of sulfhydryl groups in the catalytic site of adenosine deaminase. The enzyme is inhibited by ethylene diamine tetraacetate indicating that adenosine deaminase is a metalloenzyme. MnCl2 and MgCl2 had a remarkable activating effect, whereas HgCl2, CaCl2 and ZnSO4 showed an inhibitory effect on enzyme activity. Dialyzing the extracts for 24 hours significantly increase deaminase activity by about 33%. The apparent K(m) value was calculated for adenosine and found to be 3.63 x 10(-3) M, which indicates high affinity of adenosine deaminase for its substrate adenosine.
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PMID:Deamination of adenosine by extracts of Penicillium politans NRC-510. 1581 56

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