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:3.1.3.5 (5'-nucleotidase)
3,167 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. The kinetic properties of the 5'-nucleotidase (EC 3.1.3.5) present in the cytosol of rat liver were investigated in relation to the conversion of adenine nucleotides into uric acid, with particular reference to the stimulation of this process by fructose. The enzyme was assayed by the release of Pi and by a new and more sensitive radiochemical procedure. 2. When IMP was used as substrate, the partially purified enzyme displayed almost hyperbolic kinetics (h = 1.1) with S0.5 = 1.2 mM. Similar kinetics were observed with GMP and other nucleoside 5'-monophosphates, except AMP. 3. Vmax. of the enzyme for AMP was about the same as for IMP, but the kinetics were sigmoidal (h = 1.6) with S 0.5 = 10 mM. 4. The hydrolysis of IMP was inhibited competitively by GMP. IMP, at concentrations up to 0.5 mM, had a paradoxical stimulatory action on the hydrolysis of 2-5 mM-AMP and was inhibitory at higher concentrations. 5. The activity of the enzyme towards AMP and IMP was stimulated by ATP and GTP, and inhibited by Pi. Activators and inhibitor approximately cancelled each others' effects. At pH 7.4, the enzymic activity with 0.2 mM-AMP was undetectable under physiological conditions. 6. It is concluded that, in the liver cell, AMP is not hydrolysed by the soluble 5'-nucleotidase, but that its degradation requires prior deamination to IMP.
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PMID:A kinetic study of the soluble 5'-nucleotidase of rat liver. 1 87

Hepatocytes and Kupffer cells were separated from rat liver after prelabeling the Kupffer cells with colloidal iron and perfusion of the liver with digestive enzymes. The activity of several enzymes from Kupffer cells and hepatocytes was compared to validate this method of cell separation. The ratios of hepatocyte to Kupffer cell specific activities of glucose-6-phosphatase, 5'-nucleotidase, adenylate cyclase, and acid phosphatase were 20, 0.39, 0.18, and 0.078, respectively. Adenylate cyclases from hepatocytes and Kupffer cells were stimulated by fluoride ion, GTP, and catecholamines. Hepatocyte adenylate cyclase was also stimulated by glucagon, secretin, vasoactive intestinal polypeptide, and by prostaglandin E1, whereas, the Kupffer cell enzyme was completely insensitive to these hormones. The stimulation of hepatocyte adenylate cyclase by combinations of glucagon plus secretin, or glucagon plus vasoactive intestinal polypeptide, were equivalent to the sum of the individual stimulations. This suggests that the hepatocyte has specific receptors for glucagon and for vasoactive intestinal polypeptide and secretin. Prostaglandin E1 stimulation of hepatocyte adenylate cyclase was not additive to the stimulation caused by polypeptide hormones or catecholamines, nor did prostaglandin E1 decrease stimulation caused by these hormones. Although prostaglandin-sensitive adenylate cyclase was recovered with hepatocytes, 40 to 50% of the total liver prostaglandin-sensitive activity was recovered in a fraction of cell debris mixed with small cells which did not phagocytize colloidal iron.
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PMID:Stimulation of adenylate cyclase from isolated hepatocytes and Kupffer cells. 17 Dec 69

Tod determine whether changes in unsaturation of fatty acids in rat liver plasma membranes might alter activities of membrane-associated enzymes, liver plasma membranes were prepared from rats fed purified diets lacking or supplemented with essential fatty acids. Two methods of membrane purification were used. A similar degree of purification was obtained with both methods for both depleted and control membranes, as indicated by marker enzyme purification. The proportion of essential fatty acids of the linoleate series was significantly lower in phospholipids from depleted rats. The specific activity of 5'-nucleotidase was lower, and the activity, V and apparent Km for total (Na+ +K+ +Mg2+)-ATPase were higher in the depleted liver plasma membranes. Arrhenius plots of total ATPase activity showed a discontinuity at the same temperature for both the depleted and control membranes. Activity with the depleted membranes was higher at all temperatures tested. Supplementation of deficient rats with a source of essential fatty acids (corn oil) restored V and apparent Km values to normal. Adenylate cyclase activity in the presence of fluoride, glucagon or glucagon plus GTP was significantly lower in the depleted plasma membranes.
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PMID:Liver plasma membranes from essential fatty acid-deficient rats. Isolation, fatty acid composition, and activities of 5'-nucleotidase, ATPase and adenylate cyclase. 17 79

A concomitant increase in the activity of LH-senstive adenylate cyclase and in the number of LH/hCG binding sites was induced in ovaries of immature rats upon administration of pregnant mare serum gonadotropin (PMSG), a hormone preparation known to have predominantly follicle stimulation (FSH-like) activity. When an optimal dose of PMSG (15 i.u./rat) was administered to 25-day-old rats, specific activity of LH-dependent adenylate cyclase and the number of binding sites for LH/hCG per mg protein remained unchanged during the first 24h, but 48h after injection a 2-to 4-fold increase in both parameters was observed. By contrast, there was no change in basal adenylate cyclase activity or in the response of the enzyme to the stimulatory action of guanosine-5'-(beta gamma-imino) triphosphate (Gpp (NH)p), GTP, or NaF. Specific activity of succinate cytochrome c reductase, glucose-6-phosphatase and 5'-nucleotidase were found to be unaffected by the hormonal pretreatment, although total protein determined in these homogenates increased 3-fold in the course of this treatment. It is inferred that during follicular maturation, FSH enhances the responsiveness of ovarian adenylate cyclase to LH by stimulating the insertion of LH/hCG-receptors into the cell membrane.
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PMID:Synchronous generation of ovarian hCG binding sites and LH-sensitive adenylate cyclase in immature rats following treatment with pregnant mare serum gonadotropin. 19 42

The subcellular localization of guanylate cyclase was examined in rat liver. About 80% of the enzyme activity of homogenates was found in the soluble fraction. Particulate guanylate cyclase was localized in plasma membranes and microsomes. Crude nuclear and microsomal fractions were applied to discontinuous sucrose gradients, and the resulting fractions were examined for guanylate cyclase, various enzyme markers of cell components, and electron microscopy. Purified plasma membrane fractions obtained from either preparation had the highest specific activity of guanylate cyclase, 30 to 80 pmol/min/mg of protein, and the recovery and relative specific activity of guanylate cyclase paralleled that of 5'-nucleotidase and adenylate cyclase in these fractions. Significant amounts of guanylate cyclase, adenylate cyclase, 5'-nucleotidase, and glucose-6-phosphatase were recovered in purified preparation of microsomes. We cannot exclude the presence of guanylate cyclase in other cell components such as Golgi. The electron microscopic studies of fractions supported the biochemical studies with enzyme markers. Soluble guanylate cyclase had typical Michaelis-Menten kinetics with respect to GTP and had an apparent Km for GTP of 35 muM. Ca-2+ stimulated the soluble activity in the presence of low concentrations of Mn-2+. The properties of guanylate cyclase in plasma membranes and microsomes were similar except that Ca-2+ inhibited the activity associated with plasma membranes and had no effect on that of microsomes. Both particulate enzymes were allosteric in nature; double reciprocal plots of velocity versus GTP were not linear, and Hill coefficients for preparations of plasma membranes and microsomes were calculated to be 1.60 and 1.58, respectively. The soluble and particulate enzymes were inhibited by ATP, and inhibition of the soluble enzyme was slightly greater. While Mg-2+ was less effective than Mn-2+ as a sole cation, all enzyme fractions were markedly stimulated with Mg-2+ in the presence of a low concentration of Mn-2+. Triton X-100 increased the activity of particulate fractions about 3- to 10-fold and increased the soluble activity 50 to 100%.
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PMID:Localization of particulate guanylate cyclase in plasma membranes and microsomes of rat liver. 23 12

Insulin treatment of isolated liver plasma membranes induced the release of 5'-nucleotidase and alkaline phosphatase. This effect was maximal at physiological hormone concentrations, being 36% and 17% for 5'-nucleotidase and alkaline phosphatase respectively, and was fully mimicked by the phosphatidylinositol specific phospholipase C (PI-PLC), thus confirming the presence of a glycosylphosphatidylinositol anchoring-system for these exofacial enzymatic proteins. The complete inhibition of insulin dependent enzyme release by neomycin is strongly supportive of an involvement of membrane-located PI-PLC activity. In addition, the insulin-like effect on enzyme release induced by the GTP non-hydrolysable analog, GTP-gamma-S, and its sensitivity to the pertussis toxin are in favour of a mediatory role exerted by the G proteins system, in the transduction of some actions of insulin.
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PMID:Insulin-dependent release of 5'-nucleotidase and alkaline phosphatase from liver plasma membranes. 133 52

Effects of adenosine and nucleotides on the release of previously stored [3H]-noradrenaline were studied in rabbit brain cortex slices. The slices were stimulated twice, in most experiments by 6 electrical field pulses delivered at 100 Hz. Adenosine and the nucleotides AMP, ADP, ATP, AMPS, ADP beta S, ATP gamma S, beta,gamma-imido-ATP and beta,gamma-methylene-ATP all reduced the evoked overflow of tritiated compounds. For purines for which concentration-response curves were determined, the order of potency was adenosine greater than ATP approximately ATP gamma S approximately beta,gamma-imido-ATP approximately ADP greater than beta,gamma-methylene-ATP. AMP 30 mumol/l and AMPS 30 mumol/l were approximately equieffective with 30 mumol/l of adenosine and ATP gamma S, and ADP beta S 30 mumol/l was approximately equieffective with 30 mumol/l of ADP. alpha,beta-Methylene-ADP, 2-methylthio-ATP, UTP and GTP gamma S did not change the evoked overflow of tritium. alpha,beta-Methylene-ATP caused an increase; however, the increase was small and became significant only after 59 min of exposure to alpha,beta-methylene-ATP or when the slices were stimulated by 30 pulses, 10 Hz. Neither adenosine deaminase (100 U/l) nor the blocker of 5'-nucleotidase, alpha,beta-methylene-ADP (10 mumol/l), attenuated the inhibition caused by ATP, ATP gamma S and beta,gamma-methylene-ATP, despite the fact that adenosine deaminase abolished the effect of adenosine. 8-Cyclopentyl-1,3-dipropylxanthine (DPCPX, 10 nmol/l) shifted the concentration-response curves of adenosine, ATP gamma S, beta,gamma-imido-ATP and beta,gamma-methylene-ATP to the right by very similar degrees. 8-(p-Sulphophenyl)-theophylline (30 and 300 mumol/l) also markedly antagonized the inhibition produced by ATP gamma S. alpha,beta-Methylene-ATP (10 and 30 mumol/l) and suramin (100 mumol/l) did not modify the effects of adenosine, ATP gamma S and beta,gamma-methylene-ATP. It is concluded that nucleotides themselves can inhibit the release of noradrenaline in the rabbit brain cortex. The nucleotides and adenosine seem to act at the same site, i.e., the A1 subtype of the P1-purinoceptor. The results support the notion that metabolically stable, phosphate chain-modified nucleotides such as ATP gamma S, beta,gamma-imido-ATP and beta,gamma-methylene-ATP can be potent P1 agonists. No evidence was found for presynaptic P2x-, P2y- or P3-purinoceptors.
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PMID:Stable adenine nucleotides inhibit [3H]-noradrenaline release in rabbit brain cortex slices by direct action at presynaptic adenosine A1-receptors. 144 82

A GTP-binding protein with an apparent molecular weight of 25 kDa was detected in hepatocyte extracts using SDS-PAGE and [alpha-32P]GTP. p21ras proteins could only be detected by immunological analysis. The amounts of p21ras proteins present in isolated hepatocytes and in a highly purified preparation of liver plasma membrane vesicles were 0.3 and 4 ng p21ras protein/micrograms membrane protein, respectively. In comparison with the total cell extract, the degree of enrichment of plasma membrane vesicles with p21ras was similar to that of 5'-nucleotidase. The p21ras proteins were tightly associated with the membrane. Treatment of [3H]choline-labelled plasma membranes with an excess concentration of the anti-p21ras antibody Y13-259 failed to inhibit either basal or guanosine 5'-[gamma-thio]triphosphate (GTP[S])-stimulated [3H]choline release. It is concluded that in hepatocytes (a) the majority of p21ras is bound to the plasma membrane and (b) p21ras is not directly involved in the activation by GTP[S] of phospholipase D.
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PMID:Low molecular weight GTP-binding proteins in hepatocytes and an assessment of the role of p21ras proteins in the activation of phospholipase D. 177 29

Ribavirin enhances the anti-human immunodeficiency virus activity of 2',3'-dideoxyinosine (ddIno) in MT-4, CEM and peripheral blood lymphocyte cells. Ribavirin causes an increase in the levels of IMP, the presumed phosphate donor for the conversion of ddIno to ddIMP by 5'-nucleotidase. Consequently, ribavirin stimulates the conversion of ddIno to its antivirally active metabolite ddATP. Ribavirin also causes a marked depletion of the guanine nucleotide pools. The increase in IMP pool levels may result from (i) a direct inhibitory effect of ribavirin 5'-monophosphate on IMP dehydrogenase (which converts IMP to XMP) and (ii) an indirect inhibition of adenylosuccinate synthetase by the decreased GTP and dGTP pools (since GTP is an obligatory cofactor in the conversion of IMP to succinyl AMP). GTP depletion plays a key role in the accumulation of IMP and the resultant higher rate of ddIno phosphorylation to ddIMP and eventually ddATP. Our findings are in agreement with the observations that guanosine and 2'-deoxyguanosine, but not 2'-deoxyadenosine, reverse (i) the stimulatory effect of ribavirin on the anti-human immunodeficiency virus activity of ddIno and (ii) the accumulation of endogenous IMP pools as well as accumulation of [3H]IMP from exogenous [3H]hypoxanthine in ribavirin-treated cells.
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PMID:Mechanism of the potentiating effect of ribavirin on the activity of 2',3'-dideoxyinosine against human immunodeficiency virus. 193 81

Carbovir (CBV) is a highly selective carbocyclic nucleoside inhibitor of HIV replication in human lymphocytes and is potentially useful in the treatment of AIDS [Vince et al. (1988) Biochem. Biophys. Res. Commun. 156, 1046-1053]. Using human lymphoid cells severely deficient in nucleoside kinases, we were able to identify the route of activation of CBV metabolism. The present studies have demonstrated that CBV is anabolized to the mono-, di-, and triphosphates and to guanosine 5'-triphosphate in CCRF-CEM cells. Conversion to GTP amounted to 15-20% of the total analogue nucleotides formed in the cells and may arise from CBV through depurination and salvage via HGPRT. Evidence was obtained that neither deoxycytidine kinase, adenosine kinase, or mitochondrial deoxyguanosine kinase is primarily involved in the initial step of phosphorylation of CBV in CCRF-CEM cells. In contrast, earlier studies [Johnson & Fridland (1989) Mol. Pharmacol. 36, 291-295] showed that a cytosolic 5'-nucleotidase catalyzes the activation of CBV to the monosphosphate. Other biochemical effects examined showed that the nucleobases hypoxanthine and adenine, but not guanine, their respective nucleosides, and the dideoxynucleosides 2',3'-dideoxyinosine, 2',3'-dideoxyguanosine, and 3'-azido-3'-deoxythymidine produced significant increased accumulation of CBV nucleotides in CEM cells. The exact mechanism for this potentiation of CBV phosphorylation has not been elucidated but may be due to a modulating effect of intracellular nucleotides on 5'-nucleotidase activity.
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PMID:Metabolism of the carbocyclic nucleoside analogue carbovir, an inhibitor of human immunodeficiency virus, in human lymphoid cells. 227 22


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