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)

To examine whether activation of polymorphonuclear leukocytes attenuates release of adenosine through attenuation of their own ecto-5'-nucleotidase activity, human polymorphonuclear leukocytes were incubated with and without exposure to either N-formyl-methionyl-leucyl-phenylalanine (FMLP) or complement C5a. Ecto-5'-nucleotidase activity of polymorphonuclear leukocytes was attenuated by both FMLP and complement C5a (22.7 +/- 3.6 vs 9.7 +/- 2.6 nmol/min per 10(7) cells at 10(-6) M FMLP, P < .05; 21.5 +/- 2.2 vs 10.2 +/- 1.2 nmol/min per 10(7) cells at 5 x 10(-7) g/mL complement C5a, P < .001), whereas cytosolic 5'-nucleotidase activity was not affected by either FMLP or complement C5a. These reductions of ecto-5'-nucleotidase activity that were caused by both FMLP and complement C5a were dose and time dependent and were inhibited by superoxide dismutase. Desferrioxamine did not inhibit the decreases in ecto-5'-nucleotidase. In accordance with the decreases in ecto-5'-nucleotidase activity, release of adenosine was attenuated in the FMLP-pretreated and complement C5a-pretreated polymorphonuclear leukocytes, which were restored by concomitant administration of superoxide dismutase. The viability of FMLP-pretreated and complement C5a-pretreated polymorphonuclear leukocytes was markedly decreased compared with the untreated group after 60 minutes of hypoxia followed by 60 minutes of reoxygenation. Thus, we conclude that: (1) activation of polymorphonuclear leukocytes attenuates their own ecto-5'-nucleotidase activity and thereby reduces adenosine release, (2) reduction of ecto-5'-nucleotidase activity is attributable to generated superoxide anion in polymorphonuclear leukocytes, and (3) viability of polymorphonuclear leukocytes after hypoxia and reoxygenation largely depends on the extents of decreases in ecto-5'-nucleotidase activity.
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PMID:Attenuation of ecto-5'-nucleotidase activity and adenosine release in activated human polymorphonuclear leukocytes. 834 95

The myoblast cell surface activity of ecto-5'-nucleotidase was stimulated by a laminin substrate, whereas fibronectin and gelatin did not increase the AMPase activity of ecto-5'-nucleotidase. This increase was related to a higher expression of ecto-5'-nucleotidase on the surface of cells seeded on a laminin substrate, but without the mobilization of an intracellular pool of enzyme. Furthermore, laminin and its fragments E'1 and E8 modified the AMPase activity of the ecto-5'-nucleotidase purified from chicken striated muscle and reconstituted in liposomes. Over the range of concentrations used, intact laminin and its fragment E8, consisting of the distal half of the long arm, stimulated the AMPase activity of ecto-5'-nucleotidase. By contrast, the large fragment derived from the short arms, designated E'1, inhibited the AMPase activity. Furthermore, the monoclonal anti-ecto-5'-nucleotidase antibody, CG37, abolished the stimulatory effect of fragment E8 on the AMPase activity of ecto-5'-nucleotidase but did not reverse the inhibitory effect of fragment E'1. In conclusion, laminin stimulates the AMPase activity of ecto-5'-nucleotidase by two mechanisms: inducing the expression of ecto-5'-nucleotidase to the cell surface and direct modulation of the enzymatic activity.
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PMID:Dual mechanism of laminin modulation of ecto-5'-nucleotidase activity. 839 52

The components of the ectonucleotidase pathway at the immunoaffinity-purified striatal cholinergic synapse have been studied. The ecto-ATPase (EC 3.6.1.15) had a Km of 131 microM, whereas the ecto-ADPase (EC 3.6.1.6) had a Km of 58 microM, was Ca(2+)-dependent, and was inhibited by the ATP analogue 5'-adenylylimidodiphosphate (AMPPNP). The ecto-5'-nucleotidase (EC 3.1.3.5) had a Km of 21 microM, was inhibited by AMPPNP and alpha,beta-methylene ADP, and by a specific antiserum. The Vmax values of the ATPase, ADPase, and 5'-nucleotidase enzymes present at this synapse were in a ratio of 30:14:1. Very little ecto-adenylate kinase activity was detected on these purified synapses. The intraterminal 5'-nucleotidase enzyme, which amounted to 40% of the total 5'-nucleotidase activity, was inhibited by AMPPNP, alpha,beta-methylene ADP, and the antiserum, and also had the same kinetic properties as the ectoenzyme. The time course of ATP degradation to adenosine outside the nerve terminals showed a delay, followed by a period of sustained adenosine production. The delay in adenosine production was proportional to the initial ATP concentration, was a consequence of feedforward inhibition of the ADPase and 5'-nucleotidase, and was inversely proportional to the ecto-5'-nucleotidase activity. The function and characteristics of this pathway and the central role of 5'-nucleotidase in the regulation of extraterminal adenosine concentrations are discussed.
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PMID:Production of adenosine from extracellular ATP at the striatal cholinergic synapse. 841 43

1. Studies in rat polymorphonuclear leucocytes have suggested that 5'-deoxy-5'-isobutylthioadenosine (IBTA), an inhibitor of the IMP-selective cytosolic 5'-nucleotidase, may be used to test its role in adenosine formation in intact cells. We investigated adenosine formation in neonatal and adult rat cardiomyocytes. 2. 2-Deoxyglucose (30 mM) with oligomycin (2 micrograms/ml) induced a 90-100% fall in ATP concentration in 10 min in neonatal and 60 min in adult heart cells. Adenosine accumulation was substantially increased, accounting for 13% of the fall in ATP concentration in neonatal cells and 56% in adult cells. 3. Anti-(rat liver ecto-5'-nucleotidase) serum did not inhibit adenosine accumulation. Furthermore, dipyridamole (10 microM), a nucleoside-transport blocker, inhibited by 80% the appearance of the newly formed adenosine in the medium, showing that adenosine is produced intracellularly by both adult and neonatal-rat myocytes in response to inhibition of oxidative metabolism. 4. IBTA (3 mM) inhibited by 80% the appearance of adenosine in the medium, but did not inhibit total adenosine accumulation by neonatal-rat myocytes and only modestly inhibited total adenosine accumulation by adult myocytes. 5. IBTA, like dipyridamole, inhibited incorporation of extracellular adenosine (10 microM) into neonatal and adult ventricular myocyte nucleotides by 60-70%. Transport of IBTA (100 microM) into the cells did not appear to be inhibited by dipyridamole (30 microM). 6. We conclude that IBTA acted primarily to inhibit adenosine release from myocytes. The small effect on adenosine formation rates implies that the IMP-selective cytosolic 5'-nucleotidase plays a minor role in this tissue.
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PMID:Effect of 5'-deoxy-5'-isobutylthioadenosine on formation and release of adenosine from neonatal and adult rat ventricular myocytes. 848 9

This study was aimed to determine whether singlet oxygen (1O2) attenuates 5'-nucleotidase activity in the ischemic myocardium. Isolated rat hearts were exposed to either exogenous 1O2 produced by irradiating rose bengal or 40-min ischemia and reperfusion. Ecto-5'-nucleotidase activity was inhibited by exogenous 1O2 (3.74 +/- 0.38 mumol/min/g dry weight), when compared with normal control (7.52 +/- 0.41 mumol/min/g dry weight; P < 0.05). The enzymatic activity was significantly preserved by histidine (25 mM)--a 1O2 scavenger (7.04 +/- 0.61 mumol/min/g dry weight; P < 0.05 v rose bengal group). After ischemia, the activity of ecto-5'-nucleotidase was greatly reduced (2.51 +/- 0.25 mumol/min/g dry weight), when compared with normal control. Histidine significantly enhanced ecto-5'-nucleotidase activity (6.55 +/- 0.52 mumol/min/g dry weight, P < 0.05 v ischemic control). Adenosine release was consistent with ecto-5'-nucleotidase activity. The time course studies of effects of 1O2 on coronary flow, cardiac function, and LDH release revealed that the damage by 1O2 to ecto-5'-nucleotidase activity and adenosine release primarily accounted for impaired coronary flow, cardiac dysfunction, and impaired cardiac metabolism. Lipid peroxidation induced by exogenous 1O2 or ischemia was in parallel with ecto-5'-nucleotidase deactivation by 1O2. It is concluded that 1O2 causes inactivation of ecto-5'-nucleotidase and attenuation of adenosine release which could possibly be one of the important mechanisms of oxygen radical-mediated myocardial injury.
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PMID:Interaction of singlet oxygen with 5'-nucleotidase in rat hearts. 859 96

Ochratoxin A is a mycotoxin produced by Aspergillus ochraceus and is a natural contaminant of moldly food. Ochratoxin A has a number of toxic effects, some of which may be related to the changes in the cell membrane. We measured the activities of 5 pancreatic, membrane bound enzymes in female Fisher rats that were given low oral doses of ochratoxin A (120 micrograms/kg body weight per day) during 20-35 days. The amount of toxin corresponds to 1.5 mg/kg in the feed, daily. These doses are in the range of natural contamination found in feed. The enzymes studied were alanine aminopeptidase, alkaline phosphatase, ecto-Ca2+/Mg(2+)-ATPase, gamma-glutamyl transferase and ecto-5'-nucleotidase. Treatment lasting 20 days caused a strong decrease in the activity of alanine aminopeptidase, Ca2+/Mg(2+)-ATPase and alkaline phosphatase to 0.76 +/- 0.04, 0.53 +/- 0.03 and 0.30 +/- 0.02 of the control values, respectively (p < 0.05). No significant changes in the activity of gamma-glutamyl transferase and 5'-nucleotidase were observed. However, activity of alanine aminopeptidase returned to normal values after 35 days of treatment, suggesting an adaptation of the organism, or a substitution of a released enzyme. Activities of alkaline phosphatase and Ca2+/Mg(2+)-ATPase remained significantly reduced to 0.42 +/- 0.03 and 0.52 +/- 0.04, respectively (p < 0.01). We conclude that treatment of rats with low doses of ochratoxin A resulted in reduction of the activities of the membrane bound enzymes, most probably by inducing their release, as a result of the impairment of the functional integrity of cell membranes.
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PMID:Ochratoxin A impairs activity of the membrane bound enzymes in rat pancreas. 860 90

Various 5'-nucleotidases (EC 3.1.3.5) exist in vertebrate tissues. The sequence and cDNA cloning of the membrane-bound ecto-5'-nucleotidase (e-N) and one of the cytosolic isoenzymes, IMP-preferring (c-N-II), but not the cytosolic AMP-preferring form (c-N-I), have been reported. While c-N-II has a broad tissue distribution, c-N-I is found only in vertebrate heart. The published data on substrate specificity involve mainly the naturally occurring nucleoside monophosphates, without a systematic structure-activity relationship study. In the present study we have used a series of AMP and IMP analogues to examine the structure-activity relationship for c-N-I and c-N-II in detail. The rank order of activity of the test compounds differed substantially between c-N-I and c-N-II. c-N-I and c-N-II varied with respect to the following interactions with substrate: (1) hydrogen-bond formation with the substituent in the 6-position of the purine ring (a donor-type with c-N-I and an acceptor-type with c-N-II); and (2) hydrophobic attraction of the 6-position unsubstituted purine ring (more pronounced with c-N-I than with c-N-II). No better substrate than 5'-AMP was found for c-N-I. We propose that c-N-I functions as an AMP-binding protein in the myocardial cell with an important role during ischaemic ATP breakdown when AMP accumulates rapidly.
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PMID:Structure-activity relationship of cytoplasmic 5'-nucleotidase substrate sites. 861 51

The distribution of ecto-5'-nucleotidase, a glycosyl phosphatidylinositol anchored membrane protein capable of hydrolysing extracellular nucleoside monophosphates, was investigated by immunocytochemistry in cultures of rat cerebellar cells obtained at postnatal days 6 and 8. The enzyme was expressed at the surface of granule cells including their neurites as well as on other neurons in the culture. The distribution of 5'-nucleotidase matched that of the synaptic vesicle protein 2. Oligodendroglial cells were identified by their immunoreactivity for 2',3'-cyclic nucleotide 3'-phosphodiesterase. Their entire surface was labelled for 5'-nucleotidase. In contrast, only a subset of astrocytes immunopositive for the glial fibrillary acidic protein revealed surface-located immunoreactivity for 5'-nucleotidase. Antibody-binding of the labelled-astrocytes was enhanced at restricted surface domains. Endothelial cells that avidly bind Lycopersicon esculentum lectin were the most intensely anti-5'-nucleotidase-labelled cell type of the culture. Double labelling revealed an exact match of surface-located antibody binding sites for 5'-nucleotidase and laminin. Immunoreactivity for 5'-nucleotidase was essentially absent from fibroblasts that could be identified by their immunoreactivity for fibronectin. All cell types that carried surface-bound 5'-nucleotidase also revealed a cytoplasmic pool of the enzyme. Our results provide the first immunocytochemical demonstration of the surface-location of 5'-nucleotidase in neurons. They suggest that the broad distribution of the enzyme at the surface of neurons and other cells types from neonatal brain reflects its functional importance in the differentiation of the nervous system.
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PMID:Immunocytochemical localization of ecto-5'-nucleotidase in cultures of cerebellar granule cells. 884 51

The purpose of this study was to evaluate the relative contributions of AMP-specific cytosolic 5'-nucleotidase and ecto-5'-nucleotidase to cardiac adenosine production and its regulation by ADP and Mg2+. 5'-Nucleotidase activity was measured spectrophotometrically in the total homogenate, the 150,000-g supernatant fraction (cytosolic 5'-nucleotidase), and the membrane pellet fraction (ecto-5'-nucleotidase) of dog left ventricles. Increasing [MgCl2] over a range from 0 to 6 mmol/l increased 5'-nucleotidase activity in both the supernatant and pellet; only cytosolic 5'-nucleotidase exhibited an absolute requirement for Mg2+. ADP, (20-480 mumol/l) activated supernatant and inhibited membrane-bound 5'-nucleotidase activity. At 80 mumol/l ADP, 5 mmol/l MgCl2, 100 mumol/l AMP, and pH 7.3, the average 5'-nucleotidase activities of the supernatant vs. pellet were 74% of total and 26% of total, respectively. Total adenosine production in unfractionated samples of ventricular homogenates decreased an average of 73% by specific inhibition of cytosolic 5'-nucleotidase, using antibodies against the cytosolic enzyme, and 46% by specific inhibition of ecto-5'-nucleotidase with alpha, beta-methylene adenosine 5'-diphosphate (AOPCP). These findings support the hypotheses that 1) both cytosolic and ecto-5'-nucleotidase contribute to cardiac adenosine production in dog heart homogenates; 2) AMP-specific cytosolic 5'-nucleotidase activity exceeds ecto-5'-nucleotidase activity at physiological concentrations of ADP, AMP, and Mg2+; and 3) Mg2+ is an important regulator of cardiac adenosine production via activation of both ecto- and AMP-specific cytosolic 5'-nucleotidases.
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PMID:Contribution of cytosolic and membrane-bound 5'-nucleotidases to cardiac adenosine production. 894 37

At the neuromuscular junction and possibly also at the synaptic level in the brain, the main sequence of events (see Fig. 5) that involves purines in modulation of ACh release includes the following observations: (1) storage of ATP and its release either together with, or independently of acetylcholine. ATP is also released from the post-junctional component. Adenosine as such is released either from the motor nerve terminals or from the post-junctional component. (2) There is extracellular hydrolysis of ATP to adenosine, which is the active substance to modulate transmitter release. The key enzyme in the conversion of AMP into adenosine is the ecto 5'-nucleotidase. When ecto-5'-nucleotidase is not available (e.g. in cholinergic nerve terminals of the cerebral cortex) ATP as such exerts the neuromodulatory role normally fulfilled by adenosine. (3) Both the inhibition and the excitation induced by adenosine on ACh release in the rat is inactivated through up-take and deamination. (4) Adenosine-induced inhibition of ACh release is mediated via A1 receptors and the excitation via A2a receptors. The A2a receptors are positively coupled to the adenylate cyclase/cyclic AMP system, whereas the presynaptic A1 receptors (a) may be negatively linked to adenylate cyclase and (b) to phospholipase C, and, upon stimulation, (c) increase potassium conductance and (d) decrease calcium conductance. (5) Activation of A2a receptors is essential for substances that facilitate ACh release (e.g. CGRP, forskolin) to exert their effects, as well as for induction of nicotinic autofacilitatory receptor desensitization. (6) There are interactions between A1 and A2a receptors. Thus, the net adenosine neuromodulatory response is the resultant, at each moment, of the relative degree of activation of each one of these receptors. This relative activation depends upon the intensity (frequency, pulse duration) of stimulation of the motor nerve terminals. (7) Adenosine released as such seems to preferentially activate A1 receptors, whereas the adenosine formed from metabolism of adenine nucleotides prefers to activate the A2a receptors. In conclusion, to find out precisely what occurs with ACh in transmitting its message at the synaptic level, one has to consider the subtle ways used by purines to modulate the ACh response. It therefore appears of interest that pharmacological and therapeutic strategies use this knowledge to approach cholinergic transmission deficiencies based upon reduction of ACh release.
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PMID:Purinergic regulation of acetylcholine release. 900 12


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