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)

Although the intracellular fatty acid binding proteins have been investigated for nearly two decades and purified proteins are now available, little is known regarding the function of these proteins in intact cells. Therefore, L-cell fibroblasts transfected with cDNA encoding for rat liver fatty acid binding protein (L-FABP) were examined as to whether L-FABP expression in intact cells modifies plasma membrane enzyme activities, fluidity, and lipids. Plasma membrane Na/K-ATPase activity was 65.9 +/- 18.7 and 38.6 +/- 22.8 (P less than 0.001) nmol/mg protein x min for control and high-expression transfected cells, respectively. Consistent with this observation, [3H] ouabain binding to whole cells was significantly decreased from 3.7 +/- 0.3 to 2.0 +/- 0.8 pmol ouabain bound/mg cell protein in control and high-expression cells, respectively, whereas the cell's affinity for ouabain was not significantly altered. Unexpectedly, Western blot analysis indicated that transfected cells had higher levels of Na+, K(+)-ATPase protein; in contrast, the activities of 5'-nucleotidase and Mg-ATPase were unaltered. The effects of L-FABP expression on plasma membrane Na/K-ATPase function appeared to be mediated through alterations in plasma membrane lipids and/or structure. The plasma membrane cholesterol/phospholipid ratio decreased and the bulk plasma membrane fluidity increased in the high-expression cells. In conclusion, plasma membrane Na/K-ATPase activity in L cells may be regulated in part through expression of cytosolic L-FABP.
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PMID:Na pump and plasma membrane structure in L-cell fibroblasts expressing rat liver fatty acid binding protein. 132 53

Cholesterol oxidation products (oxysterols), such as cholestan-3 beta,5 alpha,6 beta-triol (Triol), may be atherogenic by altering the barrier function of the vascular endothelium. We have shown that incubation of endothelial cell monolayers with Triol increased transendothelial albumin transfer (i.e., decreased barrier function) in a concentration- and time-dependent manner. Such dysfunction of endothelium could result from alterations in membrane characteristics, including changes in membrane-associated enzyme activities. To test this hypothesis, endothelial monolayers were treated with 20 microM Triol and the activities of selected membrane enzymes were measured at 0, 2, 4, 6, 12 and 24 hours. Calcium-adenosine triphosphatase (Ca(++)-ATPase) and sodium, potassium, magnesium-adenosine triphosphatase (Na+, K+, Mg(++)-ATPase) activities were significantly increased after 4 or 2 hours incubation with 20 microM Triol, respectively. 5'-nucleotidase activity was significantly elevated only after a 24-hour exposure to Triol, whereas there was no change in angiotensin-converting enzyme (ACE) activity in response to 20 microM Triol treatment at any time studied. Compared with all concentrations tested 40 microM Triol increased Ca(++)-ATPase activity most markedly, with a significant increase already after a 2-hour exposure. No major morphological changes were noted until 12 hours of exposure to 20 microM Triol; obvious cellular damage was observed by 24 hours. Cultures treated with Triol for 24 hours showed significant signs of toxicity, measured by an elevated [3H]adenine release, compared with control cultures. These data demonstrate that Triol alters the activity of certain membrane-bound enzymes, particularly Na+, K+, Mg(++)-ATPase and Ca(++)-ATPase.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Oxysterol-induced endothelial cell dysfunction in culture. 133 99

Homogenates and plasma membranes were isolated from the livers of male Fischer 344 rats ranging in age from 19 hr to 92 days postnatal. These plasma membranes exhibited comparable levels of purity: protein yields were 2-2.5%; relative specific activities of 5'-nucleotidase and ouabain-sensitive Na+/K(+)-ATPase were from 8-11 and from 12-19, respectively. 5'-nucleotidase and ouabain-sensitive Na+ K(+)-ATPase displayed distinct and different developmental patterns. The activity of gamma-glutamyltranspeptidase was found to be at exceptionally high levels in isolated plasma membranes immediately after birth and to decline precipitously thereafter achieving and maintaining low levels from days 3-21 postnatal. Liver plasma membrane gamma-glutamyltranspeptidase activity was observed to increase 9.2 fold from this low point, first rising on day 21, peaking on day 40 and returning to low levels by day 56. From day 56 day to 92 postnatal, gamma-glutamyltranspeptidase activity was expressed at a uniformly low level but a level 2 fold higher than that preceding the rise at day 40. The hormone determinants of these developmental changes in gamma-glutamyltranspeptidase activity are discussed.
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PMID:An extended developmental study of gamma-glutamyltranspeptidase in rat liver plasma membranes: identification of specific patterns of changes in activity in the adult as well as the neonatal state. 135

The arrival of the nerve impulse to the nerve endings leads to a series of events involving the entry of sodium and the exit of potassium. Restoration of ionic equilibria of sodium and potassium through the membrane is carried out by the sodium/potassium pump, that is the enzyme Na+,K(+)-ATPase. This is a particle-bound enzyme that concentrates in the nerve ending or synaptosomal membranes. The activity of Na+,K(+)-ATPase is essential for the maintenance of numerous reactions, as demonstrated in the isolated synaptosomes. This lends interest to the knowledge of the possible regulatory mechanisms of Na+,K(+)-ATPase activity in the synaptic region. The aim of this review is to summarize the results obtained in the author's laboratory, that refer to the effect of neurotransmitters and endogenous substances on Na+,K(+)-ATPase activity. Mention is also made of results in the field obtained in other laboratories. Evidence showing that brain Na+,K(+)-ATPase activity may be modified by certain neurotransmitters and insulin have been presented. The type of change produced by noradrenaline, dopamine, and serotonin on synaptosomal membrane Na+,K(+)-ATPase was found to depend on the presence or absence of a soluble brain fraction. The soluble brain fraction itself was able to stimulate or inhibit the enzyme, an effect that was dependent in turn on the time elapsed between preparation and use of the fraction. The filtration of soluble brain fraction through Sephadex G-50 allowed the separation of two active subfractions: peaks I and II. Peak I increased Na+,K(+)- and Mg(2+)-ATPases, and peak II inhibited Na+,K(+)-ATPase. Other membrane enzymes such as acetylcholinesterase and 5'-nucleotidase were unchanged by peaks I or II. In normotensive anesthetized rats, water and sodium excretion were not modified by peak I but were increased by peak II, thus resembling ouabain effects. 3H-ouabain binding was unchanged by peak I but decreased by peak II in some areas of the CNS assayed by quantitative autoradiography and in synaptosomal membranes assayed by a filtration technique. The effects of peak I and II on Na+,K(+)-ATPase were reversed by catecholamines. The extent of Na+,K(+)-ATPase inhibition by peak II was dependent on K+ concentration, thus suggesting an interference with the K+ site of the enzyme. Peak II was able to induce the release of neurotransmitter stored in the synaptic vesicles in a way similar to ouabain. Taking into account that peak II inhibits only Na+,N(+)-ATPase, increases diuresis and natriuresis, blocks high affinity 3H-ouabain binding, and induces neurotransmitter release, it is suggested that it contains an ouabain-like substance.
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PMID:In search of synaptosomal Na+,K(+)-ATPase regulators. 136 48

The effect of various dietary fats on membrane lipid composition, fatty acid profiles and membrane-bound enzyme activities of rat cardiac sarcolemma was assessed. Four groups of male weanling Charles Foster Young rats were fed diets containing 20% of groundnut, coconut, safflower or mustard oil for 16 weeks. Cardiac sarcolemma was prepared from each group and the activities of Na+, K(+)-ATPase, 5'-nucleotidase, Ca(2+)-ATPase and acetylcholinesterase were examined. ATPase activities were similar in all groups except the one fed coconut oil, which had the highest activities. Acetylcholinesterase activity was also similar in all the groups, however, it was significantly higher in the group fed mustard oil. No significant changes were observed among the groups in 5'-nucleotidase activity, in the cholesterol-to-phospholipid molar ratio and in sialic acid content. The coconut, safflower and mustard oil diets significantly increased cholesterol and phospholipid contents and the lipid-to-protein ratio of cardiac sarcolemma as compared to feeding the groundnut oil diet. The fatty acid composition of membrane lipids was quite different among the various groups, reflecting the type of dietary fat given. The total unsaturated-to-saturated fatty acid ratio was not different among the various groups; however, the levels of some major fatty acids such as palmitic (16:0), oleic (18:1) and linoleic (18:2) acids were significantly different. Cardiac sarcolemma of the group fed safflower oil had the highest polyunsaturated fatty acid content. The results suggest that dietary fats induce changes not only in the fatty acid composition of the component lipids but also in the activities of sarcolemmal enzymes involved in the regulation of cardiac function.
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PMID:Effect of dietary fats on some membrane-bound enzyme activities, membrane lipid composition and fatty acid profiles of rat heart sarcolemma. 140 62

1. A model is presented for adenosine transport and metabolism in different steady states. The model considers steady-state equations for metabolic enzymes based on information from the literature on their kinetic behaviour. 2. Assuming that extracellular adenosine and inosine are translocated by three transporters, we have devised rate equations for these nucleoside transporters which are valid when both nucleosides are present. Since the Na(+)-independent transporter can either incorporate nucleosides into the cell or release them, various conditions have been simulated in which inosine was either incorporated or released. 3. Control analyses are reported which show that the fluxes towards intracellular adenine nucleosides are controlled by ecto-5'-nucleotidase in some circumstances and by the nucleoside transporters in others. The nucleoside transporter is responsible for five fluxes (two Na+ dependent adenosine transport mechanisms, a Na(+)-dependent inosine transport, a Na(+)-independent adenosine transport and a Na(+)-independent inosine influx or efflux) but the control is not always positive for all these fluxes. The control patterns of these five fluxes indicate that, in the presence of extracellular adenosine and inosine, the intracellular metabolism of adenine derivatives would be highly dependent on the extracellular and intracellular concentrations of both nucleosides, on the ectoenzymes (5'-nucleotidase and adenosine deaminase) and on the transporter. 4. Predictions of the model were examined. The results indicate that a change in one independent variable (extracellular AMP concentration) makes the system evolve towards a new steady state which is far from the initial one and has a different control pattern. In contrast, simulation of inhibition of the carriers produces only slight modification of the fluxes since the concentrations of the metabolites change to counteract the effect. Thus, for instance, a 50% inhibition of the three carriers does not affect the flux towards intracellular adenine nucleotides. Finally, our model has confirmed that the evolution of the concentration of extracellular adenosine, when an increase in extracellular AMP is produced, agrees with the behaviour expected for a neurohormone.
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PMID:A model for adenosine transport and metabolism. 144 4

5'-Nucleotidase has been purified from rat glioblastoma cells (Rugli cells). The enzyme has been solubilized from plasma membranes by using Triton X-100 and CHAPS. Two affinity chromatographies on concanavalin A and 5'-AMP-Sepharose render the purified enzyme with a high specific activity (76.36 mumol AMP.min-1.mg-1). The purified enzyme gives a single polypeptide band on SDS-PAGE with an apparent molecular mass of 74 kDa. Active forms with an apparent molecular mass of 135 kDa and 268 kDa are observed when the purified enzyme is analyzed by gel filtration in the presence of either 0.6% sodium deoxycholate or 0.1% Triton X-100, respectively. The purified 5'-nucleotidase presents optimum activity at pH 7.8-8.1 either in the presence or in the absence of Mg2+. A linear Arrhenius plot is observed in the 25-46 degrees C temperature range and an activation energy of 33.7 KJ/mol is calculated. The enzyme is inhibited by EDTA; the activity is partially restored by different divalent cations as Zn2+, Mn2+, and Co2+. The hydrolysis of nucleosides 5'-monophosphate shows Michaelis kinetic. The enzyme is inhibited by nucleosides di- and triphosphate. 5'-Nucleotidase is a glycoprotein, being its activity inhibited at different extent by various lectins.
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PMID:Isolation and characterization of the ecto-5'-nucleotidase from a rat glioblastoma cell line. 148 Jan 62

A preliminary examination for the purification and characterization of 5'-nucleotidase of fish muscle was carried out and the following results were obtained. 1. The activities of 5'-nucleotidase in the muscles of marine vertebrates and invertebrates (total 11 species) were determined. The highest activity of 5'-nucleotidase was found in Blackrock fish Sebastes inermis, which was then used as a material for estimation of subcellular distribution and solubilization of the enzyme. 2. The 5'-nucleotidase of ordinary muscle of the fish Sebastes inermis was found in nuclear, microsomal and cytosolic fractions. About half of the total activity was found in the nuclear fraction, whereas the highest specific activity was observed in the microsomal fraction. 3. Complete solubilization of the enzyme was attained by using a high concentration of detergent such as Triton X-100, CHAPS, octylglucoside, octylthioglucoside and sodium deoxycholate, suggesting that the enzyme was tightly bound to the membrane. 4. Based on the results of solubility and stability tests, Triton X-100 seemed suitable for solubilizing 5'-nucleotidase from the membrane. 5. Microsomal 5'-nucleotidase was an Mg(2+)-activated enzyme, and no inactivation was observed up to 50 mM of Mg2+.
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PMID:Distribution of 5'-nucleotidase in muscle of some marine fishes. 161 35

This ultrastructural study was undertaken to determine the localization of cytochemically demonstrable blood-brain barrier (BBB)-associated enzymatic activities and of some nonenzymatic constituents in goat [corrected] brain microvascular endothelial cells (ECs) growing in vitro. Positive reactions for alkaline phosphatase (AP), 5'-nucleotidase (5'N), transport ATPase (Na+,K(+)-ATPase), and adenosine diphosphatase (ADPase) were present on both apical and basolateral plasma membranes (PMs) of the ECs. The reaction for calcium-dependent ATPase (Ca(2+)-ATPase) was less intense and was restricted to basolateral PM and associated plasmalemmal pits. These cells also revealed an abundance of anionic sites labeled with cationic colloidal gold (CCG) and Ricinus communis agglutinin 120 (RCA)-binding sites, specific for beta-D-galactosyl residues, on the apical PM. The labeling of the apical PM with Ulex europaeus agglutinin (UEA)-gold complex, specific for alpha-L-fucosyl residues, was negligible. When compared with results of cytochemical examination of the ECs of goat [corrected] brain capillary in vivo, these observations indicate that although cells cultivated in vitro retain at confluence the enzymatic activities typical for BBB-type ECS, they lose their characteristic (polar) localization. This loss is interpreted as a reflection of lost functional polarity of the microvascular endothelium in vitro resulting from deprivation of the normal influence of the components of brain parenchyma.
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PMID:Ultracytochemical characteristics of cultured goat brain microvascular endothelial cells [corrected]. 165 77

The orientation of the enzyme Mg(2+)-ATPase (EC 3.6.1.3) in the transverse tubule (TT) membranes of skeletal muscle was investigated using highly purified chicken and rabbit TT vesicles. The percentage of sealed vesicles present in these preparations averaged 88 and 78%, respectively, as calculated from the detergent-induced increase in ouabain-sensitive (Na+, K+)-ATPase activity, ATP-dependent ouabain binding, and lactate dehydrogenase activity (sarcoplasmic enzyme trapped in the TT vesicles). Sidedness of the sealed vesicles, estimated from latency of 5'-nucleotidase, acetylcholinesterase, and adenylate cyclase, was predominantly right-side out (69-76%, chicken TT and 62-70%, rabbit TT). In both chicken and rabbit native vesicles, high Mg(2+)-ATPase activity was detected by addition of ATP to the extravesicular medium; this activity was increased 14-12% by alamethicin pointing to the external localization of the active site. Furthermore, the enzymatic activity resulted partially inhibited by treatment of the chicken TT vesicles with proteinase K or p-hydroxymercuribenzoate. Concanavalin A stimulated 4-fold the chicken TT Mg(2+)-ATPase activity, an effect not potentiated by detergent permeabilization of the intact vesicles, indicating that lectin-binding sites were also solvent accessible. This stimulatory effect was not observed in native or permeabilized rabbit TT vesicles. From these results we conclude that the TT Mg(2+)-ATPase is an ectoenzyme with its nucleotide-hydrolyzing site and glycosylated regions facing the extracellular space. Inhibitors of ion-motive ATPases did not modify the enzyme activity, suggesting a different physiological role for the TT Mg(2+)-ATPase which may be involved in the regulation of muscle fiber functions affected by extracellular ATP levels.
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PMID:Transverse tubule Mg(2+)-ATPase of skeletal muscle. Evidence for extracellular orientation of the chicken and rabbit enzymes. 166 Apr 76


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