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)

The exact route of metabolism of tiazofurin, a novel nucleoside with antitumor activity, is controversial. Using human cell lines severely deficient in salvage nucleotide enzymes, we were able to identify the route of activation in tiazofurin metabolism. With loss of adenosine kinase activity by mutation in two lymphoblastoid cell lines, CCRF-CEM and WI-L2, the growth sensitivity to tiazofurin decreased by 6- and 3-fold, respectively. In contrast, the mutant lines were about 3000- to 1500- and 16- to 4-fold more resistant to the structurally similar tiazofurin analogues pyrazofurin and ribavirin, respectively. Other mutants with defective deoxycytidine or uridine kinase activity showed normal sensitivity to all three analogues. Both cell lines with defective adenosine kinase activity accumulated about 50% wild-type levels of tiazofurin-5'-monophosphate and thiazole-4-carboxamide adenine dinucleotide analogue of tiazofurin at cytotoxic concentrations of the drug. Extracts of wild-type lymphoblasts catalyzed the phosphorylation of tiazofurin in the presence of adenosine 5'-triphosphate and Mg2+. Loss of adenosine kinase activity in the mutant extract eliminated this phosphorylating activity for tiazofurin consistent with the notion that adenosine kinase catalyzes phosphorylation of tiazofurin. However, an enzyme activity that catalyzed the phosphorylation of tiazofurin in the presence of inosine-5'-monophosphate as donor and Mg2+ was detected in the extracts of both wild-type cells and adenosine kinase-deficient mutants. The monophosphate donor specificity, divalent metal, high salt requirement, and nucleoside acceptor specificity of this enzyme activity paralleled that of a 5'-nucleotidase (EC 3.1.3.5) which catalyzes inosine phosphorylation. In addition, tiazofurin phosphorylation was competitively inhibited by inosine and the apparent Ki value was similar to the apparent Km value for inosine phosphorylation. These results indicate that two enzymes, adenosine kinase and a cytoplasmic 5'-nucleotidase, are functionally important anabolizing enzymes for tiazofurin in human cells.
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PMID:Tiazofurin metabolism in human lymphoblastoid cells: evidence for phosphorylation by adenosine kinase and 5'-nucleotidase. 300 May 75

Inhibition of cardiovascular Na,K-pump activity has been shown to promote an increase in the contractile activity of myocardial and vascular smooth muscle and a consequent rise in blood pressure (BP). It has also been shown that vascular Na,K-pump activity and myocardial Na+K+ATPase activity [the energy source for active sodium (Na) and potassium (K) transport] are decreased in rats with various forms of low renin hypertension including rats with reduced renal mass-saline (RRM-saline) hypertension. In the present study, left ventricular Na+K+ATPase activity from rats with RRM-saline hypertension was found to be decreased in membranes prepared by two independent methods: deoxycholate, sodium iodide (Nal)-treated microsomal fractions (method 1) and membranes prepared by the hypotonic, lithium bromide (LiBr) method (method 2). Relative to RRM normotensive control rats which drank distilled water, myocardial Na+K+ATPase activity from RRM-saline drinking rats was decreased by 18.2% in membranes prepared by method 1 and 33.6% in membranes prepared by method 2. The apparent affinities of Na+K+ATPase for K and for ouabain were unaltered relative to controls in membranes prepared from these hypertensive rats by method 1, and the sialic acid content and 5'-nucleotidase activity (two putative sarcolemmal markers) were unaltered in membranes from the hypertensive rats, prepared by methods 1 and 2 respectively. The Mg2+ATPase activity of membranes prepared by method 1 was increased in the RRM-saline hypertensive rats but because it was not increased in membranes prepared by method 2 the former observation does not appear to be of any pathophysiological importance. In other experiments, hypertension was reversed in RRM-saline hypertensive rats by restricting their salt intake (substitution of distilled water for drinking).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Decreased myocardial Na+K+ATPase activity in rats with reduced renal mass-saline hypertension. 300 89

Intact synaptosomes isolated from the electric organ of the electric ray Torpedo marmorata contain, at their surface, enzyme activities for the hydrolysis of externally applied nucleoside phosphates. The diazonium salt of sulfanilic acid, as a low-molecular-weight, slowly permeating, covalent inhibitory agent, selectively blocks these enzyme activities and leaves intracellular lactate dehydrogenase intact. The ectoenzymes comprise both a nucleoside triphosphate and diphosphate phosphohydrolase, as well as a 5'-nucleotidase. Activity of nonspecific ectophosphatases is absent. The nucleoside triphosphatase hydrolyzes almost equally well ATP, GTP, CTP, UTP, and ITP and is activated to a similar degree by Mg2+ or Ca2+. It has a high affinity for ATP (Km for ATP in the presence of Mg2+, 75 microM; in the presence of Ca2+, 66 microM). Maximal rates in the presence of Mg2+ and Ca2+ were very similar (34.8 and 32.5 nmol of Pi/min/mg of synaptosomal protein, respectively). Either Mg-ATP or Ca-ATP can act as a true substrate. ADP inhibits hydrolysis of ATP, but AMP is without effect. The nucleoside triphosphatase is not inhibited significantly by a number of inhibitors of mitochondrial Mg2+-ATPase or of Ca2+ + Mg2+-ATPases. It is, however, considerably inhibited by filipin and quercitin. The capacity of intact synaptosomes to hydrolyze also extracellular ADP, GDP, AMP, GMP, and IMP suggests that the nucleoside triphosphatase is part of an enzyme chain that causes complete hydrolysis of the respective nucleoside triphosphate to the nucleoside. We conclude that the cholinergic nerve terminals of the Torpedo electric organ can hydrolyze ATP released on coexocytosis with acetylcholine via an ectonucleoside triphosphatase activity that is different from known endogenous nerve terminal ATPases. The final product of the hydrolysis, adenosine, can then be salvaged by the nerve terminal for resynthesis of ATP. Other possible physiological functions of the ectonucleotidases are discussed.
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PMID:Ectonucleotidase activities associated with cholinergic synaptosomes isolated from Torpedo electric organ. 301 88

Membrane vesicles were prepared from CHO-K1 and alanine-resistant transport mutants, alar4 and alar4-H3.9. Alar4 is a constitutive mutant of the A system, and alar4-H3.9, derived from alar4, may be the result of amplification of a gene coding for an A-system transporter. Under conditions in which the same membrane potential (interior negative) and Na+ gradient were employed, the mutant vesicles show increases in the A system over that of the parental CHO-K1 cell line, paralleling, but not equivalent to, that found in whole cells. L-system and 5'-nucleotidase activities of these vesicles were similar, indicating that the increased A-system activity of the mutant vesicles is not due to the differential enrichment of the A system in these vesicles. The membrane potential was produced by a K+ diffusion gradient (internal greater than external) in the presence of valinomycin or by the addition of a Na+ salt of a highly permeant anion such as SCN-. Monensin was employed to study the effect of the Na+ gradient on transport and membrane potential. The latter was determined by measuring the uptake of tetraphenylphosphonium ion. A negative membrane potential determines the concentrative ability and the initial velocity of the A system in these vesicles. The concentration of external Na+ has a stimulatory effect on the initial velocity of this system. However, the Na+ gradient (external greater than internal) has no effect on the initial velocity or the membrane potential when the potential is set by valinomycin and high internal K+. Little if any ASC system could be detected in vesicles from CHO-K1.
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PMID:Amino acid transport in membrane vesicles from CHO-K1 and alanine-resistant transport mutants. 360 29

Two plasma membrane fractions from ascites tumour cells with differences in vesicle size were isolated by gel-exclusion chromatography on Sephacryl S-1000. Fraction 1 appeared in the void volume and had a vesicle diameter in the range 300-400 nm. Fraction 3, with an equilibrium constant (Kd) of 0.58, consisted of vesicles between 100 and 200 nm in diameter as measured by routine size analysis with the electron microscope and by calibration of the column with latex beads. The appearance of two plasma membrane fractions could also be confirmed by iodination of the surface membrane prior to fractionation. This gel chromatographic procedure represents a rapid and convenient method for the isolation of membrane material, which was enriched between five- and fourteen-fold based on the specific activity of the membrane-bound marker enzymes. Fraction 1 contained small amounts of lysosomal and Golgi membranes, and fraction 3 some material of the Golgi apparatus and the endoplasmic reticulum. The major portion of the contaminating membraneous material remained on the column and could be eluted with high salt buffer. The two plasma membrane fractions revealed some differences in 5'-nucleotidase specific activity and in the protein pattern, especially in the higher molecular weight range, as shown by sodium dodecyl sulphate gel electrophoresis.
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PMID:Preparation of two plasma membrane fractions from ascites tumour cells by gel chromatography on Sephacryl S-1000. 378 79

In the previous paper we presented findings which indicated that enzyme heterogeneity exists among PMN leukocyte granules. From histochemical staining of bone marrow smears, we obtained evidence that azurophil and specific granules differ in their enzyme content. Moreover, a given enzyme appeared to be restricted to one of the two types. Clear results were obtained with alkaline phosphatase, but those with a number of other enzymes were suggestive rather than conclusive. Since the approach used previously was indirect, it was of interest to localize the enzymes directly in the granules. Toward this end, we carried out cytochemical procedures for five enzymes on normal rabbit bone marrow cells which had been fixed and incubated in suspension. The localization of reaction product in the granules was determined by electron microscopy. In accordance with the results obtained on smears, azurophil granules were found to contain peroxidase and three lysosomal enzymes: acid phosphatase, arylsulfatase, and 5'-nucleotidase; specific granules were found to contain alkaline phosphate. Specific granules also contained small amounts of phosphatasic activity at acid pH. Another finding was that enzyme activity could not be demonstrated in mature granules with metal salt methods (all except peroxidase); reaction product was seen only in immature granules. The findings confirm and extend those obtained previously, indicating that azurophil granules correspond to lysosomes whereas specific granules represent a different secretory product.
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PMID:Differences in enzyme content of azurophil and specific granules of polymorphonuclear leukocytes. II. Cytochemistry and electron microscopy of bone marrow cells. 569 83

In model experiments using human erythrocytes, glycochenodeoxycholate caused extensive membrane damage (as judged by release of membrane phospholipid and acetylcholinesterase and by cell lysis) at approximately 10-fold lower concentrations than glycocholate. Chenodeoxycholate feeding had no effect upon the total protein, bile salt or phospholipid concentration of rat bile, although evidence is presented to suggest an expansion of the bile salt pool occurred. Rats fed chenodeoxycholate showed a dose-dependent enrichment of this bile acid in bile; this occurred mainly at the expense of cholate. Chenodeoxycholate feeding resulted in an increased biliary output of the plasma membrane enzymes alkaline phosphatase and 5'-nucleotidase; the hepatic activities of these enzymes were also increased. In contrast, the biliary output and hepatic activities of two other plasma membrane enzymes, alkaline phosphodiesterase I and L-leucine-beta-naphthylamidase, were unaffected by chenodeoxycholate feeding. A greater proportion of all four plasma membrane enzymes studied existed in bile of chenodeoxycholate-fed rats in a "soluble" form (as judged by their remaining in the supernatant on centrifugation of bile). These results are discussed in relation to the origin of plasma membrane enzymes in bile and to the potential toxicity of chenodeoxycholate and its conjugates to the membranes of the hepatobiliary system.
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PMID:Effect of chenodeoxycholate feeding upon the biliary output of plasma membrane enzymes in the rat. 608 20

We demonstrate that 5'-nucleotidase (5'NT), an ectoenzyme of guinea pig polymorphonuclear leukocytes, is largely excluded from phagosomal membrane, rather than internalized randomly during phagocytosis of heat-killed bacteria, latex microbeads, or zymosan particles. Cells were fixed in 0.25% glutaraldehyde (pH 6.3) at 4 degrees C for 10 min and incubated in a cytochemical medium for the demonstration of 5'NT. In the nonphagocytosing cells, 5'NT was evenly distributed on the external side of the plasma membrane. In cells phagocytosing bacteria, 5'NT appeared to be cleared from the nascent phagosomal membrane; after 5 min of phagocytosis, most of the phagocytic vacuoles containing bacteria, latex, or zymosan particles were devoid of reaction product. When phagosomes containing latex particles were isolated and biochemically assayed, they contained less than 3% of the total cellular 5'NT activity even after 60 min of phagocytosis, and at that time the total cellular 5'NT activity had not declined. When the diazonium salt of sulfanilic acid (DASA), a nonpermeable ectoenzyme inhibitor, was used to determine the distribution of extracellular and intracellular 5'NT activity, no increase in DASA-insensitive intracellular 5'NT was found after phagocytosis of latex or opsonized zymosan. Cytochemical and biochemical evidence led us to conclude that 5'NT is excluded from phagosomal membrane, and that the exclusion is due to redistribution rather than to inactivation by granule enzymes.
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PMID:Redistribution of ecto-5'-nucleotidase during phagocytosis by guinea pig polymorphonuclear leukocytes. 609 May 56

In experimental bile obstruction the serum activities of the membrane-bound liver enzymes, alkaline phosphatase, 5'-nucleotidase and gamma-glutamyltransferase are greatly increased, whereas in the liver only the alkaline phosphatase activity is elevated. After partial hepatectomy or tetrachloride poisoning the alkaline phosphatase activity in the regenerating live is increased to the same extent as in cholestasis without an accompanying elevation in serum activity. The following results support the hypothesis of a bile salt-mediated solubilization of membrane-bound enzymes in cholestatic liver: (1) 30 min after bile duct ligation the total bile acids in the liver were increased 5-fold, 2 h later as much as 10-fold. After 1 day, the bile acid concentration was still 4 times above normal. (2) Isolated plasma membranes from normal and obstructed livers were incubated in vitro with increasing amounts of tri- and dihydroxycholanic acids. At a final concentration of 1 mmol/l taurochenodeoxycholate significant amounts of membrane-bound enzymes were released into the 12,000-g supernatant. (3) In the regenerating liver, where tissue phsophatase activity was high and serum phosphatase activity unchanged, the bile salt concentration was not increased.
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PMID:Studies on the mechanism of the increase in serum alkaline phosphatase activity in cholestasis: significance of the hepatic bile acid concentration for the leakage of alkaline phosphatase from rat liver. 612 56

Several enzymes associated with the hepatocyte cell surface, alkaline phosphatase (AP), 5'-nucleotidase (5'N), Mg++- and total Na+K+Mg++-ATpase, were assayed and localized cytochemically in order to gain insight into alterations of the plasma membrane components during reassociation of hepatocytes in primary monolayer culture. During a period of 4 days the activities of 5'nucleotidase and alkaline phosphatase increased spontaneously up to three- and four-fold, respectively. Dexamethasone reinforce the rise of alkaline phosphatase activity but retarded the increase of that of 5'nucleotidase. However, after the third day the level of 5'nucleotidase activity converged with the untreated controls. The activities of Mg++- and Na+K+Mg++-ATPase, which closely paralleled each other, remained essentially unchanged throughout cultivation and were not affected by dexamethasone. Cytochemical demonstration of alkaline phosphatase, 5'nucleotidase and Mg++-ATPase, using the lead salt method, revealed the potential presence of reaction product on the whole cell surface. However, the cells did not react uniformly, particularly on bile canalicular membranes. This heterogeneity seems to be due to different stages of canalicular development and to different functional states of the cultured hepatocytes.
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PMID:Alterations in activity and ultrastructural localization of several phosphatases on the surface of adult rat hepatocytes in primary monolayer culture. 612 58


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