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:3.6.1.3 (ATPase)
65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An ATP diphosphohydrolase (EC 3.6.1.5) from the pancreas of the pig has been characterized and purified. The enzyme which has an optimum pH between 8 and 9 is specific for diphospho- and triphosphonucleosides. The Km values for ADP and ATP are 7.4 and 7.3 x 10(-4) M, respectively, and the purified enzyme has specific activities of 13 and 15.2 mumol of Pi/min/m of protein, respectively. It requires calcium or magnesium ions and it is insensitive to ATPase inhibitors, namely oligomycin, ouabain, and ruthenium red, and to levamisole, an inhibitor of alkaline phosphatase. Denaturation experiments, by heat and trypsin treatments, indicated that only one enzyme is involved. This is confirmed by the solubilization and purification process and by polyacrylamide gel electrophoresis. A 270-fold purification was obtained by centrifugation and successive column chromatography on Sepharose 4B and Affi-Gel blue. It is a glycoprotein with a molecular weight of 65,000 as estimated by polyacrylamide gel electrophoresis.
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PMID:Characterization and purification of a calcium-sensitive ATP diphosphohydrolase from pig pancreas. 624 96

The microsomal fraction (13 000 -60 000 x g pellet) from pea stem shows a very high divalent cation-dependent, diethylstilbestrol- and orthovanadate-inhibited ATPase and ADPase activity. No detectable inorganic or organic pyrophosphatase and adenylate kinase and almost negligible activities of mitochondrial ATPase and of other phosphomonoesterases are present in this preparation. The ATPase and ADPase activities of microsomes have been solubilized with NaClO4 and then purified by gel filtration and DEAE-Sephadex fractionation to a final specific activity of 71.5 and 102 mumol Pi/min per mg for ATP and ADP, respectively. The purified enzyme hydrolyzes triphosphonucleosides (ATP, CTP, GTP, UTP) and diphosphonucleosides (ADP and to a lesser extent CDP, UDP, IDP) and presents pH optima of 6 for ATP and 7 for ADP. It requires Mg2+, Mn2+ or Ca2+ and is inhibited by diethylstilbestrol and orthovanadate. The conclusion that the ATPase and ADPase activities belong to the same enzyme is based on the following results: (1) parallel effects of diethylstilbestrol and orthovanadate on both ATPase and ADPase; (2) parallel behavior of ATPase and ADPase throughout all the purification steps; (3) non-additivity of ATPase and ADPase and (4) lack of dilution of beta-32P formed from [beta-32P]-ATP by unlabelled ADP.
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PMID:Purification and characterization of a divalent cation-activated ATP-ADPase from pea stem microsomes. 626 9

Microsomal membranes from Cicer arietinum (chick-pea) roots contained an ATP phosphohydrolase activity that could be solubilized by high-ionic-strength media. The enzyme has been purified to homogeneity by affinity and ion-exchange chromatography. It has the properties of an ATP diphosphohydrolase (apyrase, EC 3.6.1.5) that hydrolyses different nucleoside di- and tri-phosphates but has no activity towards monophosphoric esters and pyrophosphate. No stimulation by K+ could be demonstrated for either the membrane-bound or the purified enzyme, and therefore it would seem not to be related to the K+ -dependent ATPase postulated to mediate K+ transport in plants.
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PMID:Purification and characterization of a membrane-bound ATP diphosphohydrolase from Cicer arietinum (chick-pea)roots. 627 35

Enzyme cytochemical and immunocytochemical techniques at the light and electron microscope levels were used to study the distribution of potential markers of chemical transformation in rodent bladders. In rat tumours induced by in vivo treatment with methylnitrosourea, alkaline phosphatase localization was normal on the external surface of the plasma membranes of some cells but abnormal in others where reaction product was seen only on intracellular membranes. 5'-Nucleotidase localization was abnormal in all cells, being seen on endoplasmic reticulum and nuclear membranes only, while in normal bladders only ectoenzyme localization was seen. Heterogeneity of alkaline phosphatase amd 5'-nucleotidase localization was seen on the plasma membranes of these tumours after 15 days in organ culture. Some cells produced enzyme and others did not; in other cells only parts of the membrane reacted heavily, while other regions were negative. In transformed cell cultures and tumours of mouse bladder derived by in vitro treatment of explants with dimethylbenz (a) anthracene, a bimodal pattern of alkaline phosphatase localization was seen. Cells had either normal ectoenzyme reaction product or abnormal intracellular membrane reaction product. 5'-Nucleotidase and ADPase were lost after transformation while cAMP-phosphodiesterase was retained as an ectoenzyme. Mg.ATPase and a cAMP-independent, calcium-insensitive 'protein phosphatase' were induced in transformed cell cultures. An epithelial antigen was detected in the cytoplasm of both normal and transformed cells associated with reticular cytoplasmic ground substance, plasma membrane vesicles and cytoskeletal elements.
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PMID:Cytochemical markers of bladder carcinogenesis. 627 42

In the plasma membranes from several mammalian tissues (including normal and tumor tissues), a Mg2+ (or Ca2+)-dependent ATP phosphohydrolase activity is present in much greater amount than the (Na+ + K+)-ATPase. The ouabain-insensitive activity can be attributed to at least two enzymes, an ATPase (EC 3.6.1.3) and an ATP diphosphohydrolase (EC 3.6.1.5). The ATPase hydrolyzes ATP and other nucleoside triphosphates and is not inhibited by azide. The ATP diphosphohydrolase hydrolyzes both ATP and ADP (and other nucleoside tri- and diphosphates) and the hydrolysis of adenine nucleotides is strongly inhibited by 10 mM azide. The ratios of these two enzymes in the various membranes (as determined by the extent of azide inhibition) vary widely. The ATP diphosphohydrolase accounts for most of the Mg2+ (or Ca2+)-dependent ATP hydrolysis activity of the plasma membranes of liver (mouse), kidney (dog), two mouse sarcomas, and a human astrocytoma (xenograft in athymic mice). The ATPase is more dominant in the plasma membranes from mouse brain and human oat cell carcinoma. The widespread presence of the ATP diphosphohydrolase in plasma membrane from various types of tissues is demonstrated for the first time and is of particular interest in view of its relatively high activity in the plasma membranes of two sarcomas. The membrane-bound ATP diphosphohydrolase is characterized with respect to its metal ion activators, substrates, and inhibitors. These results should facilitate the distinction of this enzyme from other ATP hydrolyzing enzymes of plasma membranes in future investigations.
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PMID:The common occurrence of ATP diphosphohydrolase in mammalian plasma membranes. 630 14

Twenty eight enzymatic activities and four macromolecular substances have been histochemically compared in rat and rabbit aortas, embedded in a common block. The study was carried out at different stages of development: 3 days, 3 months, 7-9 months and 17-19 months. In addition, lipase and cholinesterase were biochemically assayed in adult rat and rabbit aortas. The rat aortas (atheroresistant) had a better supply of aerobic oxidoreductases [linked to the pentose pathway (G6PD, 6PGD) as well as to the Krebs cycle (SD, ICD)], lipolytic enzymes (acid esterases, cholinesterase, lipase), lysosomal enzymes (acid PH/ase, Aryl-sulf/ase - Betaglu/ase), ADPase - ATPase - AlK Ph/ase Alpha GPD and acid lipids. Rabbit aortas (atherosensitive) were richer in metachromatic GAG, UDPGD (GAG Anabolism), glycogen, and related enzymes (phosphorylase, glycogen synthetase) as well as 5'-nucleotidase, Beta HBD, Lactate D and Aldolase. These differences support the hypothesis that arterial atherosensitivity is related to the activity and efficiency of smooth muscle cell energetic and catabolic processes, which govern the behaviour of lipids, proteins and carbohydrates as they penetrate the arterial wall. The factors that determine the proliferative and sclerogenic responses of arterial tissues to aggressions and, in particular, the response to lipids, remain, however, to be determined.
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PMID:A comparative study of the arterial tissue metabolism in atherosensitive and atheroresistant species. I. Comparison between rabbit and rat aortas. 734 89

The plasma membrane ATPase on the human umbilical vein endothelial cell line (ECV304) was demonstrated to be an ecto-enzyme. Hydrolysis of ATP was measured by monitoring the appearance of inorganic phosphorus. Hydrolysis of extracellular ATP was insensitive to oligomycin, vanadate, ouabain and N-ethylmaleimide, compounds that inhibit the intracellular ion pumping ATPases. Beta-Glycerophosphate (1-10 mM) or p-nitrophenyl phosphate (1-10 mM) did not inhibit hydrolysis of ATP, ruling out the involvement of non-specific phosphatases. Enzyme activity in buffer that had previously been incubated with cells was < 7%, showing that the enzyme activity measured did not result from release of intracellular enzymes. Consistent with this, the cell preparations used were estimated to be > 95% intact as judged by release of cytosolic enzyme lactate dehydrogenase. The enzyme activity was Ca2-/Mg2- dependent. Gramicidin S (20 microM), suramin (100-300 microM), chlorpromazine (250 microM), trifluoperazine (50-250 microM), and thioridazine (100 microM) inhibited the hydrolysis of ATP (3 mM) by 45-80%. The percentage inhibition produced by these substances was not altered in the presence of a concentration of alpha, beta-methylene ADP (10 microM) which inhibited hydrolysis of AMP (3 mM) by 90%, suggesting that these compounds inhibit ecto-ATPase and/or ecto-ADPase. Measurements of absolute amounts of ATP released from various tissues, including the heart, have been hindered because ATP is rapidly and sequentially hydrolysed to adenosine. Identification of compounds that inhibit ATP degradation would prove to be useful to overcome this problem and would lead to the development of invaluable pharmacological tools in many other aspects of purine research.
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PMID:Inhibition of extracellular ATP degradation in endothelial cells. 763 50

Antibodies against the holo ecto-adenosinetriphosphatase (ATPase) of rat liver and antibodies against COOH-terminal peptides of the long isoform of this enzyme reacted in Western blots with a 105-kDa band from small intestinal brush-border membranes. Indirect immunofluorescence revealed reactive proteins predominantly at the apical surface of enterocytes with some staining of basolateral membranes and of vascular endothelium. Similar results were obtained with monoclonal antibodies against HA4, a protein from rat liver closely related to the ecto-ATPase. Since these results suggested the presence of an ecto-ATPase, ATP hydrolysis was studied in intact, right-side-out brush-border membrane vesicles. Nearly half of ATP hydrolysis was caused by alkaline phosphatase (AP). Besides purine and pyrimidine trinucleotides, AP also hydrolyzed ADP, AMP, pyrophosphate, and 4-nitrophenylphosphate. Inactivation of AP by cleavage of its membrane anchor and by removal of the Zn2+ necessary for its function left the ecto-ATPase that was activated by Ca2+ and Mg2+ and hydrolyzed purine and pyrimidine trinucleotides and dinucleotides, but not AMP, pyrophosphate, and 4-nitrophenylphosphate. These features are characteristic of an ATP diphosphohydrolase (EC 3.6.1.5, also called apyrase). The physiological role of the small intestinal ecto-ATPase may be the degradation of nutrient nucleotides.
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PMID:Ecto-adenosinetriphosphatase in rat small intestinal brush-border membranes. 773 91

High-affinity ouabain binding to Na+/K(+)-ATPase (sodium- and potassium-transport adenosine triphosphatase (EC 3.6.1.37)) requires phosphorylation of the alpha subunit of the enzyme either by ATP or by inorganic phosphate. For the native enzyme (alpha/beta 1), the ATP-dependent reaction proceeds about 4-fold more slowly in the absence of Na+ than when saturating concentrations of Na+ are present. Hybrid pumps were formed from either the alpha 1 or the alpha 3 subunit isoforms of Na+/K(+)-ATPase and a chimeric beta subunit containing the transmembrane segment of the Na+/K(+)-ATPase beta 1 isoform and the external domain of the gastric H+/K(+)-ATPase beta subunit (alpha/NH beta 1 complexes). In the absence of Na+, these complexes show a rate of ATP-dependent ouabain binding from approximately 75-100% of the rate seen in the presence of Na+ depending on buffer conditions. Nonhydrolyzable nucleotides or treatment of ATP with apyrase abolishes ouabain binding, demonstrating that ouabain binding to alpha/NH beta 1 complexes requires phosphorylation of the protein. Buffer ions inhibit ouabain binding by alpha/NH beta 1 in the absence of Na+ rather than promote ouabain binding, indicating that they are not substituting for sodium ions in the phosphorylation reaction. The pH dependence of ATP-dependent ouabain binding in the presence or absence of Na+ is similar, suggesting that protons are probably not substituting for Na+. Hybrid alpha/NH beta 1 pumps also show slightly higher apparent affinities (2-3-fold) for ATP, Na+, and ouabain; however, these are not sufficient to account for the increase in ouabain binding in the absence of Na+. In contrast to phosphoenzyme formation and ouabain binding by alpha/NH beta 1 complexes in the absence of Na+, ATPase activity, measured as release of phosphate from ATP, requires Na+. These data suggest that the transition from E1P to E2P during the catalytic cycle does not occur when the sodium binding sites are not occupied. Thus, the chimeric beta subunit reduces or eliminates the role of Na+ in phosphoenzyme formation from ATP, but Na+ binding or release by the enzyme is still required for ATP hydrolysis and release of phosphate.
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PMID:The influence of beta subunit structure on the interaction of Na+/K(+)-ATPase complexes with Na+. A chimeric beta subunit reduces the Na+ dependence of phosphoenzyme formation from ATP. 777 54

The in vitro effects of phenylalanine and some of its metabolites on ATP diphosphohydrolase (apyrase, EC 3.6.1.5) activity in synaptosomes from rat cerebral cortex were investigated. The enzyme activity in synaptosomes from rats subjected to experimental hyperphenylalaninemia (alpha-methylphenylalanine plus phenylalanine) was also studied. In the in vitro studies, a biphasic effect of phenylalanine on both enzyme substrates (ATP and ADP) was observed, with maximal inhibition at 2.0 mM and maximal activation at 5.0 mM. Inhibition of the enzyme activity was not due to calcium chelation. Moreover, phenylpyruvate, when compared with phenylalanine showed opposite effects on the enzyme activity, suggesting that phenylalanine and phenylpyruvate bind to two different sites on the enzyme. The other tested phenylalanine metabolites phenyllactate, phenylacetate and phenylethylamine) had no effect on ATP diphosphohydrolase activity. In addition, we found that ATP diphosphohydrolase activity in synaptosomes from cerebral cortex of rats with chemically induced hyperphenylalaninemia was significantly enhanced by acute or chronic treatment. Since it is conceivable that ATPase-ADPase activities play an important role in neurotransmitter (ATP) metabolism, it is tempting to speculate that our results on the deleterious effects of phenylalanine and phenylpyruvate on ATP diphosphohydrolase activity may be related to the neurological dysfunction characteristics of naturally and chemically induced hyperphenylalaninemia.
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PMID:Effect of phenylalanine and its metabolites on ATP diphosphohydrolase activity in synaptosomes from rat cerebral cortex. 782 71


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