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)

This paper reports on 1H and 31P NMR as well as EPR measurements of the labeling reagent of ATPase sites, "Co(III)-(phen)-ATP." This complex is found to be paramagnetic, as deduced both from its EPR spectrum and from the significant broadening, though almost unshifted, proton and phosphorus resonances. This paramagnetism is a result of the incorporation of the superoxide free-radical anion in the coordination sphere of the trivalent cobalt ion. Evidence for the presence of superoxide in the complex is based on competition experiments with cyanide, which is able to displace the superoxide anion. The latter was identified by its inducing effect on the photoreactivity of luminol. The displacement of superoxide by cyanide was accompanied by the abolition of the paramagnetism of the complex. The relative distances between the protons and phosphorus atoms of ATP and the superoxide anion in the complex were calculated using the NMR line-broadening data. Structural models compatible with the experimental results are proposed. Under conditions of excess of adenine nucleotides or phenanthroline, the coordinated ATP molecule becomes exchangeable. This phenomenon is attributed to the labilization of the cobaltic ion ligands induced by the superoxide anion.
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PMID:Structural and exchange properties of "Co(III)-phenanthroline-ATP": a labeling reagent for the active site of ATPases. 21 Aug 47

In order to study the action of the divalent cation which is essential for phosphorylation of sodium- and potassium-transport adenosine triphosphatase, magnesium ion, the normal ligand, was replaced with calcium ion, which had properties diffeerent from those of Mg2+, Mn2+, Fe2+, Co2+, Ni2+, or Zn2+. Phosphorylation of the enzyme from ATP at pH 7.4 in the presence of Na+ and Ca2+ yielded a Ca.phosphoenzyme (60% of the maximal level) with a normal rate of dephosphorylation following a chase with unlabeled Ca.ATP (PK = 0.092S-1 at 0 degrees C). In contrast, after a chase by a chelator, namely ethylenediaminetetraacetic acid, 1,2-cyclohexylenedinitrilotetraacetic acid, or ethylene glycol bis-(beta-aminoethyl ether)N,N'-tetraacetic acid, dephosphorylation slowed within 5 s and half of the initial phosphoenzyme remained with a stability about 5-fold greater than normal. Three states of the phosphoenzyme were distinguished according to their relative sensitivity to ADP or to K+ added during a chase. Normally prepared Mg.phosphoenzyme was sensitive to K+ but not to ADP; Ca.phosphoenzyme was sensitive either to ADP or to K+; and the stabilized phosphoenzyme prepared from Ca.phosphoenzyme by addition of a chelator was sensitive neither to ADP nor to K+ nor to both together. Addition of Ca2+ to the stabilized phosphoenzyme restored the reactivity to that of Ca.phosphoenzyme. Addition of Mg2+ to the stabilized phosphoenzyme changed the reactivity to that of Mg.phosphoenzyme. Therefore, this unreactive, stabilized state of the phosphoenzyme appeared to be a divalent cation-free phosphoenzyme. With respect to sensitivity to ouabain, Ca.phosphoenzyme was as sensitive as Mg.phosphoenzyme but calcium-free phosphoenzyme was much less sensitive. It was concluded that the divalent cation required for phosphorylation normally remains tightly bound to the phosphoenzyme and is required for normal reactivity. Calcium ion was almost unique in dissociating relatively easily from the phosphoenzyme. Strontium ion appeared to act similarly to Ca2+.
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PMID:Binding of divalent cation to phosphoenzyme of sodium- and potassium-transport adenosine triphosphatase. 21 Nov 32

The central nervous systems of web-building spiders (Araneidae, Agelenidae) and hunting spiders (Lycosidae, Salticidae) were tested for non-specific and specific phosphatases. Acid phosphatase exhibited weakly to moderately positive reactions in the neuronal cell bodies and in the neuropile fibre mass of all species investigated. Alkaline phosphatase could only be demonstrated in the external and internal neural lamellae of the brain and ventral cord of several specimens of the araneid species investigated. Tests for thiamine pyrophosphatase were negative with both the lead and calcium-cobalt methods. Distinctive positive reactions for adenosine triphosphatase were visible in the nervous system of all the species used, being especially strong in the optic ganglia of the hunting spiders. The demonstration of adenosine triphosphatase was only possible when applying the calcium-cobalt method after Padykula and Herman, while the lead method after Wachstein and Meisel did not produce any staining reaction at all. Controls of the histochemical reaction showed that the enzyme was activated by Ca2+ and inhibited by sulphydryl destroying reagents (e.g. PCMB), but was insensitive to ouabain. It could be probably classified as a mitochondrial proton-translocating adenosine triphosphatase.
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PMID:Phosphatases in the central nervous system of spiders (Arachnida, Araneae). 21 10

The nucleoside triphosphatase activities of the nuclear envelopes from rat liver, pig liver and simian-virus-40-transformed mouse-embryo 3T3 cells were shown to exhibit similar parperties. All three preparations hydrolyse ATP, 2'-dATP, 3'-dATP, GTP, CTP and UTP in the presence of Mg2+, Ca2+, Mn2+ and Co2+ with a pH optimum of 8.0, are sensitive to inhibition by mercurials, arsenicals, quercetin, proflavin and adenosine 5'-[gamma-thio]triphosphate and are partially inactivated by exposure to high ionic strength. The kinetic behaviour is similar for all substrates irrespective of the source of material. The typical Eadie-Hofstee plot, which is concave upwards at pH 8.0 when the ionic strength is 20mM, becomes linear when the pH is increased to 8.5 or the ionic strength to 160mM. The overall evidence, particularly the labelling of only one polypeptide by [gamma-32P]ATP, suggests that under the conditions of preparation and assay used only one class of nucleoside triphosphatase active sites is detectable in nuclear envelopes. The importance of these results for an understanding of the role of the enzyme in vivo is discussed.
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PMID:Properties of mammalian nuclear-envelope nucleoside triphosphatase. 22 21

1. A stable ATPase complex with sensitivity to dicyclohexylcarbodiimide (TFo-F1) was purified from the membranes of the thermophilic aerobic bacterium PS3, by ion exchange chromatography in the presence of Triton X-100. 2. The ATPase of TFo-F1 was maximal at 70 degrees at pH 8.6 and was stable after monomerization in 4 M urea and 0.5% Triton X-100 at 25 degrees. The activity was dependent on Mg2+, Co2+, or Mn2+, and it became insensitive to dicyclohexylcarbodiimide when Ca2+ or Cd2+ was added instead. 3. TFo-F1 required P-lipids of this bacterium contained branched fatty acyl groups but no unsaturated groups and were stable against oxidation and heat. 4. Studies by electron microscopy, gel electrophoresis, and use of anti-ATPase antibody and [3H]acetyl-ATPase indicated that the TFo-F1 complex was composed of an ATPase moiety (TF1, five different subunits) and a hydrophobic moiety (TFo, three different subunits. TFo conferred TF1 with sensitivity to dicyclohexylcarbodiimide. 5. Vesicles catalyzing 32Pi-ATP exchange and ATP-driven enhancement of fluorescence of anilinonaphthalene sulfonate were reconstituted by dialyzing pure TFo-F1 and P-lipids together, and were active even at 50-75 degrees. The vesicles reconstituted from TFo-F1 and bacterial P-lipids were more stable than those reconstituted from TFo-F1 and soybean P-lipids.
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PMID:Purification and properties of a dicyclohexylcarbodiimide-sensitive adenosine triphosphatase from a thermophilic bacterium. 24 Aug 43

The subcellular distributions of alkaline phosphates I (the major activity of prepubertal mouse ovaries) and alkaline phosphatase Ib (a kinetically distinct isoenzyme induced in large amounts by injection of human chorionic gonadotropin or luteinizing hormone) were studied by differential rate centrifugation and discontinuous density gradient centrifugation of ovarian homogenates from control and gonadotropin-treated mice. The distributions of the two alkaline phosphatases were alike and were similar to those of nucleotidase, Mg2+ -dependent ATPase and Co2+ -stimulated naphthylamidase activities, suggesting that they were associated with plasma-membrane vesicles.
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PMID:Subcellular distribution of a gonadotropin-induced form of mouse ovarian alkaline phosphatase. 100 1

V79 Chinese hamster cells were used as a model for the characterization of the Co(II) uptake into mammalian cells as well as the mechanisms involved. Co(II) was taken up in a dose and time dependent manner. The uptake was exponential without saturation in the tested concentration range up to 400 microM CoCl2. Furthermore, there was a high intracellular cobalt accumulation at elevated extracellular Co(II) doses (up to 16 fold at 200 microM). The time course of Co(II) uptake showed a maximum after about 8-12 h with no further change after the longest tested incubation time (24 h). The uptake of Co(II) into V79 cells seems to be mediated by multiple mechanisms: active, energy consuming transport like ion pumps and endocytosis, since the Co(II) uptake was significantly reduced by ouabain (an inhibitor of the Na+/K+ATPase), N-ethylmaleinimide (an inhibitor of the Ca2+/Mg2+ATPase and the Na+/K+ATPase), chlorpromazine (a calmodulin antagonist and inhibitor of the Ca2+/Mg2+ ATPase) as well as by the endocytosis inhibitor chloroquine. Furthermore, the two agents iodoacetate and potassium cyanide, which produce ATP depletion, resulted in a diminution of the intracellular cobalt concentration. An uptake through anion channels could be excluded, since 4,4'-diisothiocyanostilbene-2,2'-disulphonic acid was not inhibitory.
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PMID:Mechanisms of cobalt(II) uptake into V79 Chinese hamster cells. 128 1

Hydrolysis of extracellular ATP and other nucleoside phosphates by A-431 human epidermoidal carcinoma cells was studied. The hydrolysis of extracellular ATP by these cells required either Mg2+ or Ca2+, and either cation could be replaced by Co2+, Fe2+, or Mn2+. Nucleoside triphosphates (ATP, GTP, CTP, UTP, and dTTP), but not nucleoside diphosphates, were hydrolyzed by the cells with Km and Vmax values similar to those for ATP (0.9-1.1 mmol/l and 6-10 nmol Pi formed/10(6) cells, respectively). The hydrolysis of ATP was inhibited strongly by ATP-gamma S and AMPPNP, and weakly by AMPCPP and ADP-beta S, but not by AMPCPP or AMPCP. Since the hydrolysis of [gamma-32P]ATP was inhibited by all these nucleoside triphosphates, the binding site for ATP is presumed to be the same as that for the other nucleoside triphosphates. All these results indicate that ecto-ATPase activity associated with A-431 cells is due to ecto-nucleoside triphosphatase. The nucleotide specificity shown in the present study indicates that ecto-nucleoside triphosphatase associated with A-431 cells is a molecule different from P2-purinergic receptors which can be stimulated specifically with nucleoside phosphates like ATP, ADP, UTP, UDP, and GTP, but not by other nucleotides.
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PMID:Characterization of ecto-nucleoside triphosphatase on A-431 human epidermoidal carcinoma cells. 129 31

We have studied extracellular ionic changes induced by iontophoretic application of excitatory amino acids in rat hippocampal slices. In contrast to kinetics of changes in [Ca2+]o, kinetics of changes in [K+]o, [Na+]o, [Cl-]o as well as in extracellular space size were comparable for different glutamate receptor agonists. Thus, alpha-amino-3-hydroxy-5-methylisoxazolepropionic acid (AMPA), quisqualate (quis), and kainate caused reductions in [Ca2+]o followed by an increase of [Ca2+]o above baseline, whereas glutamate, aspartate, N-methyl-D-aspartate (NMDA), and DL-homocysteic acid caused only reductions in [Ca2+]o. After blocking the NMDA receptors with ketamine and 2-amino-5- phosphonovaleric acid (2-APV), glutamate-induced decreases in [Ca2+]o were followed by an overshoot. Reduction of the transmembrane Na+ gradient by lowering [Na+]o, blocking of the Na(+)-K+ ATPase by lowering [K+]o, and application of ouabain blocked the overshoots after quis application, whereas vanadate, a blocker of the Ca(2+)-Mg2+ ATPase, had no effects. Lithium enhanced the reductions in [Ca2+]o and blocked the overshoots. Amiloride also reduced the overshoots. All organic Ca2+ entry blockers diminished reductions of [Ca2+]o but increased the overshoots. Inorganic Ca2+ antagonists had variable effects. Ni2+ had similar effects as the organic Ca2+ entry blockers while Cd2+ reduced both the [Ca2+]o decreases as well as the subsequent overshoots. Co2+ had initially a similar action as Ni2+. With prolonged application, [Ca2+]o decreases became augmented and, during wash, overshoots could no longer be elicited. We suggest that the overshoots in [Ca2+]o are due to a combined effect of extracellular space shrinkage and activation of the Na+/Ca2+ exchangers. This would imply that NMDA receptor activation blocks extrusion of Ca2+ from the cells. We tested the hypothesis that quis-induced intracellular Ca2+ release and extrusion of Ca2+ from the cells contributed to the overshoots. Dantrolene was without effect on the quis-induced signals, while ryanodine reduced the overshoots. Caffeine on the other hand diminished the [Ca2+]o decreases with no effects on the overshoots. To test for possible second messenger routes by which NMDA receptor activation might slow Ca2+ extrusion from cells, we investigated the effects of arachidonic acid and N-monomethyl-D- arginine on the quis-induced signals. While these agents reduced decreases in [Ca2+]o, they had no clear effects on the overshoots. Thus a possible route by which NMDA receptor activation may affect Ca2+ extrusion from cells has still to be elucidated.
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PMID:Pharmacological properties of excitatory amino acid induced changes in extracellular calcium concentration in rat hippocampal slices. 129 71

Both phosphointermediate- and vacuolar-type (P- and V-type, respectively) ATPase activities found in cholinergic synaptic vesicles isolated from electric organ are immunoprecipitated by a monoclonal antibody to the SV2 epitope characteristic of synaptic vesicles. The two activities can be distinguished by assay in the absence and presence of vanadate, an inhibitor of the P-type ATPase. Each ATPase has two overlapping activity maxima between pH 5.5 and 9.5 and is inhibited by fluoride and fluorescein isothiocyanate. The P-type ATPase hydrolyzes ATP and dATP best among common nucleotides, and activity is supported well by Mg2+, Mn2+, or Co2+ but not by Ca2+, Cd2+, or Zn2+. It is stimulated by hyposmotic lysis, detergent solubilization, and some mitochondrial uncouplers. Kinetic analysis revealed two Michaelis constants for MgATP of 28 microM and 3.1 mM, and the native enzyme is proposed to be a dimer of 110-kDa subunits. The V-type ATPase hydrolyzes all common nucleoside triphosphates, and Mg2+, Ca2+, Cd2+, Mn2+, and Zn2+ all support activity effectively. Active transport of acetylcholine (ACh) also is supported by various nucleoside triphosphates in the presence of Ca2+ or Mg2+, and the Km for MgATP is 170 microM. The V-type ATPase is stimulated by mitochondrial uncouplers, but only at concentrations significantly above those required to inhibit ACh active uptake. Kinetic analysis of the V-type ATPase revealed two Michaelis constants for MgATP of approximately 26 microM and 2.0 mM. The V-type ATPase and ACh active transport were inhibited by 84 and 160 pmol of bafilomycin A1/mg of vesicle protein, respectively, from which it is estimated that only one or two V-type ATPase proton pumps are present per synaptic vesicle. The presence of presumably contaminating Na+,K(+)-ATPase in the synaptic vesicle preparation is demonstrated.
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PMID:Characterization of the P-type and V-type ATPases of cholinergic synaptic vesicles and coupling of nucleotide hydrolysis to acetylcholine transport. 131 70


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