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)

Modification of soluble mitochondrial ATPase (factor F1) by spin-labelled iodoacetamide and spin-labelled methyleneketone does not cause and change in the catalytic properties of the enzyme. The temperature dependence of tau corr. of labels bound to factor F1 testifies to conformational changes in the enzyme at temperatures of 18--20 degrees C and 34--37 degrees C. At these temperature intervals, breaks are observed in the temperature dependence of the ATPase reaction rate in the Arrenius plot. The results obtained indicate that the thermally induced conformational changes in factor F1 affect large areas of the protein molecule. The interaction of factor F1 with the hydrophobic spin probes, namely fatty acid derivatives, was studied. It was shown that the interaction of foctor F1 with Mg2+, Mg-ATP, Mg-ADP and ADP, results in an increase in the ability of the enzyme to adsorb spin probes.
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PMID:[Conformational changes in soluble mitochondrial ATPase by the spin probe method]. 15 72

1. The distribution of ATPase and several marker enzymes was examined after differential and sucrose gradient centrifugation of yeast homogenates. 2. An ATPase activity not sensitive to oligomycin is found exclusively associated with a particulate fraction equilibrating at densities of 1.23-1.25. This particulate material shows the chemical and enzymatic characteristics of the yeast plasma membrane. 3. The pH optimum of the plasma membrane ATPase is 5.6, as compared with 8.5 for the mitochondrial ATPase. In addition to oligomycin, the enzyme is not sensitive to other inhibitors of the mitochondrial ATPase as azide, dicyclohexylcarbodiimide and the mitochondrial ATPase inhibitor protein. It is inhibited by p-chloromercuryphenyl sulfonate, fluoride, quercetin and by the antibiotic Dio-9 but is not affected by ouabain. 4. The plasma membrane ATPase shows a high affinity for ATP (Km = 0.1 mM) and is very specific for this compound, hydrolyzing other nucleotide triphosphates less than 25% as rapidly. No activity was detected with ADP. 5. The enzyme requires a divalent cation for activity and Mg2+ is the most effective. It is not significantly stimulated by K+ or bicarbonate and Ca2+ is inhibitory. 6. The activity cannot be assayed in intact cells unless they are permeabilized with toluene. This suggest that the active site is on the cytoplasmic side of the plasma membrane.
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PMID:Characterization of the plasma membrane ATPase of Saccharomyces cerevisiae. 15 59

The phospholipid requirement of membrane-bound enzymes may depend on several reasons. In our laboratory we have investigated lipids (1) as a bidimensional medium required for the movement of Coenzyme Q, a lipid-soluble cofactor of the mitochondrial respiratory chain, and (2) as a hydrophobic environment necessary to impose the proper conformation to membrane-bound enzymic proteins. We have found that Coenzyme Q, once reduced by NADH dehydrogenase, must cross the inner mitochondrial membrane; only quinones having long isoprenoid side chains can easily cross phospholipid bilayers, and this is the reason why a short chain quinone such as CoQ-3 inhibits NADH oxidation. The incapability of short quinones to cross lipid bilayers is due to their disposition in the lipid bilayer, stacked within the phospholipids. The conformational role of lipids has been investigated indirectly observing the kinetics of membrane-bound enzymes, e.g. the mitochondrial ATPase, and directly by circular dichroism. Lipid removal or lipid perturbation with organic solvents induce a decrease of alpha-helical content in mitochondrial proteins, and give rise to a series of kinetic changes in ATPase, including uncompetitive inhibition, increased activation energy, and loss of cooperativity in oligomycin inhibition. The recognition of a conformational role of lipids has allowed us to postulate a working hypothesis for the mechanism of action of general anesthetics. Such drugs have been found by us, by means of spin labels and fluorescent probes, to disrupt lipid protein interactions in several membranes, including synaptic membranes. The loosening of such interactions is believed to induce conformational changes, which will alter ion transport systems necessary to the propagation of neural impulses. Conformational changes induced by anesthetics have been found by us both directly by circular dichroism and indirectly by enzyme kinetics. The conformational effect of anesthetics is not directly exerted on the proteins but is mediated through the lipids. In agreement with this hypothesis we have found that membrane-bound acetylcholinesterase is inhibited by anesthetics, whereas the solubilized enzyme is not inhibited. However, binding of the solubilized enzyme to phospholipids restores anesthetic inhibition.
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PMID:Biophysical studies on agents affecting the state of membrane lipids: biochemical and pharmacological implications. 15 58

Incubation of mitochondria from Neurospora crassa and Saccharomyces cerevisiae with the radioactive ATPase inhibitor [14C]dicyclohexylcarbodiimide results in the irreversible and rather specific labelling of a low-molecular-weight polypeptide. This dicyclohexylcarbodiimide-binding protein is identical with the smallest subunit (Mr 8000) of the mitochondrial ATPase complex, and it occurs as oligomer, probably as hexamer, in the enzyme protein. The dicyclohexylcarbodiimide-binding protein is extracted from whole mitochondria with neutral chloroform/methanol both in the free and in the inhibitor-modified form. In Neurospora and yeast, this extraction is highly selective and the protein is obtained in homogeneous form when the mitochondria have been prewashed with certain organic solvents. The bound dicyclohexylcarbodiimide label is enriched in the purified protein up to 50-fold compared to whole mitochondria. Based on the amino acid analysis, the dicyclohexylcarbodiimide-binding protein from Neurospora and yeast consists of at least 81 and 76 residues, respectively. The content of hydrophobic residues is extremely high. Histidine and tryptophan are absent. The N-terminal amino acid is tyrosine in Neurospora and formylmethionine in yeast.
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PMID:The dicyclohexylcarbodiimide-binding protein of the mitochondrial ATPase complex from Neurospora crassa and Saccharomyces cerevisiae. Identification and isolation. 15 5

The effect of organic solvents on the beef heart mitochondrial ATP-base-catalyzed ATP and ITP hydrolysis was examined. It was observed that numerous organic solvents stimulated ATP hydrolysis while ITP hydrolysis was inhibited. Methanol at 20% (v/v) was found to stimulate ATP hydrolysis by over 300%, while at the same methanol concentration ITP hydrolysis was inhibited approximately 50%. In the presence of 20% methanol, ATP hydrolysis exhibited linear plots of 1/[ATP] vs. 1/v, while in the absence of methanol negative cooperativity was observed. These data can be interpreted to imply that the catalytic and regulatory sites of the mitochondrial ATPase are being dissociated 20% methanol. The effect of methanol on the hydrolysis of ATP and ITP was examined as a function of pH. It was found that, at high pH in totally aqueous solutions, the hydrolysis of ATP and ITP was inhibited, while the presence of 20% methanol either caused the hydrolytic rate to peak and remain constant above pH 8 (with ATP as substrate) or caused the rate of hydrolysis to continue to increase above pH 8 (when ITP was the substrate). These data are interpreted to indicate that an acidic group in the active site may be ionizing, limiting the ATPase-catalyzed hydrolytic rate, and, with 20% methanol, this ionization was inhibited.
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PMID:Effect of organic solvents on the beef heart mitochondrial adenosine triphosphatase. 15 24

1. F1-ATPase has been extracted by the diphosphatidylglycerol procedure from mitochondrial ATPase complexes that differ in ATPase activity, cold stability, ATPase inhibitor and magnesium content. 2. The ATPase activity of the isolated enzymes was dependent upon the activity of the original particles. In this respect, F1-ATPase extracted from submitochondrial particles prepared in ammonia (pH 9.2) and filtered through Sephadex G-50 was comparable to the enzyme purified by conventional procedures (Horstman, L.L. and Racker, E. (1970) J. Biol. Chem. 245, 1336--1344), whereas F1-ATPase extracted from submitochondrial particles prepared in the presence of magnesium and ATP at neutral pH was similar to factor A (Andreoli, T.E., Lam, K.W. and Sanadi, D.R. (1965) J. Biol. Chem. 240, 2644--2653). 3. No systematic relationship has been found in these F1-ATPase preparations between their ATPase inhibitor content and ATPase activity. Rather, a relationship has been observed between this activity and the efficiency of the ATPase inhibitor-F1-ATPase association within the membrane. 4. It is concluded that the ATPase activity of isolated F1-ATPase reflects the properties of original ATPase complex provided a rapid and not denaturing procedure of isolation is employed.
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PMID:F1-ATPase from different submitochondrial particles. 15 27

The effects of a photoaffinity derivate of ATP, arylazido-beta-alanyl-ATP, 3'-O-(3-[N-(4-azido-2-nitrophenyl)amino]propionyl) adenosine 5'-triphosphate, on submitochondrial particles and the partially purified ATPase complex of beef heart mitochondria have been investigated. In the absence of light the ATP analogue has been found to be a substrate for the E132PA1P1-ATP exchange reaction of submitochondrial particles. When photoirradiated in the presence of arylazido-beta-alanyl-ATP, the ATPase activity and the the the [32P]Pi-ATP exchange reaction are inhibited maximally 80%. Arylazido-beta-alanyl-ATP following photolysis is a noncompetitive inhibitor with respect to ATP while arylazido-beta-alanine, the azido-containing adjunct of the ATP analogue, has no inhibitory effect under the same conditions. The inactivating effect of arylazido-beta-alanyl-ATP is prevented in part by the presence of ATP, or ADP and pyrophosphate. Photolabeling produces a covalent binding of the derivative with the F1ATPase being the major protein labeled. The binding of 0.22 mumol of arylazido-beta-alanyl-ATP/mg of mitochondrial protein is associated with a maximal inhibitory effect. The ATPase activity of the partially purified ATPase complex is also sensitive to photoirradiation in the presence of arylazido-beta-alanyl-ATP. When the ATPase complex is associated with liposomes there is an increase in the specific ATPase activity with a 10-fold increase in Vmax and a 4-fold decrease in KmATP associated with a parallel increase in the apparent affinity and maximal inhibitory effect of the arylazido-beta-alanyl-ATP. The photoinhibition of the ATPase complex in the presence of arylazido-beta-alanyl-ATP results in covalent binding of 1.6 mumol of arylazido-beta-alanyl-ATP/mg of protein. The alpha and beta subunits are the only components of the ATPase complex labeled by the [3H]arylazido-beta-alanyl-ATP. The relationship between the arylazido-beta-alanyl-ATP-labeled sites and the nucleotide binding sites on the mitochondrial ATPase is discussed.
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PMID:The use of arylazido-beta-alanyl-ATP as a photoaffinity label for the isolated and membrane-bound mitochondrial ATPase complex. 15 61

Mixed anhydrides of nucleoside triphosphates and mesitylenecarboxylic acid inhibit soluble mitochondrial ATPase (adenosine triphosphatase), but do not inhibit ATPase of submitochondrial particles. Inhibition of soluble mitochondrial ATPase by the mixed anhydride of epsilon-ATP and mesitylenecarboxylic acid is followed by the covalent binding of one nucleotide residue to a molecule of the protein. It is suggested that this covalent binding occurs in the catalytic site of the mitochondrial ATPase. The mixed anhydride of ADP and mesitylenecarboxylic acid inhibits the ATPase activity of submitochondrial particles and has no effect on the activity of soluble mitochondrial ATPase. After separation of the submitochondrial particles from the mixed anhydride of ADP and mesitylenecarboxylic acid, their ATPase activity is restored to its original value (half-time of reactivation 3--4 min). Incubation of submitochondrial particles or soluble mitochondrial ATPase with the mixed anhydride of ADP and mesitylenecarboxylic acid results in AMP formation.
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PMID:Mixed anhydrides of nucleotides and mesitylenecarboxylic acid as new specific inhibitors of mitochondrial adenosien triphosphatase. 15 22

We investigated the kinetics of mitochondrial ATPase in bovine heart mitochondria and submitochondrial particles upon treatment with phospholipase A2, or upon addition of n-butanol to perturb the lipid protein interactions. The changes observed are the following: (1) Lipid removal or perturbation with butanol is accompanied by loss of ATPase activity with decrease of both V and of the KM for ATP. (2) There are changes of activation energy of ATPase activity at temperatures above the discontinuity normally observed for membrane-bound enzymes in mitochondria. In particular, butanol abolishes the discontinuity, and induces a constant activation energy of about 32 kcal/mol in the range 8--37 degrees C. (3) Butanol modifies the pH dependence of ATPase shifting the pH optimum from around 10 to less alkaline values. The optimum for Mg2+ concentrations is increased by the solvent. (4) Treatment with phospholipase A2 results in a removal of oligomycin-sensitive ATPase, whereas butanol addition prevents oligomycin inhibition of ATPase. (5) In beef heart mitochondria, a spin-labelled analog of the inhibitor, dicyclohexyl carbodiimide, did not show any change in environment upon butanol addition, unlike that found in mitochondria from Saccharomyces cerevisiae.
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PMID:Lipid protein interactions in mitochondria. VII. A comparison of the effects of lipid removal and lipid perturbation of the kinetic properties of mitochondrial ATPase. 15 58

The dicyclohexylcarbodiimide-binding protein of Aspergillus nidulans has been identified as the smallest subunit of the mitochondrial ATPase complex, and has a molecular weight of approximately 8000. It is extractable from whole mitochondria and from the purified enzyme in neutral chloroform/methanol, contains 30% polar amino acids, and the N-terminal amino acid has been identified as tyrosine. Using a double-labelling technique in the absence and presence of cycloheximide, followed by immunoprecipitation of the enzyme complex with antiserum against Neuospora crassa F1 ATPase, it has been shown that this subunit is synthesized on cytoplasmic ribosomes.
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PMID:Mitochondrial ATPase complex of Aspergillus nidulans and the dicyclohexylcarbodiimide-binding protein. 15 78


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