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)

Urea, in nondenaturing concentrations, inhibited Ca2+ uptake by sarcoplasmic reticulum vesicles with no concomitant effect on ATP hydrolysis. This inhibition was antagonized by 5 mM oxalate and 20 mM orthophosphate. At concentrations of 0.2 to 1.0 M, urea induced an increase in the Ca2+ efflux from preloaded vesicles diluted in a medium at pH 7.0 containing 2 mM ethylene glycol bis(beta-aminoethyl ether)N,N'-tetraacetic acid, 0.1 mM orthophosphate, and 0.1 mM MgCl2. The urea-induced efflux was arrested by ligands of the (Ca(2+)-Mg2+) ATPase, namely, K+, Mg2+, Ca2+, and ADP, and by ruthenium red and the polyamines spermine, spermidine, and putrescine. In the case of polyamines a dissociation between the effect on the efflux and the net Ca2+ uptake was observed, as only the efflux could be blocked by the drugs. Glycine betaine, trimethylamine-N-oxide, and sucrose antagonized the effects of urea on both the net Ca2+ uptake and the rate of Ca2+ efflux.
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PMID:The enhancement of Ca2+ efflux from sarcoplasmic reticulum vesicles by urea. 128 64

Electron microprobe analysis on freeze-dried cryosections was used to determine the effect of the loop diuretics torasemide and furosemide on intracellular electrolyte concentrations in individual cells of the outer and inner stripe of the outer medulla and on cell rubidium uptake, the latter a measure of basolateral Na-K-ATPase activity. In addition, the organic osmolytes glycerophosphorylcholine (GPC), betaine, inositol and sorbitol in cortex, outer medulla and inner medulla were measured using HPLC. Both loop diuretics significantly reduced sodium and chloride concentrations and rubidium uptake in thick ascending limb cells, but did not affect sodium concentration or rubidium uptake in the proximal straight tubule (PST) cells or in the light or dark cells of the outer medullary collecting duct (OMCD). Chloride concentrations in these cells (that is, PST cells, OMCD light and dark cells) were lowered by loop diuretics, albeit less than in thick ascending limb cells. Administration of both loop diuretics for only 20 minutes was sufficient to significantly depress tissue concentrations of GPC, betaine, and myo-inositol in the outer medulla and of GPC, betaine and sorbitol at the papillary tip. These results indicate that loop diuretics, presumably by blocking apical sodium entry, decrease thick ascending limb cellular sodium concentration and, as a consequence, reduce Na-K-ATPase activity as assessed by cell rubidium uptake. Although this has been shown previously in in vitro preparations, the present study confirms this for the first time in vivo.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Effect of loop diuretics on organic osmolytes and cell electrolytes in the renal outer medulla. 145 80

The resting potential of identified cells (Parker cells) in the abdominal ganglion of Elysia chlorotica (Gould) depolarizes by about 30 mV in response to a 50% reduction in osmolality and returns to the original potential in 20 min. Cell volume recovery requires approximately 2 h. Thus, recovery of the resting potential is not dependent on recovery of cell volume. The hypo-osmotic depolarization persists following inhibition of the electrogenic Na+/K(+)-ATPase with ouabain, and the levels of extracellular K+ and Cl- have little effect on the magnitude of the depolarization, while decreasing extracellular Na+ concentration produces a depolarization of only 10 mV. This suggests that the hypo-osmotic depolarization in Parker cells results mostly from increased relative permeability to Na+. Following transfer from 920 to 460 mosmol kg-1, Na+, Cl- and proline betaine leave the cells while intracellular K+ is conserved. Loss of intracellular Na+ and conservation of intracellular K+ are dependent on active transport by the Na+/K(+)-ATPase. Na+ and proline betaine leave the cells with a time course that is much longer than that of the hypo-osmotic depolarization. Unlike the other solutes, most of the reduction in intracellular Cl- concentration occurs coincidentally with the hypo-osmotic depolarization. However, unlike the hypo-osmotic depolarization, bulk loss of Cl- does not require the reduction in osmolality, only the reduction in extracellular ion concentrations. There is no apparent relationship between membrane depolarization and the regulation of intracellular osmolytes in Elysia neurons following hypo-osmotic stress.
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PMID:The ionic basis of the hypo-osmotic depolarization in neurons from the opisthobranch mollusc Elysia chlorotica. 155 12

Proline porter II is rapidly activated when nongrowing bacteria are subjected to a hyperosmotic shift (Grothe, S., Krogsrud, R. L., McClellan, D. J., Milner, J. L., and Wood, J. M. (1986) J. Bacteriol. 166, 253-259). Proline porter II was active in membrane vesicles prepared from bacteria grown under optimal conditions, nutritional stress, or osmotic stress. That activity was: (i) dependent on the presence of the energy sources phenazine methosulphate plus ascorbate or D-lactate; (ii) observed only when a hyperosmotic shift accompanied the transport measurement; (iii) inhibited by glycine betaine in a manner analogous to that observed in whole cells; and (iv) eliminated by lesions in proP. Membrane vesicles were able to transport serine but not glutamine and serine transport was reduced by the hyperosmotic shift. In whole cells, proline porter II activity was supported by glucose and by D-lactate in a strain defective for proline porters I and III and the F1F0-ATPase. Glucose energized proline uptake was eliminated by carbonyl cyanide m-chlorophenylhydrazone and KCN as was serine uptake. These results suggested that proline porter II was respiration-dependent and probably ion-linked. Activation of proline porter II in whole cells by sucrose or NaCl was sustained over 30 min, whereas activation by glycerol was transient. Proline porter II was activated by NaCl and sucrose with a half-time of approximately 1 min in both whole cells and membrane vesicles. Thus, activation of proline porter II was reversible. It occurred at a rate comparable to that of K+ influx and much more rapid than the genetic regulatory responses that follow a hyperosmotic shift.
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PMID:Proline porter II is activated by a hyperosmotic shift in both whole cells and membrane vesicles of Escherichia coli K12. 304 95

Cycocel stimulated the activity of partial purified choline kinase from spinach or squash leaves, but it inhibited the activity of yeast choline kinase. The activity of different Cycocel analogs on plant growth corresponded to their stimulatory effect on the isolated choline kinase. Cycocel had no effect upon the activity of a plant phosphatase which hydrolyzed phosphorylcholine nor upon adenosine triphosphatase from wheat roots or leaves. Gibberellin A(3) inhibited choline kinase activity and reversed the stimulatory effect of Cycocel on the kinase. Total choline kinase activity per squash plant was not greatly increased by Cycocel treatment. However, on the basis of fresh weight, total kinase activity was increased by Cycocel treatment. Gibberellin A(3) partially reversed these increases. Treatment with Cycocel plus indoleacetic acid resulted in a large increase in choline kinase activity. The same distribution of tracer among phosphorylcholine, choline and betaine was observed when either phosphorylcholine-C(14) or choline-C(14) was fed to barley or wheat roots. Cycocel stimulated the incorporation of choline-C(14) into the insoluble fraction and into lipids. Cycocel inhibited phosphorylcholine uptake by roots. Thus Cycocel stimulated choline kinase activity and the utilization of choline-C(14). The effect of Cycocel upon kinase activity in vivo and in vitro was reversed by gibberellin A(3).
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PMID:Effect of Cycocel derivatives and gibberellin on choline kinase and choline metabolism. 590 35

A method has been developed for calculating rate constants for dehydration of aldehydes that induce ATPase reactions by kinases and where 18O is transferred from the aldehyde or its hydrate to inorganic phosphate during the reaction. The method involves measurement of the fraction of 18O in phosphate by 31P NMR after the ATPase reaction has proceeded for several minutes with zero-order kinetics. The reaction is started by addition of the aldehyde in a small volume of H2 18O, and the speed of washout of 18O by reversible dehydration relative to the rate of the ATPase reaction allows calculation of the rate constants if the hydration equilibrium constant is known from the proton NMR spectrum of the aldehyde. Dehydration rate constants (s-1 at pH 8-8.5, 0.1 M buffer, 25 degrees C) for the following aldehydes (all over 95% hydrated) and kinases used are as follows: D-glyceraldehyde with glycerokinase, 0.03; 2,5-anhydro-D-mannose 6-phosphate with fructose-6-phosphate kinase, 0.025; 2,5-anhydro-D-mannose or 2,5-anhydro-D-talose with fructokinase, 0.029 and 0.017, respectively; D-gluco-hexodialdose with hexokinase, 0.068. With betaine aldehyde and choline kinase or glyoxylate and pyruvate kinase, no 18O was transferred to phosphate during the ATPase reactions. However, the dehydration rate constant for glyoxylate (0.007 s-1 at pH 7 extrapolated to zero buffer concentration and up to 0.11 s-1 at pH 9.0 with 0.3 M buffer) was determined by extrapolating the initial rate of reduction of the free aldehyde catalyzed by lactate dehydrogenase to infinite enzyme levels.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:A novel method for determining rate constants for dehydration of aldehyde hydrates. 609 90

Aldehyde analogues of the normal alcohol substrates induce ATPase activities by glycerokinase (D-glyceraldehyde), fructose-6-phosphate kinase (2,5-anhydromannose 6-phosphate), fructokinase (2,5-anhydromannose or 2,5-anhydrotalose), hexokinase (D-gluco-hexodialdose), choline kinase (betaine aldehyde), and pyruvate kinase (glyoxylate). Since purified deuterated aldehydes give V and V/K isotope effects near 1.0 for glycerokinase, fructokinase with 2,5-anhydro[1-2H]talose, hexokinase, choline kinase, and pyruvate kinase, the hydrates of these almost fully hydrated aldehydes are the activators of the ATPase reactions. Fructose-6-phosphate kinase and fructokinase with 2,5-anhydro[1-2H]mannose show V/K deuterium isotope effects of 1.10 and 1.22, respectively, suggesting either that both hydrate and free aldehyde may be activators (predicted values are 1.37 if only the free aldehyde activates the ATPase) or, more likely, that the phosphorylated hydrate breaks down in a rate-limiting step on the enzyme while MgADP is still present and the back-reaction to yield free hydrate in solution is still possible. 18O was transferred from the aldehyde hydrate to phosphate during the ATPase reactions of glycerokinase, fructose-6-phosphate kinase, fructokinase, and hexokinase but not with choline kinase or pyruvate kinase. Thus, direct phosphorylation of the hydrates by the first four enzymes gives the phosphate adduct of the aldehyde, which decomposes nonenzymatically, while with choline kinase and pyruvate kinase the hydrates induce transfer to water (metal-bound hydroxide or water with pyruvate kinase on the basis of pH profiles). Observation of a lag in the release of phosphate from the glycerokinase ATPase reaction at 15 degrees C supports the existence of a phosphorylated hydrate intermediate with a rate constant for breakdown of 0.035-0.043 s-1 at this temperature. Kinases that phosphorylate creatine, 3-phosphoglycerate, and acetate did not exhibit ATPase activities in the presence of keto or aldehyde analogues (N-methylhydantoic acid, D-glyceraldehyde 3-phosphate, and acetaldehyde, respectively), possibly because of the absence of an acid-base catalytic group in the latter two cases. These analogues were competitive inhibitors vs. the normal substrates, and in the latter case, the hydrate of acetaldehyde was shown to be the inhibitory species on the basis of the deuterium isotope effect on the inhibition constant.
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PMID:Mechanisms of aldehyde-induced adenosinetriphosphatase activities of kinases. 609 91

We have investigated the effects of the protein structure-perturbing and function-perturbing osmolyte urea, and one of its physiological counteracting solutes, the methylamine compound (carboxymethyl)trimethylammonium hydroxide (betaine), on the structure and function of the human erythrocyte plasma-membrane Ca(2+)-ATPase. Betaine per se promoted a conformational change in the purified ATPase as revealed by steady-state and time-resolved intrinsic fluorescence spectroscopy. The conformational change promoted by betaine was shown to be related to changes in the degree of compaction of the protein structure, as detected by fluorescence-quenching measurements using acrylamide and iodide, non-charged and charged quenchers, respectively. In contrast, urea promoted a biphasic increase in exposure of tryptophan residues of the purified ATPase to the aqueous medium. With the use of membrane-bound ATPase, increasing concentrations of urea up to 1.5 M promoted a twofold increase in the Ca(2+)-ATPase activity, and the simultaneous inhibition of Ca2+ accumulation indicated that ATP hydrolysis became uncoupled from Ca2+ transport. Higher urea concentrations promoted a pronounced inhibition of ATP hydrolysis. In the absence of urea, betaine decreased ATP hydrolysis without affecting Ca2+ transport, whereas it counteracted the strong inhibition of Ca(2+)-ATPase activity by urea concentrations as high as 7 M. Betaine also protected Ca2+ accumulation against inhibition with concentrations of urea up to 1.5 M, indicating that the methylamine is able to counteract the uncoupling of the ATPase observed at lower urea concentrations. These results suggest that betaine modifies the effects of urea on the erythrocyte Ca(2+)-ATPase, through specific solute-induced conformational changes that protect the energy-transduction capacity of the enzyme.
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PMID:Betaine counteracts urea-induced conformational changes and uncoupling of the human erythrocyte Ca2+ pump. 818 68

The OpuA transport system of Bacillus subtilis functions as a high-affinity uptake system for the osmoprotectant glycine betaine. It is a member of the ABC transporter superfamily and consists of an ATPase (OpuAA), an integral membrane protein (OpuAB), and a hydrophilic polypeptide (OpuAC) that shows the signature sequence of lipoproteins (B. Kempf and E. Bremer, J. Biol. Chem. 270:16701-16713, 1995). The OpuAC protein might thus serve as an extracellular substrate binding protein anchored in the cytoplasmic membrane via a lipid modification at an amino-terminal cysteine residue. A malE-opuAC hybrid gene was constructed and used to purify a lipidless OpuAC protein. The purified protein bound radiolabeled glycine betaine avidly and exhibited a KD of 6 microM for this ligand, demonstrating that OpuAC indeed functions as the substrate binding protein for the B. subtilis OpuA system. We have selectively expressed the opuAC gene under T7 phi10 control in Escherichia coli and have demonstrated through its metabolic labeling with [3H]palmitic acid that OpuAC is a lipoprotein. A mutant expressing an OpuAC protein in which the amino-terminal cysteine residue was changed to an alanine (OpuAC-3) was constructed by oligonucleotide site-directed mutagenesis. The OpuAC-3 protein was not acylated by [3H]palmitic acid, and part of it was secreted into the periplasmic space of E. coli, where it could be released from the cells by cold osmotic shock. The opuAC-3 mutation was recombined into an otherwise wild-type opuA operon in the chromosome of B. subtilis. Unexpectedly, this mutant OpuAC system still functioned efficiently for glycine betaine acquisition in vivo under high-osmolarity growth conditions. In addition, the mutant OpuA transporter exhibited kinetic parameters similar to that of the wild-type system. Our data suggest that the lipidless OpuAC-3 protein is held in the cytoplasmic membrane of B. subtilis via its uncleaved hydrophobic signal peptide.
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PMID:Lipoprotein from the osmoregulated ABC transport system OpuA of Bacillus subtilis: purification of the glycine betaine binding protein and characterization of a functional lipidless mutant. 933 65

The ability of Streptococcus mutans, a bacterial pathogen associated with dental caries, to tolerate rapid drops in plaque pH (acidurance), is considered an important virulence factor. To study this trait, Tn917 mutants of S. mutans strain JH1005 which display acid sensitivity have been isolated and partially characterized. In this paper, the characterization of one of these mutants, AS17, is reported. Preliminary sequence analysis revealed that the transposon insertion in AS17 occurred in the intergenic region of a two-gene locus which has been named sat for secretion and acid tolerance. This locus displays a high degree of homology to the ylxM-ffh operon of Bacillus subtilis. The sat+ locus was cloned by complementation of a conditional Escherichia coli ffh mutant with an S. mutans genomic library. Sequencing of the complementing clone identified the intact ylxM and ffh genes as well as a partial ORF with homology to the proUlopuAC gene of B. subtilis which encodes the binding protein of the ProU/OpuA osmoregulated glycine betaine transport system. RNA dot blot experiments indicated steady-state levels of ffh mRNA in the mutant that were approximately eightfold lower compared to parental levels. This suggests a partial polar effect of the sat-1::Tn917 mutation on ffh expression. Upon acid shock (pH 5), wild-type ffh mRNA levels were found to increase approximately four- to eightfold compared to unstressed (pH 7.5) levels. Mutant levels remained unaltered under the same conditions. Experiments designed to investigate the origins of the acid-sensitivity of the mutant revealed a lack of an acid-adaptive/tolerance response. Assays of proton-extruding ATPase (H+/ATPase) specific activity measured with purified membranes derived from acid-shocked AS17 showed twofold lower levels compared to the parent strain. Also, AS17 was found to be unable to ferment sorbitol although it was able to grow in glucose and a variety of other sugar substrates. These findings suggest that Ffh may be involved in the maintenance of a functional membrane protein composition during adaptation of S. mutans to changing environmental conditions.
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PMID:Streptococcus mutans ffh, a gene encoding a homologue of the 54 kDa subunit of the signal recognition particle, is involved in resistance to acid stress. 1007 18


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