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 electrokinetic model was developed to calculate the time course of electrical parameters, ion fluxes, and intracellular ion activities for experiments performed in airway epithelial cells. Model variables included cell [Na], [K], [Cl], volume, and membrane potentials. The model contained apical membrane Cl, Na, and K conductances, basolateral membrane K conductance, Na/K/2 Cl and Na/Cl symport, and 3 Na/2 K ATPase, and a paracellular conductance. Transporter permeabilities and ion saturabilities were determined from reported ion flux data and membrane potentials in intact canine trachea. Without additional assumptions, the model predicted accurately the measured short-circuit current (Isc), cellular conductances, voltage-divider ratios, open-circuit potentials, and the time course of cell ion composition in ion substitution experiments. The model was used to examine quantitatively: (a) the effect of transport inhibitors on Isc and membrane potentials, (b) the dual role of apical Cl and basolateral K conductance in cell secretion, (c) whether the basolateral symporter requires K, and (d) the regulation of apical Cl conductance by cAMP and Ca-dependent signaling pathways. Model predictions gave improved understanding of the interrelations among transporting systems and in many cases gave surprising predictions that were not obvious without a detailed model. The model developed here has direct application to secretory or absorptive epithelial cells in the kidney thick ascending limb, cornea, sweat duct, and intestine in normal and pathophysiological states such as cystic fibrosis and cholera.
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PMID:Model of ion transport regulation in chloride-secreting airway epithelial cells. Integrated description of electrical, chemical, and fluorescence measurements. 169 71

A method is presented for the micro-scale isolation and characterization of erythrocyte membrane Ca(2+)-ATPase from small samples (7 mL) of whole human blood. Ca(2+)-ATPase isolated by this technique was more than 92% pure and showed calcium-activation characteristics similar to enzyme purified by standard macroscale procedures--viz maximal velocity of activation (VCA2+) = 15.5 +/- 1.2 mumol ATP hydrolysed/mg/min, and reciprocal of apparent affinity (KCa2+) = 0.73 +/- 0.15 microM free calcium (mean +/- SEM; n = 9). Using the isolation procedure described, purified Ca(2+)-ATPase could be prepared and assayed in a single working day. When the calcium-activation kinetics of cystic fibrosis erythrocyte membrane Ca(2+)-ATPase were reassessed using enzyme purified by this technique, VCa2+ and KCa2+ were not significantly different from normal values.
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PMID:Purification and analysis of erythrocyte membrane Ca(2+)-ATPase from small samples of patient blood: application to cystic fibrosis. 183 18

Cystic fibrosis (CF) airway epithelia exhibit raised transepithelial Na+ transport rates, as determined by open-circuit isotope fluxes and estimates of the amiloride-sensitive equivalent short-circuit current (Ieq). To study the contribution of apical and basolateral membrane paths to raised Na+ transport in CF, CF nasal epithelial cultures were studied with double-barreled Na(+)-selective microelectrodes and the Ussing chamber technique. Intracellular Na+ activity (acNa) was 24.1 +/- 1.5 mM (n = 36), a value similar to acNa of normal nasal epithelial cells. Reduction of luminal [Na+] to 3 mM abolished Ieq and reduced acNa. Amiloride (10(-4) M) abolished Ieq but increased acNa from 20 +/- 2 to 36 +/- 7 mM (n = 10). Amiloride-induced increase in acNa was not affected by serosal [Na+] reduction but was blocked by preexposure to reduced luminal [Na+]. Amphotericin B increased Ieq during amiloride exposure, indicating that amiloride did not inhibit NA(+)-K(+)-ATPase. Ouabain abolished Ieq and slowly raised acNa. Reduction of serosal [Na+] led to a decrease in acNa that was blocked by bumetanide. It is concluded that 1) CF airway epithelia exhibit an increased apical membrane Na+ permeability, 2) acNa is regulated to a normal level in CF cells despite increased transcellular Na+ fluxes, 3) the abnormal increase in acNa in response to amiloride is dependent on luminal Na+, 4) Na+ is transported across the basolateral membrane by a bumetanide-sensitive cotransport mechanism, and 5) ouabain inhibits the basolateral Na(+)-K(+)-ATPase, causing slow dissipation of the chemical and electrical gradients across the cell membranes.
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PMID:Transcellular sodium transport in cultured cystic fibrosis human nasal epithelium. 187 75

The mucosa that lines the airways is covered with a fluid film forming a hypophase between mucus and cell surface. To study the function of this epithelium aims at describing the mechanisms by which fluid is normally produced. Another goal to be pursued consists in looking for the origin of pathological situations, such as cystic fibrosis, in which the functioning of epithelial cell is altered. The elucidation of transport mechanisms present in the apical and in the basolateral membrane results in a conceptual model that illustrates the asymmetrical functioning of epithelial cells. Recent discoveries enlarge our understanding of membrane transport processes; in particular, a concerted, reciprocal regulation of the activity of both membranes was shown to be exerted via the intracellular composition. The tracheal epithelium absorbs Na+ and secretes Cl-. These two transports are active and electrogenic; their sum corresponds approximately to the short-circuit current measured in vitro. Na+ absorption is sensitive to amiloride from the luminal side and also to ouabain added to the serosal compartment. The process is a primary active transport, analogous to that found in amphibian epithelia or in mammalian colon. Cl- secretion is abolished by furosemide (or bumetanide), by ouabain or by Na+ suppression in the serosal incubation solution. The mechanism is a secondary active transport: Cl- influx across the basolateral membrane is coupled to Na+ (probably through Na+, K+, Cl- symport); energy is dissipated by the Na+-K+-ATPase localised in the basolateral membrane. Thus, Na+ is recirculated across that membrane by the pump activity, which maintains a favorable gradient for influx via the symport. Cl- efflux takes place by diffusion through the luminal membrane. This model applies to other epithelia in which Na+-coupled Cl- secretion was shown to take place. It is confirmed by isotopic fluxes measurements and by electrophysiologic properties of the apical and the basolateral membrane. Various agents are known to influence ion transports. In particular Cl- secretion is stimulated by substances that increase the intracellular concentration of cyclic AMP. At the membrane level, the number of active Cl- channels in the apical membrane is primarily controlled, then the basolateral membrane K+ permeability. Yet, species differences are worth to note: the trachea of the cow is barely sensitive to agents that exert a marked action on dog trachea. The tracheal epithelium is used as an experimental model for studying cystic fibrosis, a disease in which the apical membrane is almost devoid of functional Cl- channels, so that Cl- permeability is quite low.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:[Physiology of the tracheal epithelium]. 246 15

Phosphoinositide content was measured in erythrocyte membranes from 11 patients with cystic fibrosis (CF) and from 12 control subjects to determine whether altered levels of phosphatidylinositol-4-phosphate (Ptdlns4P) or phosphatidylinositol-4,5-bisphosphate (Ptdlns(4,5)P2) are responsible for the decrease in Ca2+-adenosine triphosphatase (Ca2+-ATPase) activity in this disorder. Isolated membranes were extracted with an acidified chloroform-methanol solvent system. The recovered lipids were separated by one-dimensional thin-layer chromatography and quantified with a colorimetric assay for phosphorus. The results are expressed in molar percent, moles of phosphoinositide times 100 divided by the total number of moles of phospholipid per membrane. The means +/- SEM of Ptdlns(4,5)P2, Ptdlns4P, and phosphatidylinositol (Ptdlns) in CF membranes (1.07 +/- 0.18, 1.02 +/- 0.22, and 2.32 +/- 0.36 molar percent, respectively) were indistinguishable from controls (0.91 +/- 0.14, 0.85 +/- 0.12, and 2.21 +/- 0.32 molar percent, respectively) (P greater than 0.20 for all three pairs). The accuracy of quantitative recovery throughout the procedure was determined by adding a radioactive internal standard, L-3-phosphatidyl[2-3H]inositol to 10 membrane preparations. Although quantitative recoveries, as determined by percent radioactivity recovered, varied from 54% to 92%, mean Ptdlns(4,5)P2, Ptdlns4P, and Ptdlns levels appropriately corrected from tracer loss were still indistinguishable between the two groups. We conclude that absolute phosphoinositide levels are not altered in cystic fibrosis erythrocyte membranes and that the differences in Ca2+-ATPase activity cannot be explained on this basis.
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PMID:Phosphoinositide content of erythrocyte membranes in cystic fibrosis. 283 Mar 55

The reduction in (Ca2+ + Mg2+)-ATPase activity in the cystic fibrosis red blood cells can be attributed to a reduction in the number of active Ca2+ pumps per red blood cell and an altered interaction of calcium ions with the pump. Despite this, the normal free intracellular [Ca2+] is preserved due to a lower rate of passive calcium entry.
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PMID:The nature of the Ca2+-pump defect in the red blood cells of patients with cystic fibrosis. 293 Nov 15

There are several conflicting reports regarding a defective (Ca2+ + Mg2+)-ATPase in tissues from cystic fibrosis (CF) patients. The work presented in this paper represents an independent assessment of (Ca2+ + Mg2+)-ATPase function in CF and, at the same time, provides a somewhat more detailed analysis of the enzyme from CF patients than has previously existed. We found no significant differences in either the Km value for Ca2+ or the Vm value of the (Ca2+ + Mg2+)-ATPase in membrane preparations from CF patients and control subjects when the red cell membranes were prepared by methods which utilize Tris-glycylglycine-Mg2+ buffers. In contrast, the Vm value of the (Ca2+ + Mg2+)-ATPase in the preparations from CF patients was found to be lower than that from control subjects when the membranes were prepared by a series of washes with EDTA-containing buffers (i.e., 1-10 mmol/1 EDTA). The EDTA treatment, however, did not produce any significant difference in the Km between the two groups. The fluorescent ATP analogue, trinitrocyclohexadienylidine-ATP, appeared to interact with erythrocyte ghosts as evidenced from an enhancement of its fluorescence. This enhancement was greater in control preparations than in samples from CF patients. In addition, the kinetic profiles, with respect to ATP, were quite different between the two enzyme populations. The overall results suggest that the lower rate of ATP hydrolysis observed with membranes from CF patients may reflect an impaired utilization of the substrate by the (Ca2+ + Mg2+)-ATPase from these individuals.
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PMID:Probe of the (Ca2+ + Mg2+)-ATPase in erythrocyte membranes of cystic fibrosis patients. 294 33

The (Ca2+ + Mg2+)-ATPase present per mg of protein in erythrocyte membranes of controls and patients with cystic fibrosis (CF) was determined by estimation of the levels of its phosphoprotein. In the presence of 10 mM free Ca2+, which inhibits phosphoprotein decomposition, significantly less phosphoprotein intermediate, ECaP, was found in erythrocyte membranes from CF patients than in age- and sex-matched controls; this correlated with a significant decrease in (Ca2+ + Mg2+)-ATPase activity. These observations indicate a decrease in the number of functional (Ca2+ + Mg2+)-ATPase molecules in erythrocyte membranes from CF patients or an alteration in either the structure of the pump protein or the composition of its environment.
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PMID:Investigation of (Ca2+ + Mg2+)-ATPase phosphoprotein formation in erythrocyte membranes of patients with cystic fibrosis. 294 Nov 49

Conditioned culture media taken from fibroblast cell lines derived from skin biopsies of control or of patients with Cystic Fibrosis (CF) were incubated with membranes of rat submandibular glands. The Na/K - ATPase activity of these membranes was inhibited when treated with CF-media, including both ouabain sensitive and insensitive activities. However, the membrane associated Mg-ATPase, Ca-ATPase, and both basal and hormone-stimulated adenylate cyclase activities were relatively unaffected. Thus, a factor or factors produced by CF-fibroblasts was shown to be active in a cell-free system derived from an exocrine gland.
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PMID:Conditioned media from cultured cystic fibrosis fibroblasts inhibits Na/K ATPase activity. 300 62

The properties of the Ca2+, Mg2+-ATPase of erythrocyte membranes from patients with cystic fibrosis (CF) were extensively compared to that of healthy controls. Following removal of an endogenous membrane inhibitor of the ATPase, activation of the enzyme by Ca2+, calmodulin, limited tryptic digestion or oleic acid, as well as inhibition by trifluoperazine, were studied. The only properties found to be significantly different (CF cells vs controls) were calmodulin-stimulated peak activity (90 vs 101, P less than 0.02) and trypsin-activated peak activity (92 vs 102, P less than 0.02). No significant difference could be measured in the steady-state Ca2+-dependent phosphorylation of CF and control erythrocyte membranes indicating similar numbers of enzyme molecules per cell. The functional state of Ca2+ homeostasis in intact erythrocytes was investigated by measuring the resting cytosolic free Ca2+ levels using quin-2. Both CF and control erythrocytes maintained cytosolic free Ca2+ between 20 to 30 nM. Addition of 50 uM trifluoperazine resulted in an increase in erythrocyte cytosolic free Ca2+ to about 50 nM in both CF and control cells. Estimates of erythrocyte membrane permeability using the steady-state uptake of 45Ca into intact erythrocytes revealed no differences between CF and control cells. These results confirm that there is a small decrease in the calmodulin-stimulated activity of the erythrocyte Ca2+, Mg2+-ATPase in CF. However, this deficit is apparently not large enough to impair the ability of the CF erythrocyte to maintain normal resting levels of cytosolic free Ca2+.
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PMID:Erythrocyte cytosolic free Ca2+ and plasma membrane Ca2+-ATPase activity in cystic fibrosis. 316 Apr 70


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