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)

Muscle actin is, in most cases, prepared from an acetone-dried powder of the myosin-removed myofibrils under low-salt conditions in the presence of ATP. In this paper, it is shown that G-actin can be directly extracted from the myosin-removed myofibrils without acetone treatment. The extraction conditions are the same as those used for the extraction of G-actin from the dried powder: extraction of the myosin-removed myofibrils for 1 h with 2 mM Tris-HCl, pH 8.0, in the presence of 0.5 mM ATP. However, the crude G-actin directly extracted from the myosin-removed myofibrils loses its polymerizability after prolonged extraction. Measurements of inorganic phosphate and thin layer chromatography of the adenine nucleotides of the crude G-actin solution show that free ATP added to the extraction buffer is sequentially hydrolyzed to ADP and AMP, and then finally converted to IMP. The instability of the G-(ADP)-actin, depolymerized from the ends of actin filaments, explains the loss in polymerizability of G-actin during the extraction. Residual ATPase, adenylate kinase, and deaminase contained in the myofibrils may account for the decomposition of ATP.
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PMID:Direct extraction of G-actin from the myosin-removed myofibrils under the conditions of low ionic strength. 716 Dec 61

Endocytic vesicles are acidified by an electrogenic proton pump and a parallel chloride conductance; however, acidification might be decreased if electrogenic transporters, such as Na,K-ATPase, that increase vesicle interior-positive membrane potential were also present. We examined this issue in early rat liver endosomes using ion substitution and inhibitors to alter Na,K-ATPase activity. These early endosomes, labeled for 2 min with the fluorescent fluid-phase marker fluorescein isothiocyanate-dextran, consistently acidified faster than endosomes similarly labeled for a 10-min period. In chloride-free media initial rates of acidification of early endosomes were faster in K+ media than in Na+ medium, although addition of K+ to Na+ or Na+ to K+ media to allow Na,K-ATPase to function did not decrease the rate of acidification. In chloride-containing media, rates were the same regardless of cation composition. The Na,K-ATPase inhibitor vanadate was prepared from orthovanadate by several methods, all of which inhibited liver ATPase activity. Two hundred mumol/L vanadate, prepared Cl(-)-free, tended to decrease rates of acidification in all media tested and these effects achieved statistical significance in Cl(-)-free media containing 150 mmol/L K+ or mixtures of Na+ and K+ and in 145 mmol/L KCl/5 mmol/L NaCl medium. Vanadate stocks pH-adjusted with hydrogen chloride increased rates of acidification in sodium gluconate buffers, probably as a result of the effects of the included Cl-. Five mmol/L ouabain (loaded into vesicles by endocytosis) and the membrane-permeable analog strophanthidin (2 mmol/L) both markedly inhibited endosome acidification, regardless of buffer ion composition. Collectively, these results suggest that Na,K-ATPase does not regulate acidification of rat liver early endocytic vesicles, that vanadate may modestly inhibit endosome acidification and that ouabain at high concentrations may inhibit acidification from the vesicle interior face.
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PMID:Role of Na,K-ATPase in regulating acidification of early rat liver endocytic vesicles. 751 Nov 27

Brain pial microvessels have previously been demonstrated to have blood-brain barrier properties. The potential difference (PD) across exposed brain pial microvessels, 20-60 microns in diameter and superfused with artificial CSF, has been measured in anaesthetised rats using glass microelectrodes. The PD on insertion into venous vessels, V(in), was 3.2 mV lumen negative, and in arterial vessels it was higher at 4.5 mV. Superfusion with high K(+)-CSF, made by replacing Na+ with K+, caused a positive deflection in PD, VK+, whereas reducing the Na+ alone, by replacing Na+ by Tris-HCl, made the lumen more negative. These two effects were additive. Studies on venous vessels showed that ouabain had no effect on V(in) and only affected VK+ under conditions of low Na pre-exposure. Neither histamine nor cimetidine had any effect on V(in) or VK+ whereas tetraethylammonium, a K(+)-channel blocker, reduced VK+ by 20%. These experiments demonstrate that changes in PD caused by changing abluminal Na+ or K+ are due predominantly to movement of ions through channels in the endothelial cell membranes, and that actions that alter the activity of the Na+,K(+)-ATPase or reduce the resistance of the paracellular pathway in parallel with increased membrane permeability have less effect on the PD.
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PMID:Transendothelial electrical potential across pial vessels in anaesthetised rats: a study of ion permeability and transport at the blood-brain barrier. 752 22

The transfer of radioactive phosphatidylcholine (PC*) from liposomes to rabbit jejunal and renal brush-border membrane vesicles (BBMVs) was measured with a fast-sampling, rapid-filtration apparatus. PC* uptake by jejunal and renal BBMVs was favoured when liposomes were made from soybean phosphatidylcholine (azolectin, AZO), whereas PC* uptake could not be quantitatively assessed from egg yolk phosphatidylcholine (PC) liposomes even after a 22-h period of incubation. The increased turbidity of BBMV dispersion following the addition of CaCl2 or HCl to AZO-treated BBMVs suggested that negatively charged lipids and phosphatidylethanolamine are transferred during the process. These data and the analysis of PC*-uptake time measurements, using an algorithm simulating aggregation phenomena, indicated that the reaction mechanism involved liposome aggregation to BBMVs rather than specific lipid transfer. The constants of the dimerization reaction between AZO liposomes and BBMVs were evaluated to be 0.016 +/- 0.006 min-1 for jejunal and 0.095 +/- 0.02 min-1 for renal preparations. IntraveSICULAR D-ASPartic acid accumulation in the presence of a NA+ gradient indicated that vesicles were still closed after coincubation with liposomes. In contrast, 70-85% of rabbit jejunal and renal Na(+)-D-glucose cotransporter activities were lost after overnight incubation with either AZO liposomes or buffered solution. Further, H(+)-ATPase activity in rabbit renal BBMVs largely decreased after coincubation with AZO liposomes, while brush-border membrane associated enzymes remained stable. These results demonstrate that coincubation of BBMV with liposomes of different composition may represent a useful approach to study the influence of lipidic environment on various membrane protein functions.
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PMID:Characteristics of exogenous lipid uptake by renal and intestinal brush-border membrane vesicles. 757 91

We examined the effects of YM020 (3-cyanomethyl-2-methyl-8-[(3-methyl-2-butenyl)oxy]-imidazo[1,2- a]pyridine), a novel H+,K(+)-ATPase inhibitor, on gastric acid secretion and experimental gastroduodenal lesions in rats and dogs. Intraduodenal, subcutaneous and oral YM020 inhibited basal gastric acid secretion in pylorus-ligated rats with ED50 values of 9.1, 9.1 and 9.5 mg/kg, respectively. Oral pretreatment with YM020 5 hr before ligation still suppressed acid secretion, with a potency a little less than that of omeprazole. In anesthetized dogs, intravenous YM020 inhibited histamine-, methacholine- and pentagastrin-induced gastric acid secretion with ED50 values of 0.05, 0.01 and 0.08 mg/kg, respectively. In Heidenhain pouch dogs, although oral YM020 (3 mg/kg) inhibited histamine-induced acid secretion, acid output returned to control levels faster than in dogs treated with omeprazole. Oral YM020 inhibited the formation of water-immersion restraint stress-, indomethacin-, absolute ethanol-, 0.7 N hydrochloric acid- and cysteamine-induced gastric or duodenal lesions with ED50 values of 2.9, 4.3, 2.0, 11.7 and 8.4 mg/kg, respectively. Moreover, subcutaneous YM020 also suppressed the formation of ethanol- and HCl-induced gastric lesions. These results suggest that YM020 has an antisecretory effect almost the same as or 2 to 3 times weaker than those of omeprazole and that its duration is not as long as that of omeprazole in rats and dogs. Furthermore, YM020 possesses a cytoprotective effect and the mechanism of YM020 may be different to that of omeprazole.
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PMID:Antisecretory and antiulcer effects of YM020, a new H+,K(+)-ATPase inhibitor, in rats and dogs. 774 46

The present experiments were undertaken to extent our earlier observations (J Physiol Pharmacol 1991, 42, 367-79) relating membrane potential with membrane recycling of parietal cells. Studies were performed in vitro using gastric glands that were isolated through the use of rat stomachs transformed into "everted sacs" and filled with hyperosmolar NaCl-EDTA solution. Acid production was indirectly determined by accumulation of 14C-aminopyrine (AP) and its translocation by measurement of acridine orange fluorescence. H+/K(+)-ATPase activity was assayed by measurement of K(+)-stimulated p-nitrophenylphosphatase (pNPPase) of the proton pump. Morphologic state of parietal cells in relation to their functional activity was observed using electron microscopy. Changes in the membrane potential were obtained by the treatment of gastric glands with protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP) in the incubation media of different pH. CCCP caused time-dependent decrease in AP accumulation by parietal cells from the medium of pH 6.6 but not that of pH 7.8. pNPPase activity increased in aplical and decreased in tubulovesical membrances prepared from CCCP treated glands which were incubated in the medium being more acidic than cell cytoplasm. Electron microscopic assessment showed morphological transformation of resting parietal cells treated with CCCP in pH 6.6 from nonsecreting to secreting state. CCCP acting in acidic incubation medium also caused the decrease in acridine orange fluorescence in the cytoplasm of parietal cells with some temporary increase of its fluorescence in the lumen o gastric glands. These findings support our hypothesis that changes in parietal cell membrane potential by protonophore CCCP may translocate HCl from tubulovesicles to secretory canaliculi. While the above explanation is suggestive, the exact mechanisms controlling a membrane recycling during the secretory response of parietal cells in vitro remain to be elucidated.
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PMID:Functional and structural changes of isolated rat parietal cells during membrane potential modulation. 784 48

Basolateral uptake of chloride by the HCl-secreting parietal cells of the gastric (oxyntic) glands is most likely mediated by a HCO3-/Cl- anion exchange mechanism. Circumstantial evidence indicates that in rodents the anion exchange proceeds through an anion exchanger 2(AE2)-like membrane protein. In the present study, we raised antibodies against a bacterial fusion protein expressing a approximately 26-kDa portion of the human AE2 sequence. These antibodies were used to identify and localize AE2 in the human stomach. Here we report that the mucosa of the human stomach expresses an approximately 160-kDa immunoreactive form of AE2 containing the AE2-specific exoplasmic domain (Z-loop) as identified by polymerase chain reaction. Immunostaining specific for AE2 was restricted to the basolateral membrane domain of parietal cells and was also detected in small epithelial cells localized in the glandular isthmus region. The latter cells most likely represent pre-parietal cells. Parietal cells were identified by simultaneous and sequential labelling with antibodies against the gastric H+,K(+)-ATPase and actin, respectively. Both actin and the H+,K(+)-ATPase were localized along the apical membrane of parietal cells and the membrane of their secretory intracellular canaliculi. In addition, actin was shown to be colocalized with AE2 along the basolateral cell surface. Discontinuous staining for AE2 coincided with infoldings of the basolateral plasma membrane labelled by the actin antibody. These observations indicate that AE2 might be placed at specialized (folded) microdomains of the basolateral cell surface by linkage to the actin-based cytoskeleton.
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PMID:Basolateral localization of anion exchanger 2 (AE2) and actin in acid-secreting (parietal) cells of the human stomach. 784 88

We have examined the intrinsic fluorescence properties of a highly purified chloroplast H(+)-ATPase (CF0F1) preparation [R. D. Kirch and P. Graber (1992) Acta Physiol. Scand. 746, 9-12). Unlike the catalytic CF1 portion alone, CF0F1 fluorescence was dominated by tryptophan fluorescence both at 277-nm excitation, favoring tyrosine excitation, and at 295-nm excitation, favoring tryptophan excitation. A broad tryptophan fluorescence peak was observed with a maximum at around 335 nm and a broad shoulder around 350 nm. Denaturation of the enzyme complex with guanidine-HCl resulted in a significant increase (approximately 40%) in tyrosine fluorescence. The fluorescence spectrum (lambda ex = 295 nm) of the inhibitory epsilon subunit isolated from CF1 resembled that of CF1, indicating the presence of two tryptophan species located in different environments. Fluorescence quenching by potassium iodide indicated a substantial increase in the solvent accessibility of one of the two tryptophans following isolation of epsilon from CF1. Thus, when epsilon binds to CF1, a tryptophan residue becomes partially buried, probably at an interface between epsilon and another (possibly gamma) CF1 subunit. Removal of the epsilon subunit from CF1 leads to an increase in tyrosine fluorescence of a magnitude similar to that obtained upon denaturation of the CF0F1 complex. The results suggest that the reversible association of the epsilon subunit with CF0F1 or with isolated CF1 may be monitored by following changes in the intrinsic fluorescence of the enzyme complex.
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PMID:Intrinsic fluorescence of the chloroplast H(+)-ATPase. 787

The effect of leminoprazole ((+-)-2-[[2-(isobutyl-methylamino)benzyl]sulfinyl]-1H-benzimidazol e, NC-1300-O-3, CAS 104340-86-5), a new compound being developed as an inhibitor of the gastric mucosal proton pump (H+,K(+)-ATPase), on gastric mucus secretion was studied by a biochemical method measuring the gastric mucin content in rats. Oral administration of leminoprazole (30 mg/kg) strongly inhibited the hemorrhagic lesions induced by 60% ethanol containing 0.15 mol/l HCl (acid-ethanol) administered 1 h later. Leminoprazole significantly inhibited the acid-ethanol-induced reduction of the mucin content in the surface mucosa including the mucus gel layer, but no significant effect could be obtained on the reduction of mucin present in the deep layer of the corpus and antral mucosa. Leminoprazole given to rats not treated with acid ethanol accelerated the secretion of deep mucosal mucus and increased significantly the content of soluble mucus which was recovered from the gastric luminal contents to about 200% of control, but failed to produce any significant change in the mucus content present in the surface mucosal and the mucus gel layers. The effect of leminoprazole on gastric mucus secretion might contribute to healing the peptic ulcer diseases and may be involved in the cytoprotective mechanism of this drug.
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PMID:Effect on gastric mucus of the proton pump inhibitor leminoprazole and its cytoprotective action against ethanol-induced gastric injury in rats. 794 16

In vivo ethanol exposure reduces in vitro Na+,K(+)-adenosine triphosphatase (Na+,K(+)-ATPase) sensitivity to ethanol in some animal models, but very little is known about the effects of ethanol on human brain Na+,K(+)-ATPase. Cerebral cortex homogenates from 13 male alcoholic and 9 control subjects were assayed for K(+)-p-nitrophenylphosphatase (K(+)-pNPPase, a measure of Na+,K(+)-ATPase) and Mg(2+)-pNPPase activities at 37 degrees for 20 min in 75 mM imidazole-HCl (pH 7.4), 5 mM p-nitrophenylphosphate, 5 mM MgCl2, and 20 mM KCl, with or without 1 mM ouabain. Native K(+)-pNPPase activites were similar in control and alcoholic brains (61.5 +/- 3.5 vs 55.3 +/- 3.1 nmol/mg/min). In vitro exposure to a near lethal ethanol level (0.5%, or 110 mM) was without effect, whereas 5% ethanol inhibited K(+)-pNPPase activity by about 28% (P < 0.001) in both groups. Both 0.5 and 5% ethanol in vitro significantly stimulated Mg(2+)-pNPPase activity (1-2% and 19-20%, respectively). By comparison, mouse brain K(+)-pNPPase was inhibited significantly by in vitro ethanol, and Mg(2+)-pNPPase activity was unaffected. Ethanol levels attainable in humans may not be sufficient to alter significantly brain Na+,K(+)-ATPase activity.
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PMID:Effects of in vitro ethanol on the brain cation pump in alcoholics and controls. 805 41


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