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)

Hexanol at 7 degrees C stimulates the activity of the Ca-ATPase of sarcoplasmic reticulum (SR). Time-resolved phosphorescence spectroscopy studies of SR whose Ca-ATPase is covalently labeled with erythrosin isothiocyanate (ERITC) indicate that at 7 degrees C hexanol (1) cause a concentration-dependent increase in the rate of decay of phosphorescence anisotropy, (2) causes larger oligomers of Ca-ATPase to dissociate into smaller oligomers, and (3) increases the rotational mobility of Ca-ATPase in all its oligomeric states. Electron paramagnetic resonance (EPR) spectroscopy of spin-labeled stearic acid (SASL) in SR suggests that at 7 degrees C hexanol diminishes the fraction of SR lipids in the boundary lipid domain and disorders and fluidizes both the boundary lipid and the unrestricted lipid domain. In protein-free liposomes of extracted SR lipids hexanol increases fluidity and decreases order to a greater extent near the center of the lipid bilayer than near the polar head groups. At 25 degrees C hexanol has biphasic effects on Ca-ATPase activity: at 10 and 20 mM hexanol increases activity, but at 30 mM and especially at 40 mM there is inhibition of Ca-ATPase activity. The influence of hexanol at 25 degrees C on the oligomeric state of Ca-ATPase is also biphasic. At 10 and 20 mM, hexanol promotes the dissociation of larger oligomers into smaller ones, whereas at higher concentrations, 30 and 40 mM, hexanol causes larger oligomers to be formed from smaller ones. Lidocaine at 25 degrees C inhibits Ca-ATPase activity and causes dramatic slowing of the decay of phosphorescence anisotropy of ERITC-labeled SR by causing the formation of larger oligomers of Ca-ATPase from smaller ones. In protein-free liposomes of SR lipids at 25 degrees C, lidocaine disorders and fluidizes the acyl chains near the center of the bilayer (as did hexanol), but has opposite effects near the polar head groups. The opposite effects of hexanol and lidocaine on the oligomeric state of the SR Ca-ATPase provide a new molecular explanation for the opposite effects of hexanol and lidocaine on the activity of the Ca-ATPase. We conclude that the biphasic effects of hexanol on the activity of Ca-ATPase can be accounted for by biphasic effects of hexanol on the oligomeric state of the Ca-ATPase. This study supports the view that anesthetics can alter interactions between membrane proteins.
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PMID:Hexanol and lidocaine affect the oligomeric state of the Ca-ATPase of sarcoplasmic reticulum. 794 28

The effects of sodium (Na+) and chloride ions (Cl-) on blood pressure were studied in rats treated with deoxycorticosterone acetate (DOCA). Four groups were prepared, each consisting of male Wistar rats that underwent heminephrectomy and administration of DOCA: the control group was maintained with tap water, the NaCl group with tap water containing 1% sodium chloride, the NaCit group with tap water containing 1.67% sodium citrate (including an equivalent dose of Na+ to 1% NaCl), and the ChoCl group with tap water containing 1.15% choline chloride (including an equivalent dose of Cl- to 1% NaCl). The time-course of systolic blood pressure showed only slight change in blood pressure in the control and ChoCl groups, and in the NaCl and NaCit groups. The rotational correlation time, an index of the fluidity of erythrocyte membrane, with spin-labeling of 16-doxyl-stearic acid, was significantly (p < 0.05) higher in the NaCl and NaCit groups than in the control group, indicating an increase in the membrane fluidity, i.e., membrane fragility. The sodium, potassium ions-activated adenosine triphosphatase (Na+,K(+)-ATPase) activity of the erythrocyte membrane was decreased to 22% (P < 0.01) and 24% (P < 0.01) in the NaCl and NaCit groups, respectively, compared with the control groups; this activity was decreased to 43% in the ChoCl group (P < 0.05). The Ca(2+)-ATPase activity showed similar changes. In contrast, there were no marked differences in the erythrocyte electrolyte level between the groups.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Effects of sodium and chloride ions on blood pressure in deoxycorticosterone acetate-treated rats. 796 82

Endothelial cell integrity has been suggested to play a role in the development of atherosclerosis. The effects of fatty acids on endothelial barrier function were tested by measuring albumin transport across endothelial monolayers cultured on polycarbonate filters. Compared with control cultures, a 24-h exposure to 90 mumol/L lauric (12:0) and linoleic acid (18:2) but not to butyric (4:0), hexanoic (6:0), octanoic (8:0), decanoic (10:0), myristic (14:0), palmitic (16:0) or stearic acid (18:0) caused an increase in albumin transfer across endothelial monolayers. Selective enrichment of a "physiological" serum fatty acid mixture (FA-Mix; 90 mumol/L) with 90 mumol/L of 12:0 or 18:2 significantly increased albumin transfer, whereas enrichment with 90 mumol/L of 4:0, 16:0 or 18:0 significantly decreased albumin transfer relative to 180 mumol/L FA-Mix. Only 12:0- or 18:2-treated cultures showed increased Ca(++)-ATPase activity and the presence of lipid droplets. Fatty acids (60 mumol/L) extracted from butter fat and beef tallow had no effect on albumin transfer, whereas fatty acids extracted from chicken fat and corn oil consistently disrupted endothelial barrier function. This fat-induced disruption of endothelial barrier function seems to be related to the amount of 18:2 present in each fat source. These data indicate that unsaturated fats cause cellular perturbations that result in a decrease in endothelial barrier function in this model system, and that high dietary levels of unsaturated fats may be detrimental to cell integrity.
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PMID:Selective disruption of endothelial barrier function in culture by pure fatty acids and fatty acids derived from animal and plant fats. 832 May 62

We have used time-resolved phosphorescence anisotropy and electron paramagnetic resonance (EPR) spectroscopy to detect the rotational dynamics of the Ca-ATPase and its associated lipids in dog cardiac sarcoplasmic reticulum (DCSR), in comparison with rabbit skeletal SR (RSSR), in order to obtain insight into the physical bases for different activities and regulation in the two systems. Protein rotational motions were studied with time-resolved phosphorescence anisotropy (TPA) of erythrosin isothiocyanate (ERITC) and saturation-transfer EPR (ST-EPR) of a maleimide spin-label (MSL). Both labels were attached selectively and rigidly to the Ca-ATPase. Lipid rotational motions were studied with conventional EPR of stearic acid spin-labels. As in previous studies on RSSR, the phosphorescence anisotropy decays of both preparations at 4 degrees C were multiexponential, due to the presence of different oligomeric species. The rotational correlation times for the different rotating species were similar for the two preparations, but the total decay amplitude was substantially less for cardiac SR, indicating that more of the Ca-ATPase molecules are in large aggregates in DCSR. ST-EPR spectra confirmed that the Ca-ATPase is less rotationally mobile in DCSR than in RSSR. Lipid probe mobility and fatty acid composition were very similar in the two preparations, indicating that the large differences observed in protein mobility are not due to differences in lipid fluidity. We conclude that the higher restriction in protein mobility observed by both ST-EPR and TPA is due to more extensive protein-protein interactions in DCSR than in RSSR.
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PMID:Protein and lipid rotational dynamics in cardiac and skeletal sarcoplasmic reticulum detected by EPR and phosphorescence anisotropy. 839 31

We have used spin-label EPR spectroscopy to examine possible alterations in protein-protein interactions that accompany the activation of the cardiac sarcoplasmic reticulum (SR) Ca-ATPase following the phosphorylation of phospholamban (PLB). Using a radioactive derivative of a maleimide spin label (MSL), we have developed conditions for the selective spin-labeling of the Ca-ATPase in both native cardiac and skeletal sarcoplasmic reticulum membranes. The rotational dynamics of the cardiac and skeletal Ca-ATPase isoforms in native SR membranes were measured using saturation transfer EPR. We report that the phosphorylation of PLB in cardiac SR results in a (1.8 +/- 0.2)-fold reduction in the overall rotational mobility of the Ca-ATPase. The alteration in the rotational dynamics of the Ca-ATPase is the direct result of the phosphorylation of PLB, and is not related to the phosphorylation of the Ca-ATPase or any other SR proteins since no alteration in the ST-EPR spectrum is observed as a result of conditions that phosphorylate the cardiac Ca-ATPase with ATP. Neither do the use of conditions that activate the Ca-ATPase in cardiac SR result in the alteration of the rotational dynamics or catalytic properties of the Ca-ATPase in skeletal SR where PLB is not expressed. Measurements of the rotational dynamics of stearic acid spin labels (SASL) incorporated into cardiac SR membranes with a nitroxide at the 5- and 12-positions using conventional EPR indicate that there is virtually no difference in the lipid acyl chain dynamics in cardiac SR membranes upon the phosphorylation of PLB. These results indicate that the decrease in the rotational dynamics of the Ca-ATPase in cardiac SR membranes associated with the phosphorylation of PLB is related to enhanced interactions between individual Ca-ATPase polypeptide chains due to (i) an alteration in the spatial arrangement of cardiac Ca-ATPase polypeptide chains within a defined oligomeric state or (ii) increased protein-protein associations. We suggest that altered interactions between Ca-ATPase polypeptide chains and PLB serves to modulate the activation barrier associated with calcium activation of the Ca-ATPase in cardiac SR membranes.
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PMID:Phosphorylation of phospholamban by cAMP-dependent protein kinase enhances interactions between Ca-ATPase polypeptide chains in cardiac sarcoplasmic reticulum membranes. 878 78

The rotational diffusion of Ca2(+)-ATPase [Ca2+,Mg2(+)-activated ATP phosphohydrolase E.C. 3.6.1.38] was studied in native sarcoplasmic reticulum membrane by saturation transfer ESR spectroscopy after covalent labelling of intramembranous sulfhydryl groups with nitroxyl derivative of maleimide (5-MSL) as a function of sucrose and glycerol in the suspending medium. The relative enzymatic activity of sarcoplasmic reticulum was followed by increasing the viscosity of the aqueous phase. The ATP hydrolysing activity of the enzyme decreased differently on adding sucrose and glycerol. In the case of sucrose the reciprocal of power dependence of viscosity was observed, whereas for glycerol an exponential decay law was obtained, indicating solvent-protein interaction. On increasing the viscosity of the aqueous phase by either sucrose or glycerol, no changes were observed in the intramembranous viscosity as measured using intercalated spin-labelled stearic acid (16-SASL). The effective rotational correlation time of the protein was measured, as a mobility parameter, using saturation transfer ESR spectroscopy and found to be increased linearly with the viscosity of the sucrose containing medium and for the extramembranous size a height of 6.8 nm was obtained, indicating that approx. 82% of the volume of Ca2(+)-ATPase protein is external to the sarcoplasmic reticulum. The addition of glycerol probably promoted protein-protein interaction, as indicated by the larger changes in rotational diffusion and non-linear viscosity dependence.
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PMID:Rotational mobility of Ca2+-ATPase of sarcoplasmic reticulum in viscous media. 921 50

Benzophenone (BP) was used as a photosensitizer to initiate lipid peroxidation in model and native biological membranes at concentrations of BP that do not perturb bilayer structure, as assessed by stearic acid spin label dynamics. Illumination of BP partitioned into sarcoplasmic reticulum membranes (SR) results in an exponential decay of BP and a linear accumulation of conjugated dienes and other products of lipid peroxidation as observed previously for micelles of linoleic acid [Marcovic and Patterson. Photochem. Photobiol. 58:329-334, 1993]. Lipid peroxidation was substantially inhibited in the presence of membrane-spanning proteins in SR compared to protein-free lipid vesicles, suggesting the competitive reaction of the initiator (triplet BP) and BP-derived radical species with protein groups. Modification of the predominant integral membrane protein, the Ca(2+)-ATPase, was demonstrated by changes in Ca(2+)-ATPase amino acid composition as well as by its functional inhibition. The rate of calcium transport showed an immediate exponential decay to completion, while calcium-dependent ATPase activity exhibited an initial lag before modest inactivation. These results are consistent with the respective localization of calcium transport sites within membrane-spanning peptides and the ATP-binding site within the cytosolic domain of the Ca(2+)-ATPase, further suggesting that photosensitization of BP models oxidative stress inside the hydrophobic interior of the SR membrane.
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PMID:Benzophenone-sensitized photooxidation of sarcoplasmic reticulum membranes: site-specific modification of the Ca(2+)-ATPase. 935 44

Arachidonic acid, phosphatidic acid, and other lipids inhibit the catalytic fragment of neurofibromin more potently than that of p120 guanosine triphosphatase-activating protein (GAP). The effects of fatty acids other than arachidonic acid on full-length neurofibromin and p120 GAP, to our knowledge, have not been studied. In this study, we analyzed the effects of eight nutritionally relevant fatty acids on guanosine triphosphatase (GTPase) stimulatory activity of full-length neurofibromin and p120 GAP. The fatty acids tested were saturated stearic acid, monounsaturated oleic acid, and three n-6 and three n-3 polyunsaturated fatty acids. Analysis was performed by Ras immunoprecipitation GTPase assay. The full-length p120 GAP expressed in insect Sf9 cells and immunoaffinity-purified full-length neurofibromin were used. In contrast to neurofibromin, which was readily inhibited by stearic and oleic acid, p120 GAP was only weakly inhibited even at high concentrations (> 80 microM). Neurofibromin was also two- to threefold more sensitive to inhibition by other fatty acids tested. A chimeric protein in which the neurofibromin catalytic domain was fused to the NH2-terminal sequences of p120 GAP was used to determine that differential sensitivity to fatty acid inhibition maps to the catalytic domain of the proteins. These results indicate that nutritionally relevant fatty acids can modulate the GTPase function of c-Ha-Ras protein by inhibiting GTPase stimulatory activity of two Ras regulators, full-length neurofibromin and p120 GAP, at physiologically relevant concentrations in vitro.
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PMID:Differential regulation of neurofibromin and p120 GTPase-activating protein by nutritionally relevant fatty acids. 958 27

The selectivity of the lipid-protein interactions in trypsinised Na, K-ATPase membranes from Squalus acanthias has been determined by using EPR spectroscopy with different lipid probes spin-labelled on the 14-C atom of the fatty acid chain. From measurements at low ionic strength and different pH values, the pattern of selectivity is: (stearic acid)->(phosphatidylserine)->(stearic acid)0>(phosphatidylcholine)+/-, where superscripts indicate the formal electrostatic charge on the lipid headgroup. This is in the same order as that determined with native Na,K-ATPase membranes [M. Esmann, D. Marsh, Biochemistry 24 (1985) 3572-3578]. The selectivity for phosphatidylserine is independent of pH, over the range pH 6.0-9. 0, as found also for native membranes. For membranes trypsinised in the presence of Rb+ ions, and in the presence of Na+ (which allows more extensive proteolysis), the relative association constants, Kr, of all lipids are the same as for control membranes, with the exception of ionised (stearic acid)- that shows the highest specificity. Therefore, both the stoichiometry and the principal determinants of the specificity of lipid-protein interaction are preserved on extensive trypsinisation of Na,K-ATPase membranes. This has implications for the location and arrangement of those amino acid side chains that determine the lipid selectivity of the native Na,K-ATPase.
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PMID:Selectivity of lipid-protein interactions with trypsinized Na, K-ATPase studied by spin-label EPR. 963 Jun 3

The interaction of the tertiary amine drugs chlorpromazine and dibucaine in their cationic form with carboxyl groups at the membrane surface is studied at concentrations relevant to anesthesia. Spin-labeled stearic acid is used both to provide the carboxyl groups and to monitor binding and ionization behavior in egg lecithin liposomes. Membrane anesthetic concentrations are spectrophotometrically obtained. They are shown to determine the drug influence on carboxyl groups at the membrane surface, independently of aqueous concentrations. The intramembrane association constants (related to the usual aqueous phase ones through the partition coefficient) of the drugs with fatty acids are determined. The same value (10(2) M-1) is obtained for both drugs, suggesting that it is approximately the same for all tertiary amine local anesthetics. pH titrations of anesthetic-treated spin-labeled membranes are performed. The observed shifts in the fatty acid pK are higher than can be produced assuming uniform distribution of the drug in the membrane surface, implying that there is an increased affinity of local anesthetics for superficial carboxyl. This affinity could account for the resting block of voltage-gated Na+ channels. Under these considerations, local anesthetic binding sites at voltage-gated Na+ channels and at sarcoplasmic reticulum Ca(2+)-ATPase are proposed.
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PMID:Carboxyl groups at the membrane interface as molecular targets for local anesthetics. 974 84


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