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)

To investigate the molecular mechanisms of the inhibition of Na+,K(+)-adenosine triphosphatase (Na+,K(+)-ATPase) in diabetes mellitus, we incubated Na+,K(+)-ATPase purified from human placenta of six healthy nondiabetic women with plasma from six insulin-dependent diabetic (IDDM) men and six healthy controls and with different concentrations of lysophosphatidylcholine (LPC). We determined the enzyme activity, anthroyl ouabain-binding capacity, dissociation constant (Kd), and average lifetime values (tau) by the static and dynamic fluorescence of anthroyl ouabain. The lipid annulus of the enzyme was studied by static and dynamic fluorescence of 1-(4-trimethylamino-phenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH). Moreover, we studied the lipid microenvironment surrounding the Na+,K(+)-ATPase purified from the placentas of six healthy women and six insulin-dependent diabetic women, determining the percent composition of phospholipids of the lipid annulus. The addition of total and protein-free IDDM plasma to normal Na+,K(+)-ATPase significantly inhibited the enzymatic activity even at the lowest concentration studied (1: 100), whereas the ouabain-binding capacity, Kd, and tau were not affected by IDDM plasma. The fluorescence polarization and lifetime values of TMA-DPH were significantly decreased by diabetic plasma. The incubation of Na+,K(+)-ATPase with LPC caused an inhibition of the enzymatic activity without modifications of the anthroyl ouabain-binding capacity and dissociation constant. The fluorescence polarization and lifetime values of TMA-DPH were significantly decreased by 5 mumol/L LPC. The study of the phospholipids surrounding Na+,K(+)-ATPase demonstrated a significant increase in the percent LPC content in IDDM patients compared with controls together with a concomitant decrease in phosphatidylcholine. These observations indicate that the inhibition caused by diabetic plasma on Na+,K(+)-ATPase is not dependent on a modification of the ouabain-binding site and that it seems to mimic the effect of LPC addition. A link between modification of the lipid moiety of the enzyme and Na+,K(+)-ATPase inhibition might be hypothesized.
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PMID:Modifications induced by plasma from insulin-dependent diabetic patients and by lysophosphatidylcholine on human Na+,K(+)-adenosine triphosphatase. 966 19

Catalytically important motions of the Ca-ATPase, modulated by the physical properties of surrounding membrane phospholipids, have been suggested to be rate-limiting under physiological conditions. To identify the nature of the structural coupling between the Ca-ATPase and membrane phospholipids, we have investigated the functional and structural effects resulting from the incorporation of the lysophospholipid 1-myristoyl-2-hydroxy-sn-glycerol-3-phosphocholine (LPC) into native sarcoplasmic reticulum (SR) membranes. Nonsolubilizing concentrations of LPC abolish changes in fluorescence signals associated with either intrinsic or extrinsic chromophores that monitor normal conformational transitions accompanying calcium activation of the Ca-ATPase. There are corresponding decreases in the rates of calcium transport coupled to ATP hydrolysis, suggesting that LPC may increase conformational barriers associated with catalytic function. Fluorescence anisotropy measurements of the lipid analogue 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH) partitioned into SR membranes indicate that LPC does not significantly modify lipid acyl chain rotational dynamics, suggesting differences in headgroup conformation between LPC and diacylglycerol phosphatidylcholines. Complementary measurements using phosphorescence anisotropy of erythrosin isothiocyanate at Lys464 on the Ca-ATPase provide a measure of the dynamic structure of the phosphorylation domain, and indicate that LPC restricts the amplitude of rotational motion. These results suggest a structural linkage between the cytosolic phosphorylation domain and the conformation of membrane phospholipid headgroups. Thus, changes in membrane phospholipid composition can modulate membrane surface properties and affect catalytically important motions of the Ca-ATPase in a manner that suggests a role for LPC generated during signal transduction.
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PMID:Lysophosphatidylcholine modulates catalytically important motions of the Ca-ATPase phosphorylation domain. 1019 82

The aim of the present study was to evaluate the action of plasma from insulin-dependent diabetic (IDDM) pregnant women on nitric oxide synthase (NOS) activity in cultured human umbilical vein endothelial cells (HUVECs). We also studied the effect of the plasma on cytosolic calcium and on Na+/K+-adenosine triphosphatase (ATPase) activity. Dynamic fluorescence studies of membrane fluidity were contemporarily performed to detect a direct effect of plasma on the endothelial cell membrane. We observed a significant increase in NOS activity, intracellular calcium, and Na+/K+-ATPase activity in cultured HUVECs exposed to IDDM plasma. Our dynamic fluorescence study showed a different microenvironmental organization of the cellular membrane after incubation with plasma from IDDM pregnant women, with a marked decrease in microheterogeneity as evaluated in terms of 1-(4-trimethylaminophenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH) lifetime distribution width. The present investigation suggests that plasma from IDDM pregnant women can cause a generalized disturbance in the function of endothelial cells cultured from healthy subjects. Such a modification might play a central role in the pathogenesis of the vascular complications of the disease.
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PMID:A study on human umbilical cord endothelial cells: functional modifications induced by plasma from insulin-dependent diabetes mellitus patients. 1033 52

The aim of the present work was to analyze the effect of LDL obtained from type 1 diabetic patients in good metabolic control on human umbilical vein endothelial cells (HUVECs) after a short incubation period to detect possible atherogenic modifications of endothelial properties. Cultured HUVECs were incubated for 3 h with culture medium alone (control HUVEC), with native LDL from 12 healthy men (control LDL), or with native LDL from 12 type 1 diabetic men (type 1 LDL) (100 pg/ml). After the incubation, the following parameters were evaluated: nitric oxide synthase (NOS) activity, cytoplasmic Ca2+ levels, Na+-K+-ATPase activity, plasma membrane fluidity determined by means of 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1-(4-trimethylaminophenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH), and plasma membrane conjugated diene (CD) content. The same experiments were repeated after bradykinin stimulation or in the presence of the antioxidant butylated hydroxytoluene (BHT), and nitric oxide (NO) production in intact HUVECs was also evaluated. HUVECs incubated with control LDL in comparison with control HUVECs showed a decreased fluidity of the membrane surface evaluated by TMA-DPH and a higher CD content. These alterations were prevented by the presence of BHT. HUVECs incubated with type 1 LDL in comparison with both control HUVECs and cells incubated with control LDL showed 1) increased NOS and Na+-K+-ATPase activity, cytoplasmic Ca2+ levels, and CD content, and 2) decreased fluidity of the membrane surface evaluated by TMA-DPH. These modifications were blunted--but not abolished--by the presence of BHT. After bradykinin stimulation either in the absence or in the presence of BHT, both cytoplasmic Ca2+ levels and NO production were increased in control HUVECs and in HUVECs incubated with control LDL, while a reduced response was observed in HUVECs incubated with type 1 LDL. The alterations observed in the endothelial function after the cell-LDL interaction might play a central role in the atherogenic process in diabetes.
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PMID:Modifications induced by LDL from type 1 diabetic patients on endothelial cells obtained from human umbilical vein. 1053 58

Alterations in membrane lipid composition is known to result in functional and structural changes in the membrane, and dietary lipids play an important role in this change. It was of interest to study the influence of ghee feeding to the rat on membrane structure and function. The activities of membrane bound enzymes Na+ K+ ATPase and Acetylcholinesterase were studied as an index of membrane changes. Male weanling rats were fed 2.5% fresh or thermally oxidized ghee in the diet for a period of 8 weeks. The control rats were fed groundnut oil. A decrease of 28% in the membrane fluidity of erythrocyte ghost membranes was observed in the oxidized ghee fed group at 37 degrees C, by fluorescence polarization measurements using 1,6-Diphenyl-1,3,5-hexatriene as a probe. The activities of Na+ K+ ATPase and Acetylcholinesterase showed an increase of 65 and 200% respectively after feeding oxidized ghee (2.5%). Also changes in Na+, K+ and ATP kinetics were observed in these rats. Increased membrane lipid peroxidation (80%) and C/PL ratio (11%) in the oxidized ghee fed group was observed. Marginal changes in the fatty acid composition were also seen. Further, an increase in the osmotic fragility of erythrocytes was observed in the oxidized ghee fed rats. It is inferred from these experiments that consumption of oxidized ghee with the diet affects the erythrocyte ghost membrane structure and function at 2.5% level, whereas consumption of fresh ghee has no effect on the erythrocyte membrane.
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PMID:Membrane changes in rat erythrocyte ghosts on ghee feeding. 1071 25

Plasma membrane lipid disorganization takes place in cells of Saccharomyces cerevisiae grown under copper stress, as shown by fluorescence anisotropy measurements with the lipid reporter probe 1,6-diphenyl-1,3,5-hexatriene. The extent of plasma membrane disorganization, presumably due to copper-induced lipid peroxidation, was discontinuous when measured in cells grown in media supplemented with different concentrations of CuSO4. Results suggested the existence of adaptive mechanisms that cells employ to protect themselves against the deleterious effects of copper. The adaptive mechanisms examined in this study included the coordinate increase in the activities of Cu,Zn-superoxide dismutase (up to five-fold), glutathione reductase (up to 1.7-fold), and plasma membrane H+-ATPase (up to three-fold). These enzyme activities showed maximal levels in cells grown with copper supplied at intermediate concentrations, within the range that allowed growth. Significantly, at these concentrations, plasma membrane disorganization did not increase when increasing amounts of CuSO4 were supplied. However, at copper concentrations close to the maximal that allowed growth, the capacity of the yeast cell response to cope with the deleterious effects of copper was exceeded; plasma membrane lipid organization and plasma-membrane-bound H+-ATPase activity drastically declined in response to the increased levels of copper stress and the consequences on growth kinetics were even more severe. Our results clearly suggest that modification of plasma membrane H+-ATPase activity is either part of or the result of the global response of yeast to mild or high copper stress.
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PMID:Modification of plasma membrane lipid order and H+-ATPase activity as part of the response of Saccharomyces cerevisiae to cultivation under mild and high copper stress. 1081 44

The ATP-generated proton pumping across tonoplast vesicles from chilling-sensitive Boro rice (Oryza sativa L. var. Boro) cultured cells was markedly decreased by chilling at 5 degrees C for 3 d. The membrane fluidity of core hydrophobic and surface hydrophilic regions of the lipid bilayer was measured by steady-state fluorescence depolarization of 1,6-diphenyl-1,3,5-hexatriene and trimethylammonium 1,6-diphenyl-1,3,5-hexatriene and by electron spin resonance spectroscopy of 16- and 5-doxyl stearic acid, respectively. The fluidity of the surface region of the lipid bilayer of the tonoplast vesicles decreased by chilling. The fluidity of the surface region of the liposomes and the proton pumping across the reconstituted proteoliposomes with tonoplast H+-ATPase decreased with increasing content of the glycolipids. The proton pumping across chimera proteoliposomes was reduced by chilling only when it was reconstituted in the presence of tonoplast glycolipids from chilled Boro cells. These data suggest that the reduction in ATP-generated proton pumping across the tonoplast by chilling is due to the decrease in the fluidity of the surface region of the lipid bilayer of the tonoplast, which is caused by the changes in glycolipids.
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PMID:Mechanism of the chilling-induced decrease in proton pumping across the tonoplast of rice cells. 1096 40

The effect of tricyclic antidepressant drugs amitriptyline, nortriptyline, imipramine and desipramine on synaptosomal membrane and lipid bilayer was studied using steady state and time dependent fluorescence spectroscopy of lipid specific fluorescent probe 1,6-diphenyl-1,3,5-hexatriene (DPH). The synaptosomal membrane was prepared from rat brain while liposomes were prepared from dimyristoyl phosphatidyl choline (DMPC) alone and a mixture of DMPC and cholesterol. Upon treatment with amitriptyline and nortriptyline a decrease was observed in the steady state anisotropy of DPH in DMPC liposomes as well as in rat brain synaptosomes. On the other hand, imipramine and desipramine did not cause any significant change. Amitriptyline and nortriptyline also decreased the steady state anisotropy of DPH in liposomes prepared from a mixture of DMPC and cholesterol. Fluorescence decay time and time dependent anisotropy of DPH in both the membranes were measured and the decay of anisotropy was analyzed using wobbling in cone model. Amitriptyline and nortriptyline treatment decreased the limiting anisotropy and order parameter, while the cone angle increased. Imipramine and desipramine did not cause significant change in these parameters. In addition to structural alterations, these drugs inhibited the activity of Na+-K+-ATPase in synaptosomal membrane, however, the decrease was more in case of amitriptyline and nortriptyline as compared to imipramine and desipramine. Our results suggest that the perturbation in membrane order caused by antidepressant drugs could depend on the net charge on the drug molecule.
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PMID:Differential effects of tricyclic antidepressant drugs on membrane dynamics--a fluorescence spectroscopic study. 1113 48

Pluronic block copolymer, P85, inhibits the P-glycoprotein (Pgp) drug efflux system and increases the permeability of a broad spectrum of drugs in the blood-brain barrier (BBB). This study examines the mechanisms by which P85 inhibits Pgp using bovine brain microvessel endothelial cells (BBMEC) as an in vitro model of the BBB. The hypothesis was that simultaneous alterations in intracellular ATP levels and membrane fluidization in BBMEC monolayers by P85 results in inhibition of the drug efflux system. The methods included the use of 1) standard Pgp substrate rhodamine 123 to assay the Pgp efflux system in BBMEC, 2) luciferin/luciferase assay for ATP intracellular levels, and 3) 1,6-diphenyl-1,3,5-hexatriene for membrane microviscosity. Using 3H-labeled P85 and fluorescein-labeled P85 for confocal microscopy, this study suggests that P85 accumulates in the cells and intracellular organelles such as the mitochondria where it can interfere with metabolic processes. Following exposure of BBMEC to P85, the ATP levels were depleted, and microviscosity of the cell membranes was decreased. Furthermore, P85 treatment decreased Pgp ATPase activity in membranes expressing human Pgp. A combination of experiments examining the kinetics, concentration dependence, and directionality of P85 effects on Pgp-mediated efflux in BBMEC monolayers suggests that both energy depletion (decreasing ATP pool available for Pgp) and membrane fluidization (inhibiting Pgp ATPase activity) are critical factors contributing to the activity of the block copolymer in the BBB.
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PMID:Mechanism of pluronic effect on P-glycoprotein efflux system in blood-brain barrier: contributions of energy depletion and membrane fluidization. 1160 58

Sialic acid (SA) content, membrane fluidity, and Na(+)/K(+)-adenosine triphosphatase (ATPase) activity were determined in erythrocyte membrane from 10 nonpregnant women (HNPW), 16 pregnant women affected by gestational diabetes mellitus (GDM), and 25 healthy pregnant women (HPW). In GDM patients the membrane erythrocyte SA content was significantly increased compared with HNPW and membrane fluidity was significantly increased in comparison with HPW. Erythrocyte membrane Na(+)/K(+)-ATPase activity was significantly reduced in GDM patients compared both to HNPW and to HPW subjects. A significant inverse correlation was found between 1-(4-trimethylaminophenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH) anisotropy and erythrocyte membrane SA content in HNPW and in HPW, while this significant correlation was not observed in GDM. The present results indicate that in comparison with normal pregnancy GDM is characterized by deep alterations of the erythrocyte plasma membrane physicochemical properties (increased fluidity) and functional activities (reduced Na(+)/K(+)-ATPase activity). These modifications might be at the basis of the altered blood viscosity and placental perfusion observed under such conditions. Moreover, these results show that in physiological pregnancy and in the nonpregnant state, the erythrocyte surface membrane fluidity is inversely correlated with SA content, while in GDM there is an unbalance of this relation, which might be associated with the microcirculatory abnormality present in this disease.
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PMID:Sialic acid content in erythrocyte membranes from pregnant women affected by gestational diabetes. 1197 93


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