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)

The purification of axonal membranes of crustaceans was followed by measuring enrichment in [3H]tetrodotoxin binding capacity and in Na+, K+-ATPase activity. A characteristic of these membranes is their high content of lipids and their low content of protein as compared to other types of plasmatic membranes. The axonal membrane contains myosin-like, actin-like, tropomyosin-like, and tubulin-like proteins. It also contains Na+, K+-ATPase and acetylcholinesterase. The molecular weights of these two enzymes after solubilization are 280,000 and 270,000, respectively. The molecular weights of the catalytic subunits are 96,000 for ATPase and 71,000 for acetylcholinesterase. We confirmed the presence of a nicotine binding component in the axonal membrane of the lobster but we have been unable to find [3H]nicotine binding to crab axonal membranes. The binding to axonal membranes og of the sodium channel, has been studied in detail. The dissociation constant for the binding of [3H]tetrodotoxin to the axonal membrane receptor is 2.9 nM at pH 7.4. The concentration of the tetrodotoxin receptor in crustacean membranes is about 10 pmol/mg of membrane protein, 7 times less than the acetylcholinesterase, 30 times less than the Na+, K+-ATPase, and 30 times less than the nicotine binding component in the lobster membrane. A reasonable estimate indicates that approximately only one peptide chain in 1000 constitutes the tetrodotoxin binding part of the sodium channel in the axonal membrane. Veratridine, which acts selectively on the resting sodium permeability, binds to the phospholipid part of the axonal membrane. [3H]Veratridine binding to membranes parallels the electrophysiological effect. Veratridine and tetrodotoxin have different receptor sites. Although tetrodotoxin can repolarize the excitable membrane of a giant axon depolarized by veratridine, veratridine does not affect the binding of [3H]tetrodotoxin to purified axonal membranes. Similarly, tetrodotoxin does not affect the binding of [3H]veratridine to axonal membranes. Scorpion neurotoxin I, a presynaptic toxin which affects both the Na+ and the K+ channels, does not interfere with the binding of [3H]tetrodotoxin or [3H]veratridine to axonal membranes. Tetrodotoxin, veratridine, and scorpion neurotoxin I, which have in common the perturbation of the normal functioning of the sodium channel, act upon three different types of receptor sites.
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PMID:Constitution and properties of axonal membranes of crustacean nerves. 0 58

The biochemical properties of the electrically excitable sodium channels in the electroplaque of Electrophorus electricus were investigated using tritiated tetrodotoxin (TTX) as a specific membrane probe. Membrane fragments from the electroplaque were isolated essentially by differential centrifugation and characterized with respect to the plasma membrane markers acetylcholine receptors, acetylcholinesterase, (Na+ + K+)ATPase, and [3H]TTX binding. Equilibrium binding studies showed that [3H]TTX bound to a single population of noninteracting receptor sites with an apparent dissociation constant of 6 +/- 1 X 10(-9) M. The toxin-membrane complex dissociated with a first-order rate constant of 0.012 sec-1. Studies on the pH dependence of complex formation demonstrated the requirement for an ionizable, functional group with a pK of 5.3 and this group has been shown to be a carboxyl. Treatment of the membranes with trimethyloxonium tetrafluoroborate, a carboxyl group modifying reagent, resulted in an irreversible loss in the binding of [3H]TTX, which could be prevented by low concentrations of TTX or saxitoxin. This decrease was due to a reduction in the total number of binding sites and not to a decrease in toxin binding affinities. The relative binding affinities of various monovalent alkali metal and polyatomic cations for the TTX-receptor site showed that this site displayed cation discrimination properties which were similar to those reported previously for the electrically excitable sodium channel in intact nerve fibers. A possible role for this site in the ion selectivity of the sodium channel is proposed.
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PMID:Properties of the tetrodotoxin binding component in plasma membranes isolated from Electrophorus electricus. 0 13

Erythrocyte ankyrin is a member of a family of proteins that mediate the linkage between membrane proteins and the underlying spectrin-actin-based cytoskeleton. Ankyrin has been shown to interact with a variety of integral membrane proteins such as the anion exchanger, the Na+K(+)-ATPase, and the voltage-dependent sodium channel (NaCh) in brain. To understand how ankyrin interacts with these proteins and maintains its specificity and high affinity for the voltage-dependent NaCh, we have mapped the binding site on ankyrin for the NaCh by examining the binding of purified ankyrin subfragments, prepared by proteolytic cleavage, to the purified rat brain NaCh incorporated into liposomes. 125I-Labeled ankyrin and the radiolabeled 89- and 43-kDa fragments of ankyrin bind to the NaCh with high affinities and with Kd values of 34, 22, and 63 nM, respectively, and have stoichiometries of approximately 1 mol/mol NaCh. The 72-kDa spectrin binding domain is inactive and does not bind to the NaCh. Dissection of ankyrin reveals that the 43-kDa domain retains all the binding properties of native ankyrin to the NaCh. Analysis of the primary structure reveals that the NaCh binding site is confined to a domain of ankyrin consisting entirely of the 11 terminal 33-amino acid repeats and is distinct from the ankyrin domains that interact with spectrin and the Na+K(+)-ATPase.
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PMID:Mapping the binding site on ankyrin for the voltage-dependent sodium channel from brain. 131 4

To maintain alveolar air spaces relatively fluid free, the alveolar epithelium appears capable of vectorial transport of water and solutes. Active transepithelial transport of sodium by alveolar epithelial cell monolayers has previously been demonstrated, indicating that alveolar pneumocytes must possess ion transport mechanisms by which sodium can enter the cells apically for subsequent extrusion via Na(+)-K(+)-adenosinetriphosphatase activity at the basolateral surface. In this study, sodium entry mechanisms were investigated by directly measuring 22Na uptake into rat alveolar epithelial cells grown in primary culture. Cells exhibited increasing 22Na uptake with time over a 30-min interval. Total sodium uptake was compared in the presence and absence of several sodium transport inhibitors. Uptake was inhibited by the sodium channel blockers amiloride and benzamil but was not affected by two amiloride analogues (bromohexamethylene amiloride and dimethylamiloride) with diminished specificity for blocking sodium channels and enhanced specificity for inhibiting the Na(+)-H+ antiporter. Uptake was also unaffected by the chloride transport inhibitor bumetanide or by the absence of glucose. These data suggest that sodium uptake occurs primarily via sodium channel and that Na(+)-H+ antiport, Na(+)-K(+)-2Cl- cotransport, and Na(+)-glucose cotransport do not contribute significantly to sodium uptake under these experimental conditions. The presence of sodium channels in the alveolar epithelial cell membrane may provide the major entry mechanism by which sodium enters these cells for subsequent active extrusion, thereby effecting net salt and water reabsorption from the alveolar spaces.
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PMID:Evidence for amiloride-sensitive sodium channels in alveolar epithelial cells. 131 1

Both atrial natriuretic peptide (ANP) and its receptors are present in the central nervous system, but effects of ANP on brain are unclear. In the present study, we evaluated both the effects of ANP on sodium uptake, and a possible effector mechanism, the putative intracellular second messenger guanosine 3',5'-cyclic monophosphate (cGMP), in rat brain synaptosomes. In the presence of ANP (10(-7) M), the basal level of sodium uptake in synaptosomes was reduced (n = 6) from the control value of 1.90 +/- 0.06 to 1.73 +/- 0.04 (SE) nmol/mg protein at 5 min, P less than 0.05. The observed reduction of sodium uptake by ANP was not influenced by blockade of the other important pathways for sodium uptake. Addition of either a sodium channel blocker (tetrodotoxin) or an inhibitor of Na(+)-K(+)-adenosinetriphosphatase (ATPase) (ouabain) did not affect sodium uptake in the presence of ANP. However, the reduction of sodium uptake was completely blocked by addition of amiloride. These findings suggest that ANP reduced sodium uptake via inhibition of an amiloride-sensitive pathway for sodium uptake. cGMP is a major intracellular second messenger for ANP in other tissues. We found that after stimulation with 10(-7) M ANP, synaptosomal cGMP increased significantly from 58.0 +/- 9.5 to 73.5 +/- 10.6 fmol/mg protein (P less than 0.01). When an analogue of cGMP, 8-bromoguanosine 3',5'-cyclic monophosphate (8-bromo-cGMP), was added to synaptosomes, amiloride-sensitive sodium uptake was again inhibited, by a similar amount as occurred with ANP. It appears that in rat brain, ANP inhibits amiloride-sensitive sodium uptake via a pathway involving intracellular production of cGMP.
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PMID:Atrial natriuretic peptide inhibits amiloride-sensitive sodium uptake in rat brain. 132 15

Previous work has shown that excitatory amino acids inhibit agonist or depolarisation evoked formation of inositol phosphate in brain. In this paper, possible mechanisms by which this may be occurring have been investigated. The inhibition of carbachol-stimulated formation of inositol phosphate by kainic acid (KA) was abolished if the tissue was incubated in a sodium-free medium. The sodium channel activator, veratridine (10 microM) and the sodium ionophore, monensin (3 microM), also inhibited the response of inositol phosphate to carbachol; tetrodotoxin (300 nM) reversed the effect of veratridine but not monensin or KA. Incubation with cadmium (0.3 mM) or removal of extracellular calcium did not alter the effects of KA, monensin or veratridine. The effects of KA were significantly reduced with the Na+/K(+)-ATPase inhibitor, ouabain (10-100 microM). Inhibition by KA was still observed in tissue that had been prestimulated with KA and then washed to remove the agonist. Incorporation of [3H]inositol into inositol lipids was significantly reduced by KA, in the absence or presence of carbachol. It is suggested that the inhibition of the turnover of stimulated phosphoinositide, by excitatory amino acids, is related to the neurotoxic actions of these transmitters and is mediated by Na+ influx, with a consequent activation of Na+/K(+)-ATPase, depletion of cellular ATP and reduction in synthesis of inositol lipid.
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PMID:The inhibition of agonist- or depolarisation-evoked formation of inositol phosphate by excitatory amino acids in rat cerebral cortex is due to the neurotoxic action of this class of neurotransmitter and is mediated by sodium influx. 167 45

The effect of the Na/K-ATPase inhibitor ouabain on phosphoinositide (Ptdlns) hydrolysis was studied in rat brain cortical slices. Ouabain induced a dose-dependent accumulation of inositol phosphates (InsPs) which was much higher in neonatal rats (1570 +/- 40% of basal) than in adult animals (287 +/- 18% of basal). For this reason, all experiments were conducted with 7 day-old rats. Strophantidin caused a similar stimulation of Ptdlns hydrolysis, although it was less potent than ouabain. The order of potency for ouabain-stimulated InsPs accumulation in brain areas was hippocampus greater than cortex greater than brainstem greater than cerebellum. The effect of ouabain was not blocked by antagonists for the muscarinic, alpha1 -adrenergic and glutamate receptors. Also ineffective were the K+ channel blockers 4-aminopyridine and tetraethylammonium, the sodium channel blocker tetrodotoxin, and the calcium channel blocker verapamil, whereas the Na/Ca exchanger blocker amiloride partially antagonized the effect of ouabain. The accumulation of InsPs induced by ouabain was additive to that of carbachol and norepinephrine, as well as to that induced by high K+ and veratrine, but not to that of glutamate. Removal of Na+ ions from the incubation buffer completely prevented the accumulation of InsPs induced by ouabain. The effect of ouabain was also dependent upon extracellular calcium and was under negative feedback control of protein kinase C. Despite the higher effect of ouabain on Ptdlns hydrolysis of immature rats, the density of [3H]ouabain binding sites, as well as the activity of Na/K-ATPase were higher in adult animals. Furthermore, a poor correlation was found between ouabain-stimulated Ptdlns hydrolysis and [3H]ouabain binding in brain regions.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Characterization of ouabain-induced phosphoinositide hydrolysis in brain slices of the neonatal rat. 196 25

The (Na+ + K+)-ATPase from 3T3-F442A fibroblasts and adipocytes was characterized by immunoblotting, ouabain-sensitive rubidium uptake, sodium affinity, and Northern analysis. Using an antibody that cross-reacts with all three forms of the catalytic subunit of the enzyme, it was found that only the alpha 1 isozyme was present in both fibroblasts and adipocytes. This result was confirmed using an antibody specific for the alpha 2 isoform. Additionally, the ouabain dependence of Rb+ uptake in both cell types gave KI values of 0.7-1.0 X 10(-4) M, a concentration that is characteristic for the alpha 1 isoform. For both fibroblasts and adipocytes, the dependence of rubidium uptake activity on sodium concentration was characterized by K0.5 values of 9.4 and 6.2 mM, respectively, which is also diagnostic for the alpha 1 subunit in vivo. Although in fibroblasts there was no detectable message for the alpha 2 isozyme, the 3.4-kilobase message for this isozyme was present in adipocytes; this discrepancy is discussed. The (Na+ (+) K+)-ATPase was activated in fibroblasts and adipocytes by insulin at half-maximal concentrations of 11 nM and about 100 pM, respectively. Glucose uptake was also stimulated at similar concentrations of the hormone. In fibroblasts, insulin caused an increase in sodium uptake which was not inhibited by 1 mM amiloride. From these data, the presence of an insulin-sensitive sodium channel is hypothesized.
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PMID:Characterization of the (Na+ (+) K+)-ATPase from 3T3-F442A fibroblasts and adipocytes. Isozymes and insulin sensitivity. 216 48

1. The kidney taken from a rat rendered nephrotic by exposure to puromycin aminonucleoside retains sodium abnormally when perfused in isolation and has an abnormally low vascular resistance (J. D. Firth et al., Clin. Sci. 1989; 76, 387-95). In this study the relation of oxygen consumption to sodium reabsorption has been examined in the isolated nephrotic organ, which has also been exposed to a variety of natriuretic agents and to the effect of inhibition of metabolism by cooling, in an attempt to discern the transport process, or processes, responsible for abnormal tubular handling of sodium. In addition, the effects of three endogenous vasoconstrictors, noradrenaline, angiotensin II and endothelin, on the function of the isolated nephrotic kidney have been examined. 2. The ratio of mol of sodium reabsorbed by the tubules of the isolated nephrotic kidney to mol of oxygen consumed was reduced in comparison with the control kidney (means +/- SEM): 9.22 +/- 0.97 versus 15.43 +/- 1.55 (P less than 0.002). 3. In the presence of ouabain (1 mmol/l), acetazolamide (1 mmol/l), frusemide (200 mumol/l), the combination of these three agents together, hydroflumethiazide (100 mumol/l), benzamil (100 nmol/l) or atrial natriuretic peptide (1000 pmol/l), a lesser increment in sodium excretion was induced in the isolated nephrotic kidney than in the control kidney and the nephrotic organ continued to excrete less sodium in both absolute and fractional terms. 4. This suggests that enhanced tubular sodium reabsorption in the isolated nephrotic kidney does not depend upon abnormally increased activity of the Na+/K(+)-adenosine triphosphatase, bicarbonate-dependent sodium transport, Na+/K+/2Cl- co-transport, electrically neutral proportionate reabsorption of sodium and chloride (distal tubule), epithelial sodium channel (distal tubule) or atrial natriuretic peptide-sensitive sodium transport processes. 5. When isolated nephrotic kidneys and normal kidneys were cooled to 8-10 degrees C the handling of sodium became virtually identical in the two groups. On re-warming to 37 degrees C, the original differences in sodium handling between nephrotic and control kidneys were restored. This implies that the mechanism responsible for the abnormal tendency to retain sodium is temperature-sensitive; as yet it remains otherwise undefined. 6. The sensitivity of the renal vessels to noradrenaline, angiotension II and endothelin, as judged by the percentage reduction in perfusate flow rate produced by a given concentration of any of these agents, was not substantially altered in the nephrotic kidney compared with the control kidney. Increase in vascular tone was not associated with amelioration of the tendency of the isolated nephrotic organ to retain sodium. Increasing concentrations of angiotensin II caused the filtration rate to increase in the nephrotic kidney. This effect was unexpected: in the control preparation, as anticipated, angiotensin II caused the filtration rate to decrease.
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PMID:Effect of natriuretic agents, vasoactive agents and of the inhibition of metabolism on sodium handling in the isolated perfused kidney of the nephrotic rat. 217 43

Norepinephrine and carbamoylcholine stimulate accumulation of [3H]inositol phosphates from [3H]inositol-labeled guinea pig cerebral cortical synaptoneurosomes through interaction with alpha 1-adrenergic and muscarinic receptors, respectively. In addition to such agonist, a variety of natural products that affect voltage-dependent sodium channels can markedly stimulate accumulation of [3H]inositol phosphates. These include alkaloids that activate sodium channels, such as batrachotoxin, veratridine, and aconitine; peptide toxins that alter activation or slow inactivation of sodium channels, such as various scorpion toxins from Leiurus, Centruroides, and Tityus species; and agents that cause repetitive firing of sodium channel-dependent action potentials, such as pyrethroids and pumiliotoxin B. Ouabain, and agent that will increase accumulation of internal sodium by inhibition of Na+, K+-ATPase, also stimulates formation of [3H]inositol phosphates, as does monensin, a sodium ionophore. Tetrodotoxin and saxitoxin, specific blockers of voltage-dependent sodium channels, prevent or reduce the stimulatory effects of sodium channel agents and ouabain on phosphatidylinositol turnover, while having lesser or no effect, respectively, on receptor-mediated or monensin-mediated stimulation. Removal of extracellular sodium ions markedly reduces stimulatory effects of sodium channel agents, while removal of extracellular calcium ions with EGTA blocks both receptor-mediated and sodium channel agent-mediated phosphatidylinositol turnover. The results provide evidence for a hitherto unsuspected messenger role for sodium ions in excitable tissue, whereby neuronal activity and the resultant influx of sodium will cause activation of phospholipase systems involved in hydrolysis of phosphatidylinositols, thereby generating two second messengers, the inositol phosphates, which mobilize calcium from internal stores, and the diacylglycerols, which activate protein kinase C.
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PMID:Regulation of phosphatidylinositol turnover in brain synaptoneurosomes: stimulatory effects of agents that enhance influx of sodium ions. 242 64


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