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)

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

The membrane-associated V-ATPase that plays an important role in the regulation of acid-base balance by the kidney is a multisubunit enzyme that is densely packed into specialized membrane domains in intercalated cells. Intercalated cells can be separated into at least two subtypes, A-cells and B-cells, based on their morphological features, the distribution of V-ATPase, and the presence or absence of a basolateral chloride/bicarbonate anion exchanger (AE1) exclusively in B-cells. A-cells secrete protons into the tubule lumen, whereas B-cells secrete bicarbonate. The relative amounts of V-ATPase and AE1 in the plasma membranes of A- and B-cells are modulated under different acid-base conditions and provide a sensitive means by which urinary acidification can be controlled. The mechanisms governing the movement of acid-base transporting proteins between intracellular vesicles and the plasma membrane are under investigation. The microtubular apparatus of the cell is involved in maintaining both apical and basolateral polarity of the enzyme, and different isoforms of V-ATPase subunits may also be involved in the selective targeting of V-ATPase to different membrane domains.
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PMID:Polarized targeting of V-ATPase in kidney epithelial cells. 149 Dec 26

Two major types of intercalated cells (IC) have been previously defined in rabbit collecting duct: alpha-cells have a basolateral band 3-like anion exchanger and secrete H+, whereas beta-cells bind peanut agglutinin (PNA) apically and are believed to secrete HCO3-. To further define IC types, we double-labeled kidney sections with anti-H(+) -ATPase antibodies and with either an anti-band 3 antibody or PNA. We found four patterns of staining: 1) IC with apical H(+)-ATPase and basal band 3, a configuration consistent with ongoing H+ secretion, which prevailed in the inner stripe of outer medulla (OMCDi); 2) diffuse H(+)-ATPase labeling across the cell and basal band 3, which was most numerous in the outer stripe of outer medulla (OMCDo); 3) IC with "bright" apical peanut lectin, diffuse H(+)-ATPase, and no band 3, which was abundant in the cortical collecting duct (CCD) and probably represents HCO3(-)-secreting cells; and 4) "hybrid" cells with various staining combinations (e.g., apical lectin binding and apical H(+)-ATPase), which although they are uncommon, were seen in the CCD. Consistent with this immunocytochemical finding of hybrid cells, cell-sorting studies on isolated CCD IC showed that 6-18% of PNA-positive cells also stained positively for band 3. We conclude that 1) band 3-positive IC in the OMCD vary axially. Most OMCDi IC are probably active proton secretors, whereas up to one-half of OMCDo IC may be latent H+ secretors. 2) The diffuse H(+)-ATPase pattern in putative beta-cells differs from comparable results in the rat and is not consistent with a "reversed" alpha-cell. HCO3- secretion by beta-cells may be driven by an H+ extrusion mechanism other than the alpha-cell pump re-sorted to the basolateral membrane. 3) The possibility of hybrid cells that might combine alpha- and beta-cell transport proteins suggests a mechanism for functional reversal of collecting duct IC polarity.
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PMID:Colocalization of H(+)-ATPase and band 3 anion exchanger in rabbit collecting duct intercalated cells. 184 62

The F1 portion of H(+)-translocating ATPase as purified from membrane vesicles of Vibrio parahaemolyticus by a rapid procedure. The whole purification process (from culture of cells to purification of the enzyme) could be completed in 1 day. The F1-ATPase consists of five subunits (alpha, beta, gamma, delta and epsilon) like F1 of Escherichia coli and other microorganisms. The F1-ATPase of V. parahaemolyticus showed some interesting properties. Its activity was greatly stimulated by high concentrations (about 0.5 M) of SO4(2-), SO3(2-) and CH3COO-, their effects decreasing in this order. Among the anions tested, Cl- and NO3- were ineffective, or rather inhibitory, and cations had no significant effects. Ethanol (or methanol) stimulated the activity 2- to 3-fold. The activity was inhibited by 4-acetamido-4'-isothiocyanostilbene 2,2'-disulfonate (SITS) (an anion exchanger inhibitor), tetrachlorosalicylanilide (TCS) (an H+ conductor), azide and N-ethylmaleimide. Zinc inhibited the activity only slightly, although it strongly inhibited the ATPase activity in membrane vesicles.
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PMID:Rapid purification and characterization of F1-ATPase of Vibrio parahaemolyticus. 214 93

This report demonstrates that the high affinity binding of ankyrin to two well characterized ankyrin-binding proteins, the erythrocyte anion exchanger and kidney Na+K(+)-ATPase, requires interaction of these proteins with unique sites on the ankyrin molecule. Binding of 125I-labeled erythrocyte ankyrin and ankyrin proteolytic domains was measured to the anion exchanger and Na+K(+)-ATPase incorporated into phosphatidylcholine liposomes. 125I-Labeled ankyrin associated with both anion exchanger and Na+K(+)-ATPase liposomes with a high affinity (KD ranging from 10 to 25 nM), and a capacity approaching 1 mol of ankyrin/2 mol of ATPase and 1 mol of ankyrin/8 mol of anion exchanger. The 43 kDa cytoplasmic domain of the erythrocyte anion exchanger inhibited binding of ankyrin to both the anion exchanger and Na+K(+)-ATPase liposomes with a 50% reduction at approximately 90 nM for both proteins. Further binding experiments using proteolytic domains derived from ankyrin demonstrated the following differences between the anion exchanger and Na+K(+)-ATPase in interactions with ankyrin: 1) 125I-Labeled Na+K(+)-ATPase associated with both the 89-kDa domain as well as the spectrin binding domain of ankyrin, while the anion exchanger only associated with the 89-kDa domain. 2) The 125I-labeled 89-kDa domain of ankyrin associated with Na+K(+)-ATPase liposomes with at least a 20-fold lower affinity compared with intact ankyrin while this domain associated with the anion exchanger with a 2-3-fold increase in affinity compared with intact ankyrin. 3) The 125I-labeled spectrin-binding domain of ankyrin associated with the Na+K(+)-ATPase liposomes to at least an 8-fold greater extent than to anion exchanger liposomes. The data are consistent with an independent acquisition of high affinity ankyrin binding activity for the anion exchanger and Na+K(+)-ATPase proteins through a convergent evolutionary process.
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PMID:The anion exchanger and Na+K(+)-ATPase interact with distinct sites on ankyrin in in vitro assays. 217 Mar 69

When parietal cells (PC) are stimulated with histamine, the anion exchanger rate increases three to five times to compensate for alkaline loading induced by H+-K+ adenosinetriphosphatase (ATPase) and to provide Cl for acid secretion. It has been hypothesized that this increased activity is caused by the increase in intracellular pH (pHi) that often occurs in stimulated PC (from 7.1 to a maximum of 7.3). The dependence of the anion exchanger on pHi was studied using microspectrofluorimetry of the pH-sensitive dye 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF). N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES)-buffered solutions were used because the anion exchanger can transport OH- (or HCO3) in exchange for Cl- even with [HCO3]o = 200 microM. It was found that when solutions were changed either from NaCl to Cl- free or Cl- free to NaCl, rates of change of pHi (delta pH/delta t) were strongly dependent on pHi: nearly 0 at pHi 6.6 and 1.25 pH/min at pHi 8.0. To convert these pHi changes into anion flux rates, the intrinsic buffer capacity (beta i) was determined over the same pHi range by making small changes of [NH4]o to determine the resulting changes of [NH4]i and pHi (i.e., beta i = delta[NH4]i/delta pHi) in PC that had been pretreated with 1 mM amiloride and 200 microM [H2]4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) [to block Na+-H+ and Cl- -OH-(HCO3-) exchange]. beta i was also strongly dependent on pHi: at pHi 6.5 beta i = 48 mM/pH, and this decreased as pHi increased; at pHi 7.75 beta i = 8 mM/pH. The derived anion fluxes (i.e., JOH = beta i x delta pH/delta t) were roughly linearly related to pHi between 6.6 (JOH near 0) and 8.1 (JOH = 13 mM/min). Between pHi 7.1 and 7.3, the range normally observed during stimulation of PC, rates of anion exchange increased by 75%. This pHi sensitivity cannot explain the 300-500% increase in anion exchanger activity observed during secretagogue stimulation of PC.
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PMID:Intracellular pH dependence of buffer capacity and anion exchange in the parietal cell. 255 41

Interactions between integral proteins of the plasma membrane and the cytoskeleton may be important for localizing certain membrane proteins in a nonrandom fashion at specialized domains of the cell surface. Here, we show that ankyrin, the key protein for the linkage of the erythrocyte anion exchanger (band 3) to the spectrin-based membrane cytoskeleton, is also present in kidney distal tubular cells where ankyrin is precisely colocalized with Na+,K+-ATPase. Both proteins are confined to the basolateral plasma membrane and are absent from the apical membrane, the junctional complex and the membrane surface that contacts the basal lamina. Purified Na+,K+-ATPase of sheep and pig kidney contains a binding site for erythrocyte ankyrin as demonstrated by immunoprecipitation experiments. A band 3-like binding site for ankyrin is likely, since binding of ankyrin to Na+,K+-ATPase could be inhibited in a competitive fashion by the isolated cytoplasmic domain of erythrocyte band 3.
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PMID:Colocalization and coprecipitation of ankyrin and Na+,K+-ATPase in kidney epithelial cells. 283 37

Disulphonic stilbenes are effective inhibitors of an anion exchanger which is present in the plasma membranes of many cells (Cabantchik et al. 1978). In the present study, the effects of 4,4'-diisothiocyano-2,2'-disulphonic stilbene acid (DIDS) on the transport activity of the hydrochloric acid pump isolated from pig stomach (H,K-ATPase, EC 3.6.1.36) were tested. Half-maximal inhibition of proton transport carried out by the H,K-ATPase in the isolated vesicles was observed at micromolar concentrations of DIDS. The effects of DIDS on the adenosine-trisphosphatase and p-nitrophenyl-phosphatase activities of isolated H,K-ATPase were also studied and compared with those of the kinetically and structurally related Na,K-ATPase (EC 3.6.1.37). Half-maximal inhibition of the enzymatic activities of both enzymes were observed in the micromolar range of DIDS. The lipid bilayer of the gastric vesicle membrane is highly asymmetric and the original cytosolic side is facing the outside of the vesicle. Since DIDS does not readily cross the membrane, it is most likely that DIDS exerts its inhibitory effects by modifying the transport ATPases on their cytosolic sides.
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PMID:Inhibition of H,K-ATPase and Na,K-ATPase by DIDS, a disulphonic stilbene derivative. 285 43

1. Models are presented for (a) HK ATPase acting in the presence of K and Cl conductances; (b) a pH regulatory system where Na/H exchange is regulated directly by second messenger and the anion exchanger is activated secondarily to the rise in cell pH; (c) vesicle fusion and K and Cl conductances activation in the gastric parietal cell. 2. It is suggested that H transport involves protonation and deprotonation of histidine groups as well as the motion of these groups relative to the membrane barrier. 3. The HK ATPase would have a voltage generating and voltage sensitive step in the forward direction. 4. Given net electroneutrality the K transport reaction would also be charge translocating and voltage sensitive.
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PMID:Passive and active transport in the parietal cell. 290 80

Fast-performance liquid chromatography was used to purify assembly-competent tubulin from porcine brain microtubule protein prepared by two cycles of assembly-disassembly. Microtubule protein (1-100 mg at 1.5-2.5 mg/ml) in buffer consisting of 0.1 M 2-(N-morpholino)ethanesulfonic acid, 0.5 mM MgCl2, 1 mM EGTA, 0.3 M KCl, and 0.02 mM GTP (pH 6.6) was applied to the Mono Q column (anion exchanger). The microtubule-associated proteins, GTP and GDP, eluted in the void volume. The tubulin fraction eluted at 0.45-0.50 M KCl with 65-80% recovery. The tubulin fraction contained trace enzymatic activities when compared with the starting microtubule protein, i.e., less than 1 versus 60 mU/mg/min of nucleoside diphosphate kinase, 0.2 versus 7.0 nmol/mg/min of Mg-ATPase at pH 6.6, and 0.2 versus 88 mU/mg/min of adenylate kinase. Both the Mono Q-purified tubulin and the pelleted microtubules that were assembled in 0.5 mM [3H]GTP contained 0.77 mol of labeled nucleotide/tubulin dimer. The Mono Q-purified tubulin fraction was competent to assemble, i.e., the critical concentration was 0.1 mg/ml in the presence of 0.03 mM taxol and 1 mM GTP at 37 degrees C. The Mono Q-purified tubulin fraction showed trace high-molecular-weight components, which were removed on Mono S (cation exchanger) columns. Alternatively, microtubule protein in buffer was applied to the Mono S column. Tubulin, trace nontubulin proteins, and several enzymatic activities came off in the void volume. A combination of Mono Q-Mono S or Mono S-Mono Q chromatography resulted in highly purified protein.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Separation of assembly-competent tubulin from brain microtubule protein preparations using a fast-performance liquid chromatography procedure. 300 70


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