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)

Resistance of tumors to a variety of chemotherapeutic agents presents a major problem in cancer treatment. The gene responsible for multidrug resistance, termed mdr1, encodes a membrane glycoprotein (P-glycoprotein) that acts as a pump to transport various cytotoxic agents. The P-glycoprotein has been shown to bind anticancer drugs and several resistance-reversing agents including calcium channel blockers, and to be an ATPase. The P-glycoprotein was found to function in the blood-brain barrier. The physiological function of the P-glycoprotein in relation to therapy is discussed.
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PMID:[Mechanism of resistance to antitumor agents--its involvement in blood-brain barrier]. 756 98

Resistance of tumors to a variety of chemotherapeutic agents presents a major problem in cancer treatment. The gene responsible for multidrug resistance, termed mdr1, encodes a membrane glycoprotein (P-glycoprotein) that acts as a pump to transport various cytotoxic agents. The P-glycoprotein has been shown to bind anticancer drugs and several resistance-reversing agents including calcium channel blockers, and to be an ATPase. The P-glycoprotein was found to function in blood-brain barrier. The implication of the P-glycoprotein in relation to therapy is discussed.
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PMID:[Structure and function of P-glycoprotein in antitumor agent resistance; implication for clinical setting]. 853 80

Multidrug resistance (MDR) to chemically unrelated therapeutic anticancer agents in mammalian cells is mediated by the overexpression of an ATP-dependent 150- to 180-kD membrane glycoprotein P-glycoprotein (P-gp). Although the complete physiological role of P-gp is unknown, it is proposed to function in cellular detoxification of xenobiotics. In this study, we investigated whether the organophosphorus insecticide chlorpyrifos (O,O-diethyl O-3,5,6-trichloro-2-pyridinyl phosphorothioate) or its metabolites interact with P-gp. Immunohistochemical analysis of tissues from male Fischer 344 rats administered chlorpyrifos (7.6 mg/kg gavage) showed increased P-gp expression in the kidney, adrenal, liver, jejunum, and stomach (tissues associated with elimination of xenobiotics), compared to control tissues. The most prominent increase was detected in the large bile ducts of the liver and the proximal tubule region of the kidney. P-gp expression was increased throughout the adrenal medulla and cortex, while a moderate increase was detected in the epithelial layers of the stomach and jejunum. To examine further the interaction between chlorpyrifos and P-gp, we evaluated whether chlorpyrifos or its active metabolite, chlorpyrifos oxon, could inhibit [3H]azidopine labeling of P-gp in MDR1 baculovirus-infected insect Sf9 cells. A concentration-dependent inhibition of [3H]azidopine labeling of P-gp was detected with chlorpyrifos oxon, while significant inhibition was not detected with chlorpyrifos. To correlate the binding of chlorpyrifos oxon to P-gp with a biochemical effect, we examined its ability to stimulate P-gp-mediated ATPase activity in these Sf9 cells. Chlorpyrifos oxon stimulated P-gp ATPase activity 1.75 times that of the positive control (10 microM verapamil). Taken together, these results suggest that chlorpyrifos oxon interacts with P-gp, and support the hypothesis that P-gp may play a role in the cellular detoxification of insecticides in mammalian tissues. To our knowledge this is the first report of an organophosphorus insecticide interacting with and increasing the expression of P-gp.
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PMID:Chlorpyrifos oxon interacts with the mammalian multidrug resistance protein, P-glycoprotein. 860 Feb 91

Hepatocyte plasma membranes contain a glycosylated 230-kDa Ca(2+) -dependent, Mg(2+)-stimulated ATPase (pgp230), which consists of two subunits, one of 120 kDa and the other of 110 kDa. pgp230 can be enriched by the use of affinity chromatography on Concanavalin A-Sepharose, wheat germ lectin-Sepharose, and 5'-AMP-Sepharose. It has a high-affinity Ca2+ binding site. In the presence of Ca2+, it forms a phosphorylated intermediate by autocatalytic transfer of the terminal phosphate residue from ATP. Maximal Ca(2+)-dependent autophosphorylation is observed at pH 5-6. Photoaffinity labeling using 8-azido-[alpha-32P]ATP or [y-32P]ATP confirms the presence of ATP binding sites. Incubation with [alpha-32P]ATP leads to a rapid but transient labeling of pgp230. Various nucleotides, nucleotide receptor agonists, or antagonists inhibit Ca(2+)-dependent phosphorylation by [y-32P]ATP. The concentrations of half-maximal inhibition range from 10(-7) M to 10(-3) M. The rank order of inhibitory potency is: ATP > alpha,beta-methylene-ATP > CTP = TTP > y-4-amino-phenyl-ATP = 2-methyl-thio-ATP > UTP = GTP > GDP = ADP = beta,y-methylene-ATP = beta, y-methylene-TTP = beta,y-methylene-GTP = adenosine-5'-O-2-thiodiphosphate = CMP = AMP > adenosine > cytidine > guanosine = suramin > Reactive blue 2 > iso-butyl-methyl-xanthine > thymidine > uridine. These data suggest a nucleotide binding capacity of this new hepatocyte membrane glycoprotein. Further investigations should be carried out to reveal its biological function.
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PMID:Partial characterization of a new nucleotide binding glycoprotein of hepatocyte plasma membrane. 878 41

The liver converts endogenous and xenobiotic lipophilic compounds into anionic conjugates with glutathione, glucuronate, or sulfate. These conjugates are transported across the canalicular (apical) membrane into bile by a 190 kDa membrane glycoprotein that has been cloned recently. This apical conjugate-transporting ATPase has been termed canalicular multidrug resistance protein (cMRP) because of the similarity in substrate specificity and sequence with the multidrug resistance protein (MRP1), canalicular multispecific organic anion transporter (cMOAT), or multidrug resistance protein 2 (MRP2). The amino acid sequence identity of human MRP2 and MRP1 is 49%. MRP2 is predominantly expressed in hepatocytes and localized to apical membrane domains. MRP2 is not expressed in the human Dubin-Johnson syndrome, which is therefore associated with an inherited deficiency in the secretion of amphiphilic anionic conjugates into the bile. The rat homolog Mrp2 is absent in two mutant strains of rats with different point mutations in the corresponding gene. These mutant rats are hyperbilirubinemic and deficient in the ATP-dependent transport of conjugates from hepatocytes into bile. Impairment of bile flow (cholestasis) can be associated with a down-regulation of the expression of the conjugate export pump, and MRP2 contributes to bile flow as an important driving force.
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PMID:Hepatic canalicular membrane 5: Expression and localization of the conjugate export pump encoded by the MRP2 (cMRP/cMOAT) gene in liver. 921 74

We have examined the in vivo localization of extracellular ecto-ATPase and ecto-apyrase (ATPDase) in adult chicken gizzard and stomach by immunofluorescence and laser scanning confocal microscopy. In chicken gizzard, the ecto-ATPase was distributed in discrete clusters restricted to the sarcolemma of the smooth muscle cells. Anti-ecto-apyrase antibody detected a single 80-kDa band (putative apyrase) in Western blots of both chicken gizzard membrane extracts and partially purified anion exchange fractions, but the antibody did not detect ecto-apyrase in immunolabeled gizzard cryosections. In adult chicken stomach, the ecto-apyrase was observed at the apical membrane of the glandular oxyntico-peptic cells as described in previous immunoperoxidase studies (Stout, J. G., R. S. Strobel, and T. L. Kirley (1995) Biochem. Mol. Biol. Int. 36, 529-535). However, ecto-ATPase was clustered in the sarcolemma of the organized layer of circular smooth muscle and in smooth muscle cells of the septa surrounding the glandular tissue, but not in the glandular cells containing the ecto-apyrase. The findings indicate compartmentalization of the two related extracellular nucleotide hydrolyzing enzymes and suggest differential functions that are specialized for different regions of the chicken stomach. We also partially purified the ecto-apyrase of chicken stomach, an 80-kDa membrane glycoprotein. Chicken stomach membranes were solubilized in digitonin, glycoproteins were separated from solubilized proteins by lectin chromatography, and nucleotide-binding glycoproteins were selected by immobilized Cibacron blue chromatography. Further purification by size exclusion and anion exchange chromatography yielded purification of 94-fold. The ATPase specific activity of the purified stomach ecto-apyrase was 75,000 micromol of Pi/mg of protein/h, and the purified preparation consisted of a major band (55% of total protein) at 80 kDa. The purified enzyme could be deglycosylated with peptide N-glycosidase-F to a core molecular mass of 54 kDa. The N-terminal sequence of the 80-kDa stomach ecto-apyrase band (which reacted with anti-ecto-ATPDase antibodies) was determined to be: MEYKGKVVAGLLTATWV. Immunological cross-reactivity data indicate that the stomach 80-kDa protein isolated is an ecto-apyrase and is related to both the chicken liver and oviduct ecto-ATPDase enzymes characterized earlier, as well as to the human lymphoid cell activation antigen, CD39.
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PMID:Immunolocalization of the ecto-ATPase and ecto-apyrase in chicken gizzard and stomach. Purification and N-terminal sequence of the stomach ecto-apyrase. 929 5

1. P-glycoprotein, a 170-180 kDa membrane glycoprotein that mediates multidrug resistance, hydrolyses ATP to efflux a broad spectrum of hydrophobic agents. In this study, we analysed the effects of three MDR reversing agents, verapamil, cyclosporin A and [3'-keto-Bmt1]-[Val2]-cyclosporin (PSC 833), on the adenosine triphosphatase (ATPase) activity of human P-glycoprotein. 2. P-glycoprotein was immunoprecipitated with a monoclonal antibody (MRK-16) and the P-glycoprotein-MRK-16-Protein A-Sepharose complexes obtained were subjected to a coupled enzyme ATPase assay. 3. While verapamil activated the ATPase, the cyclosporin derivatives inhibited both the substrate-stimulated and the basal P-glycoprotein ATPase. No significant difference was observed between PSC 833 and cyclosporin A on the inhibition of basal P-glycoprotein ATPase activity. PSC 833 was more potent than cyclosporin A for the substrate-stimulated activity. 4. Kinetic analysis indicated a competitive inhibition of verapamil-stimulated ATPase by PSC 833. 5. The binding of 8-azido-[alpha-32P]-ATP to P-glycoprotein was not altered by the cyclosporin derivatives, verapamil, vinblastine and doxorubicin, suggesting that the modulation by these agents of P-glycoprotein ATPase cannot be attributed to an effect on ATP binding to P-glycoprotein. 6. The interaction of the cyclosporin derivatives with ATPase of P-glycoprotein might present an alternative and/or additional mechanism of action for the modulation of P-glycoprotein function.
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PMID:Interaction of cyclosporin derivatives with the ATPase activity of human P-glycoprotein. 931 31

In open monolayers of epithelial cells grown in vitro, the apical membrane domain forms on the free cell surface that faces the culture medium. However, in vivo, the apical lumenal compartment arises within groups of cells that do not have a free cell surface. We designed in vitro culture conditions, using small colonies of MDCK cells overlaid with collagen, in which formation of the apical membrane must occur de novo by remodeling existing membrane domains that are contacted by other cells or extracellular matrix. Within 12 hours of collagen overlay, the apical membrane glycoprotein gp135 is removed from the free cell surface, while lateral membrane proteins (e.g. Na+,K+-ATPase) remain at sites of cell-cell contacts. Subsequently, lumenal structures, containing gp135 and the apically secreted protein gp81, formed within these cell-cell contacts. Na+,K+-ATPase, adherens junction (E-cadherin, alpha- and beta-catenins) and tight junction (ZO-1) proteins were localized on the lateral membrane adjacent to, but excluded from the gp135-positive lumenal compartment. Therefore, each lumen represents a newly formed apical compartment on the lateral membrane. The Golgi complex (alpha-mannosidase II and Golgi beta-spectrin), centrosomes (gamma-tubulin) and microtubules reorient to a cytoplasmic position adjacent to the newly-forming apical lumenal compartments. Significantly, addition of colchicine, nocodazole or brefeldin A inhibits apical lumen formation. These results demonstrate that simple epithelial cells form an apical lumenal compartment de novo through initial intermixing, and then sorting of apical and basal-lateral membrane proteins at sites of cell-cell contacts. In addition, apical lumen formation requires an intact microtubule network, microtubule-dependent reorientation of the Golgi complex and secretory apparatus, and fully functional protein delivery from the Golgi complex to the forming apical cell surface.
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PMID:Mechanisms for de novo biogenesis of an apical membrane compartment in groups of simple epithelial cells surrounded by extracellular matrix. 942 87

Employing antisera against various subfractions of rat liver mitochondria (mitoplast, inner membrane, intermembrane, and matrix) as well as metabolically radiolabeled BRL-3A rat liver cells, we undertook a search for the presence of glycoproteins in this major cellular compartment for which little information in regard to glycoconjugates was available. Subsequent to [35S]methionine labeling of BRL-3A cells, a peptide:N-glycosidase-sensitive protein (45 kDa) was observed by SDS-polyacrylamide gel electrophoresis of the inner membrane immunoprecipitate, which was reduced to a molecular mass of 42 kDa by this enzyme. The 45-kDa protein was readily labeled with [2-3H]mannose, and indeed the radioactivity of the inner membrane immunoprecipitate was almost exclusively present in this component. Moreover, antisera directed against mitochondrial NADH-ubiquinone oxidoreductase (complex I) or F1F0-ATPase (complex V) also precipitated a 45-kDa protein from BRL-3A cell lysates as the predominant mannose-radiolabeled constituent. Endo-beta-N-acetylglucosaminidase completely removed the radiolabel from this glycoprotein, and the released oligosaccharides were of the partially trimmed polymannose type (Glc1Man9GlcNAc to Man8GlcNAc). Cycloheximide as well as tunicamycin resulted in total inhibition of radiolabeling of the inner membrane glycoprotein, and moreover, pulse-chase studies employing metrizamide density gradient centrifugation demonstrated that the glycoprotein was initially present in the endoplasmic reticulum (ER) and subsequently appeared in a mitochondrial location. Early movement of the glycoprotein to the mitochondria after synthesis in the ER was also evident from the limited processing undergone by its N-linked oligosaccharides; this stood in contrast to lysosomal glycoproteins in which we noted extensive conversion to complex oligosaccharides. Our findings suggest that the 45-kDa glycoprotein migrates from ER to mitochondria by the previously observed contact sites between the two organelles. Furthermore, the presence of this glycoprotein in at least two major mitochondrial multienzyme complexes would be consistent with a role in mitochondrial translocations.
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PMID:Identification of a glycoprotein from rat liver mitochondrial inner membrane and demonstration of its origin in the endoplasmic reticulum. 967 1

The polarized rat hepatoma/human fibroblast hybrid cell line, WIF-B, forms apical vacuoles into which cholephilic substances are secreted. We studied expression, localization, and function of the apical conjugate export pump, Mrp2, in WIF-B cells. Mrp2, the apical isoform of the multidrug resistance protein, alternatively termed canalicular Mrp (cMrp) or canalicular multispecific organic anion transporter (cMoat), is a 190-kd membrane glycoprotein mediating adenosine triphosphate (ATP)-dependent transport of glucuronides, glutathione S-conjugates, and other amphiphilic anions across the hepatocyte canalicular membrane into bile. Expression of the rat mrp2 gene in WIF-B cells was shown by reverse-transcription polymerase chain reaction (PCR), followed by sequencing of the amplified 789-bp fragment. Immunoblotting, using antibodies reacting with the amino-terminal or with the carboxyl-terminal sequence of rat Mrp2, detected the 190-kd glycoprotein in WIF-B cell homogenates. Immunofluorescence microscopy localized Mrp2 to the apical membrane domain. Preloading of WIF-B cells with a membrane-permeable ester of the calcium-dependent fluorescent indicator, Fluo-3, was followed by Mrp2-mediated secretion of the amphiphilic anion, Fluo-3, into the apical vacuoles. This transport was potently inhibited by cyclosporin A added to the culture medium. Direct measurements of ATP-dependent transport into Mrp2-containing plasma membrane vesicles in comparison with Mrp2-deficient vesicles established that Fluo-3 is transported by Mrp2 with a Km value of 3.7 micromol/L. Our results indicate that the polarized WIF-B cells express the rat ortholog of the apical conjugate-transporting ATPase, Mrp2. The function of Mrp2 as well as the action of inhibitors can thus be analyzed by use of the fluorescent amphiphilic anion, Fluo-3.
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PMID:Expression of the apical conjugate export pump, Mrp2, in the polarized hepatoma cell line, WIF-B. 979 19


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