Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.3.1 (alkaline phosphatase)
 47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A microvillus plasma membrane-enriched fraction of human placenta was obtained by a combination of differential, isopycnic, and rate-zonal centrifugation techniques. Assays for enzyme markers from mitochondria, lysosomes, peroxisomes, endoplasmic reticulum, and plasma membrane indicated a relative enrichment of plasma membrane between 10- and 20-fold over the most prominent contaminating enzyme markers. Electron microscopy verified the microvillus ultrastructure of the isolated placental membrane and the lack of significant contamination by identifiable organelles. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the microvillus membrane fraction revealed a protein and glycoprotein subunit composition. There were 16 major protein subunits and 10 major glycoprotein subunits, and apparent molecular weights are assigned to these subunits. 32P-labeling of the microvillus membrane-associated alkaline phosphatase indicated that this enzyme is one of the major glycoproteins of the human placental microvillus membrane.
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PMID:The plasma membrane of human placenta. Isolation of microvillus membrane and characterization of protein and glycoprotein subunits. 93 24

1. Alkaline phosphatase (EC 3.1.3.1) from human liver was solubilized from the homogenate using 0.2% Triton X-100 containing 0.2 M lithium 3,5-diiodosalicylate, and the pellet obtained was resolubilized with 20% n-butanol. The procedure resulting in 3842-fold purification included acetone fractionation, ammonium sulfate precipitation, DEAE-cellulose chromatography, Sephadex G-200 gel filtration, hydroxyapatite gel chromatography and further concanavalin A/Sepharose 4B affinity chromatography. 2. The highly purified enzyme showed one major protein band on acrylamide gel electrophoresis at pH 8.6, and exhibited one-seventh of the alkaline p-nitrophenylphosphatase activity in the hepatic enzyme preparation contains of the alkaline pyrophosphatase activity. 3. The highly purified enzyme was a sialic-acid containing glycoprotein. 4. Sialidase-treated hepatic enzyme clearly presented the phenomenon of delayed mobility, and the delayed enzyme fraction stained more strongly than that of non-treated hepatic alkaline phosphatase. 5. In order to investigate the role of the carbohydrate region(s) of the hepatic alkaline phosphatase molecule on substrate binding, the effect of sialidase treatment on the rate of substrate inhibition of alkaline phosphatase was studied. In the case of hepatic enzyme without sialidase, substrate inhibition of alkaline phosphatase activity was clearly shown, while in the case of the hepatic enzyme with sialidase, there was hardly any substrate inhibition in the range of 1-8 mM p-nitrophenylphosphate.
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PMID:Partial purification and some properties of human liver alkaline phosphatase. 94 51

Rat intestinal alkaline phosphatase is an heterogeneous glycoprotein that contains three protein sub-forms separable by electrophoresis. The molecular weight for the glycoprotein (i.e. the average for the three sub-forms) is 157 000-160 000. Three protein sub-forms are detectable on sodium dodecyl sulphate-polyacrylamide gel electrophoresis that migrate at rates corresponding with molecular weights of 64 000, 79 000 and 92 000. Treatment of native alkaline phosphatase with 6 M guanidine - HC1 or buffer at pH 3.0 results in a product with a molecular weight of 78 000 and 70 000, respectively. Thus it is concluded that each of the three sub-forms is a dimer of identical or closely similar subunits. Limited proteolysis results in the production of new enzymically active sub-forms separable by electrophoresis. Using a bacterial protease it is possible to convert intestinal alkaline phosphatase into a form with a molecular weight of 132 000 without causing any significant change in kinetic properties. Electrophoresis of this new form on sodium dodecyl sulphate polyacrylamide gel suggests that it is composed of 66 000-dalton subunits. The native enzyme contains at least 20% by weight of carbohydrate that probably contributes to microheterogeneity of a second degree superimposed on that stemming from the presence of three protein sub-forms. Treatment with various glycosidases has no effect on electrophoretic behaviour, however. It is suggested that the three sub-forms possibly represent different stages of a maturation process that operates by limited proteolysis of a single parent protein.
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PMID:Molecular properties of rat intestinal alkaline phosphatase. 97 5

The spectrum of plasma proteins present in human cortical bone and permanent dentine has been determined. One plasma glycoprotein, the alpha2HS-glycoprotein, was found to be present at a high concentration in both bone and dentine and was shown to be concentrated in the mineralized tissues with respect to the other plasma proteins by factors of between 30 and 100. In this respect the alpha2HS-glycoprotein is analogous to the G2B-glycoprotein and alpha-glycoprotein of bovine and rabbit b one, respectively. The binding of alpha2HS-glycoprotein and albumin to calcium phosphates generated within serum samples has been studied at different serum:precipitate ratios. In each case all the alpha2HS-glycoprotein was removed from the samples and the alpha2HS-glycoprotein was concentrated with respect to albumin by factors ranging from 370 at the highest serum:precipitate ratio to 25 at the lowest ratio. The plasma alpha2HS-glycoprotein concentrations of patients with Paget's disease of bone were shown to be substantially lower than the normal range, with significant negative correlation between the alpha2HS-glycoprotein concentration and the plasma alkaline phosphatase activity.
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PMID:Plasma proteins present in human cortical bone: enrichment of the alpha2HS-glycoprotein. 100 Mar 40

By means of DEAE-Sephadex A-50 column chromatography, Trimeresurus gramineus venom was separated into 12 fractions. Fraction 8 had marked anticoagulant action in the tests of whole blood clotting time, calcium clotting time and plasma prothrombin time. Fraction 8 was rechromatographed on Sephadex G-100, then on DEAE-Sephadex A-50 again, and finally on Sephadex G-100, and a single peak was obtained. The patterns of microzone and disc electrophoresis also showed a single band. A single symmetrical boundary with 1.70 Svedberg units was obtained by ultracentrifugation. The estimated molecular weight was 19 500. The isoelectric point was pH 4.5. Chemical analysis showed that the anticoagulant principle was a glycoprotein and that it was thermolabile. The anticoagulant activity of this purified principle was 3.5 times higher than that of the crude venom. Fraction 5 potentiated its anticoagulant activity to 10 times higher than that of the crude venom. This principle did not possess caseinolytic, tosyl-L-arginine methyl ester esterase, phospholipase A, phosphodiesterase, alkaline phosphomonoesterase, fibrinolytic, hemorrhagic or local irritating activities. The purified anticoagulant principle did not destroy fibrinogen, induce fibrinolysis, inactivate thrombin nor interfere with the interaction between thrombin and fibrinogen. However, a marked inhibition of prothrombin activation was caused by the anticoagulant principle. The inhibition of prothrombin activation was not due to the destruction of prothrombin or its activation factors, but due to an interference in the interaction between prothrombin and its activation factors because of the reversible binding of these factors with the anticoagulant principle of the venom.
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PMID:Purification and properties of the anticoagulant principle of Trimeresurus gramineus venom. 113 81

Kidney alkaline phosphatase was purified to homogeneity. It is a glycoprotein of about 172,000 molecular weight. Analyses of the subunit structure by sedimentation equilibrium in 6 M guanidine hydrochloride and by gel electrophoresis in sodium dodecyl sulfate indicate that the alkaline phosphatase is a dimer comprising two very similar or identical subunits of about 87,000 molecular weight. The native enzyme contains 4.5 +/- 0.2 g atoms of zinc per mol of protein. Reconstitution experiments from the apophosphatase show that binding of 4 Zn2+ per mol of dimer is essential for full activity. The kinetic data of Zn2+ binding to the apoprotein require at least a two-step mechanism, in which one of the steps corresponds to a conformational change within the enzyme. This paper also presents data concerning amino acid composition, sugar content, enzyme stability, absorbance index, and sedimentation velocity.
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PMID:Bovine kidney alkaline phosphatase. Purification, subunit structure, and metalloenzyme properties. 115 Jun 70

A solid-phase assay for the activity of CMPNeuAc:Gal beta 1-4GlcNAc-R alpha-2,6-sialyltransferase (2,6ST) has been developed. In the assay an acceptor glycoprotein is immobilized onto microtiter plate wells. The two glycoprotein acceptors used were asialofetuin (ASF), which contains oligosaccharides terminating in the sequence Gal beta 1-4GlcNAc-R, and a neoglycoprotein of bovine serum albumin containing covalently attached Gal beta 1-4GlcNAc-R units. Samples containing the donor CMPNeuAc and the 2,6ST were incubated with the immobilized acceptor to generate the product NeuAc alpha 2-6Gal beta 1-4GlcNAc-R. The product was detected by a biotin-streptavidin system using the biotinylated plant lectin Sambucus nigra agglutinin (SNA), which binds to sialic acid in alpha-2,6, but not in alpha-2,3, linkage. The biotinylated SNA bound to the product was then detected with streptavidin and biotinylated forms of either alkaline phosphatase or the recombinant bioluminescent protein aequorin. The assay was optimized with respect to the commercially available 2,6ST and shown to be dependent on the concentration of acceptor and CMPNeuAc and proportional to the 2,6ST activity in the range of 20 to 400 microU in a 1-h assay. The solid-phase assay also allows for the selective detection of 2,6ST activity in human and fetal bovine serum, where the activity was proportional in the range of 0.1 to 2 microliters of serum.
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PMID:A solid-phase assay for the activity of CMPNeuAc:Gal beta 1-4GlcNAc-R alpha-2,6-sialyltransferase. 128 7

The extracellular matrix glycoprotein, tenascin, is associated in vivo with mesenchyme undergoing osteogenesis and chondrogenesis, but is absent from mature bone and cartilage matrix. The expression of tenascin by osteoblastic cells in vitro has been investigated by immunoblotting and immunocytochemistry. Tenascin was secreted into the medium and deposited in the matrix by human and rat osteoblast-like cell lines, as well as by primary osteoblast-enriched cultures from chick embryo calvarial bones. In primary osteoblast-enriched cultures, extracellular tenascin was found only in cell aggregates expressing the osteoblast marker alkaline phosphatase. Chicken osteoblast cultures synthesized almost exclusively the largest tenascin subunit, whereas fibroblast cultures from periostea of chicken calvariae synthesized approximately equal amounts of all three subunits. In situ hybridization studies of developing chicken bones, using a cDNA probe that hybridizes to all chicken tenascin splice variants, showed specific labelling of both osteogenic and chondrogenic regions of developing endochondral bones. In contrast, a cDNA probe specific for the large tenascin splice variant showed specific hybridization in osteogenic but not chondrogenic regions. Within osteogenic regions, tenascin mRNA was expressed by osteoblasts. A comparison of in situ hybridization and immunohistochemical studies demonstrated that tenascin mRNA and protein were codistributed in osteogenic regions of endochondral and membrane bones, whereas protein was retained in regions of differentiating cartilage where mRNA was no longer detectable. The results presented here demonstrate that tenascin is synthesized by osteoblasts. Moreover, within developing bones, there are at least three different cell type-specific patterns of expression of tenascin splice variants.
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PMID:Tenascin in bone morphogenesis: expression by osteoblasts and cell type-specific expression of splice variants. 128 16

Saccharomyces cerevisiae contains an amphiphilic cAMP-binding glycoprotein at the outer face of the plasma membrane (M(r) = 54,000). It is converted to a hydrophilic form by treatment with glycosyl-phosphatidylinositol-specific phospholipases C and D (GPI-PLC/D), suggesting membrane anchorage by a covalently bound glycolipid. Determination of the constituents of the purified anchor by gas-liquid chromatography and amino acid analysis reveals the presence of glycerol, myo-inositol, glucosamine, galactose, mannose, ethanolamine, and asparagine (as the carboxyl-terminal amino acid of the Pronase-digested protein to which the anchor is attached). Complementary results are obtained by metabolic labeling, indicating that fatty acids and phosphorus are additional anchor constituents. The phosphorus is resistant to alkaline phosphatase, whereas approximately half is lost from the protein after treatment with GPI-PLD or nitrous acid, and all is removed by aqueous HF indicating the presence of two phosphodiester bonds. Inhibition of N-glycosylation by tunicamycin or removal of protein-bound glycan chains by N-glycanase or Pronase does not abolish radiolabeling of the anchor structure by any of the above compounds. Analysis of the products obtained after sequential enzymic and chemical degradation of the anchor agrees with the arrangement of constituents in GPIs from higher eucaryotes. Evidence for anchorage of the yeast cAMP-binding protein by a GPI anchor is strengthened additionally by the reactivity of the GPI-PLC-cleaved anchor with antibodies directed against the cross-reacting determinant of trypanosomal variant surface glycoproteins.
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PMID:The cAMP-binding ectoprotein from Saccharomyces cerevisiae is membrane-anchored by glycosyl-phosphatidylinositol. 133 92

Resistance to several cytotoxic agents, including anthracyclines, vinca alkaloids and epipodophylline derivatives (multidrug resistance, or MDR) can develop in tumor cells by overexpression of a 170-kd glycoprotein (p170) which is an essential component of a membrane transport system leading to increased drug efflux and decreased intracellular drug concentration. By means of a p170-directed monoclonal antibody (MRK-16) and immunocytochemistry (alkaline phosphatase anti-alkaline phosphatase technique), we investigated the expression of p170 in marrow blast cells of 59 cases (38 at diagnosis and 21 in relapse) of acute-non-lymphocytic leukemia (ANLL). The proportion of strongly MDR-positive cells was higher in relapse that at diagnosis (median 15.5% vs 1.5%). Out of 31 patients who were evaluable for the results of first remission induction, failure of first-line treatment (including Daunorubicin, standard-dose and high-dose Arabinosyl Cytosine, and sometimes also Mitoxantrone) occurred in 8/22 MDR-positive cases and in 1/9 MDR-negative ones (p = 0.21). Failure of first-line treatment was always associated with a progressive increase of p170 expression. Total failures (no remission plus early relapse) were more frequent (p = 0.001) among MDR-positive cases (16/22) than among the others (2/9). These data show that MDR is very frequent in ANLL also at diagnosis and suggest that MDR can contribute to early failure of standard treatment.
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PMID:Overexpression of multidrug resistance-associated p170-glycoprotein in acute non-lymphocytic leukemia. 134 49


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