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)

Human placental microsomal 5'-nucleotidase (EC 3.1.3.5) was prepared free of alkaline phosphatase by isoelectric focusing. A total of seven electrophoretic variants were isolated during the preparation of six placentas. Only three to six variants were found in a single placenta. The isoelectric pH's were 6.70, 6.44, 6.23, 6.02, 5.76, 5.63 and 5.44. These were found to be composed of variable quantities of a large, medium and low molecular weight form. The apparent molecular weights of the medium and light form of the enzyme were 86 500 and 43 500, respectively, as estimated from Stokes radius and sedimentation velocity determinations. The electrophoretic variants were not distinguishable with respect to specific activity and Michaelis constants for AMP, GMP or CMP or inhibition by ATP, CTP or adenosine. These electrophoretic variants appeared to be pseudoisozymes based upon different states of aggregation of a common primary sequence. There was a wide range of substrate specificity among nucleoside 5'-monophosphates which included 2-deoxyribose compounds. With AMP as 100, substrate activity was: CMP, 122; NMN, 74; GMP, 68: IMP, 63; XMP, 28 and UDP-glucose, 68. The Michaelis constants for AMP, GMP and CMP ranged from 12-18 muM, from 33-67 muM and from 170-250 muM, respectively. Although 5'-nucleotidase was active in the absence of divalent cation, 5 mM MgCl2 stimulated the enzyme activity to 234% of control and shifted the pH optimum of 9.8 to a plateau from pH 7.4-9.8.
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PMID:Purine catabolism in man: characterization of placental microsomal 5'-nucleotidase. 0 35

The aims of this study were to determine the effect of levels of various substances and reaction by-products, which are formed during hydrolysis of nucleic acids, on the derivatization and chromatography of nucleosides; and to investigate the silylation of mono- and dinucleotides. The effect of NaCl, KCl, MgCl2, NH4Cl, and (NH4)2SO4 on silylation and chromatography of nucleosides was studies at various molar excesses of salt. The response values for all nucleosides were studied at various molar excesses of salt. The response values for all nucleosides were significantly affected at molar excess salt present values (MSP) between 1 and 10 for KCl, NaCl, NH4Cl, (NH4)2SO4 and between 0.1 and 1 for MgCl2. It was noted that thymidine was more sensitive than other nucleosides if silylated in presence of these salts. Two chromatographic peaks at retention temperatures (RT) 240 and 251 were obtained for cytidine at MSP values of 10(-3) for NaCl, KCl, and MgCl2, and 10(-4) for NH4Cl and (NH4)2SO4. In a mixture of nucleosides the RT = 251 peak was used for quantitative analysis of cytidine as the RT = 240 peak elutes with guanosine. Thus, these salts have a significant effect on the gas-liquid chromatography of trimethylsilyl (TMS) cytidine in a mixture of nucleosides, especially the RT = 241 peak. The effect of salts on derivatization can be explained in part as follows: (a) reduced derivatization of nucleosides due to a decreased solubility in the solvent system; (b) formation of TMS anion derivatives, e.g. TMS-SO4, TMS-PO4, with a reduced molar excess of BSTFA; (c) metal chelation by Mg ions or other divalent cations with nucleosides or BSTFA; and/or (d) an increased breakdown of TMS derivatives in presence of salt in the sample or on the top 3 in. of the column packing. Also, experiments were made on the effect of other substances such as Tris, phosphate, alkaline phosphatase, and KCl on completeness of silylation. The individual impurities showed no significant effect on the relative weight response (RWR) values of nucleosides; however, when a mixture was used, significantly lower RWR values were observed for all nucleosides except thymidine when using 1000 molar excess of BSTFA greater than 1000 should be used for silylation and chromatography of nucleosides in an RNA hydrolysate. As reported earlier the best derivatization of nucleosides was achieved using closed tube silylation at 150 degrees for 15 min with 225 molar excess BSTFA and chromatography on 4% OV-11 on Supelcoport. In general, the presence of salts and other substances can be significant in quantitative work, thus it is suggested that they be removed using chromatographic cleanup methods. The stability of nucleosides as a function of concentration of HCl, at room temperature was studied and very low RWR values for nucleosides were obtained when stored for 48 h in greater than 0.001 N HCl. Trimethylsilylation of various nucleotides and dinucleotides were made at 15 min as a function of temperature, and at 150 degrees at different times...
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PMID:Derivatization and chromatography of nucleosides and nucleotides. 1 23

The binding of Ca2+ to a salivary phosphoprotein, protein C, was studied by equilibrium dialysis. In 5mM-Tris/HCl buffer, pH 7.5, protein C bound 190 nmol of Ca2+/mg of protein. The apparent dissociation constant, K, was determined to be 1.9 x 10(-4)M and the binding of Ca2+ to the protein was non-co-operative. The binding of Ca2+ to protein C apparently depends on groups which ionize above pH 5.0. Ca2+ binding decreased with increased concentration of the dialysis buffer and on addition of SrCL2, MgCl2 and MnCl2 to the dialysis buffer. Digestion of protein C with trypsin or collagenase or heating of the protein to 60 degrees or 100 degrees C had little or no effect on the Ca2+ binding. Digestion of protein C with alkaline phosphatase caused a decrease in the amount of protein-bound Ca2+. This was also found for another salivary phosphoprotein, protein A. In the absence of Ca2+ the S020,w for protein C was 1.29 S and in the presence of Ca2+ it was 1.46S. Ca2+ may cause a conformational change in the protein or an aggregation of the protein molecules. No conformational changes of protein C in the presence of Ca2+ could be detected by circular dichroism or nuclear magnetic resonance.
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PMID:The binding of calcium to a salivary phosphoprotein, protein C, and comparison with calcium binding to protein A, a related salivary phosphoprotein. 1 96

A plasma membrane-enriched fraction was prepared from homogenized rat pancreatic islets by a one-step sucrose gradient centrifugation. Using 125I-wheat germ agglutinin as a plasma membrane probe, a fraction was obtained at a sucrose density of about 1.10 that was enriched in 5'-nucleotidase, Mg2+-ATPase and alkaline phosphatase. The fraction contained little, if any, monoamino oxidase activity, insulin or DNA. Hydrolysis of 3-0-methyl-fluoresceinphosphate was stimulated by K+ (10mM) at a pH optimum of pH 8.2. Hydrolysis of ATP-gamma-32P in the presence of MgCl2 was of high specific activity and was optimum at pH 7.0 and 8.2. K+ did not affect ATP-hydrolysis. At pH 8.2, a small fraction of the total Mg2+-ATPase activity was inhibited by ouabain in the presence of Na+ and K+. Since K+-stimulated phosphatase activity does not correlate with Mg2+-ATPase, the two assay systems define separate enzymatic processes.
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PMID:Cation-dependent phosphatase activites in a rat pancreatic islet plasma membrane fraction prepared by one-step gradient centrifugation. 3 53

The conditions necessary for the secretion of phospholipase C (phosphatidylcholine cholinephosphohydrolase) by Pseudomonas aeruginosa were studied. Enzyme secretion by washed cell suspensions required a carbon source and ammonium, potassium, and calcium ions. The calcium requirement could be substituted by magnesium and strontium but not by copper, manganese, cobalt, or zinc. During growth in liquid medium, cells secreted phospholipase C during late logarithmic and early stationary phases. Secretion was repressed by the addition of inorganic phosphate but not by organic phosphates, glucose, or sodium succinate. Studies with tetracycline indicated that de novo protein synthesis was necessary for the secretion of phospholipase C and that the exoenzyme was not released from a preformed periplasmic pool. Similarly, extraction of actively secreting cells with 0.2 M MgCl2 at pH 8.4 solubilized large quantities of the periplasmic enzyme alkaline phosphatase but insignificant amounts of phospholipase C. Bacteria continued to secrete enzyme for nearly 45 min after the addition of inorganic phosphate or rifampin.
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PMID:Secretion of phospholipase C by Pseudomonas aeruginosa. 11 87

A new species of orthophosphate repressible extracellular 5'-nucleotidase (5'-ribonucleotide phosphohydrolase, EC 3.1.3.5) was found to be released into mycelial culture media when a wild type strain of Neurospora crassa was grown on limiting amounts of phosphate. The production of 5'-nucleotidase and extracellular acid and alkaline phosphatase was inhibited by the addition of rifampicin when it was added at the later stage of mycelial growth, but not when it was added at a very early stage. The 5'-nucleotidase and extracellular alkaline phosphatase were partially purified and characterized. pH optimum of the former was 6.8 and that of the latter was higher than 10.0. The 5'-nucleotidase activity was inhibited by ethylenediaminetetraacetate (EDTA) and ZnCl2 at pH 6.8 and stimulated by MnCl2 and CoCl2 at pH 4.0. Alkaline phosphatase activity was stimulated by EDTA, MgCl2, CoCl2 and MnCl2. 5'-nucleotidase activity was stimulated by EDTA, MgCl2, CoCl2 and MnCl2. 5'-nucleotidase hydrolyzed various 5'-nucletides but not 3'-nucleotides or other various phosphomono- and diester compounds. Alkaline phosphatase hydrolyzed all the phosphomonoester compounds tested. Mutants, nuc-1 and nuc-2, which were originally isolated by the inability to utilize RNA or DNA as a sole source of phosphate, were unable to produce 5'-nucleotidase or six other repressible enzymes reported previously. These mutants showed no or significantly reduced growth on orthophosphate-free nucleotide media depending on the number of conidia inoculated, mainly because of loss of ability to produce these repressible extracellular phosphatases.
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PMID:Control of the production and partial characterization of repressible extracellular 5'-nucleotidase and alkaline phosphatase in Neurospora crass. 13 48

In the course of serial passages for several years a line of uncloned HeLa cells (A line) showed a gradual decrease in plaquing efficiency by coxsackievirus A9 (CA9 virus), while subcultures prepared from the same line kept frozen at an early passage level (A original line) did not show any change. However, it was observed later that the plaque-forming ability of the A original line (A orig. line) also decreased after serial passages as was observed with the A line. Comparing the characteristics of the same cell line at two different passage levels, it was found that the efficiency of adsorption of virus to cells was nearly the same, while virus yield at 8 hours after infection was different. The activity of alkaline phosphatase of cells was also different between these two passage levels, suggesting that a high enzymatic activity is associated with the susceptibility of cell cultures to CA9 virus. Magnesium chloride at 25 mM enhanced plaque formation by CA9 virus in highly passaged less susceptible cell cultures, and a possible role of the chemical as a stabilizer of alkaline phosphatase was discussed.
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PMID:Variation in susceptibility of HeLa cell lines to coxsackievirus A9. 22 35

The existence of a specific inorganic pyrophosphatase (PPi ase), distinct from alkaline phosphatase in bone has been suggested but is often regarded as questionable. In the present investigation, several features of PPi ase activity have been demonstrated, which suggest that it represents an enzyme protein different from those splitting phosphomonoesters and ATP. PPi ase was largely destroyed during extraction with n-butanol, which facilitated the solubilization of alkaline phosphatase and ATP splitting enzymes and only partially destroyed acid phosphatase. Two major groups of phosphate esters and pyrophosphates splitting enzymes were separated by gel filtration from homogenates of rat bones. The first pool contained high ATP-ase and phosphomonoesterase activities, but only low activity against inorganic pyrophosphate (PPi) in the presence of MgCl2. The second pool was most active against PPi at pH 7.5 in the presence of excess MgCl2 and only slightly hydrolyzed phosphomonoesters or ATP. Immunodiffusion showed that these 2 pools contained 2 distinct proteins. It was concluded that there exists a specific inorganic pyrophosphatase distinct from phosphomonoesterases and ATP-ases in bone tissue.
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PMID:Inorganic pyrophosphatase activity distinct from alkaline phosphatase in rat bone. 59 70

A study was conducted to determine the feasibility of using the qualitative screening method specifying phenolphthalein monophosphate for differentiating reactivated and residual alkaline phosphatase activity. The relative increase in activity of the enzyme in the presence of MgCl2 serves to distinguish reactivated and residual alkaline phosphatase activity. Ten samples each of pasteurized (172 degrees F for 24 sec), sterilized (about 300 degrees F for a minimum of 2 sec) half-and-half and heavy cream were analyzed. Most samples yielded negative results initially but demonstrated activity after incubation 1 hr at 34 degrees C. The average values in terms of + marks, for the half and half and heavy cream in samples without MgCl2 were less than 1 and 2.4, respectively; for samples treated with MgCl2, the values were 2.18 and 4.6, respectively, indicating reactivated phosphatase activity. In samples containing various levels of raw milk, the activity observed in the diluted, Mg2+-containing samples was less than in the undiluted samples containing no Mg, indicating residual phosphatase activity.
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PMID:Rapid determination of alkaline phosphatase reactivation. 92 45

The examination of alkaline phosphatase isoenzymes by means of isoelectric focusing in polyacrylamide gel rods using the apparatus and focusing method of Righetti and Drysdale is discussed. A simultaneous coupling procedure using alpha-naphthyl phosphate and fast blue salt R in 2-amino-2-methyl-1,3-propanediol buffer, pH 9.68, containing MgCl2 and ZnSO4 proved sensitive for developing the enzyme bands. Also discussed are the effects seen with the incorporation of Triton X100 into the gel and sample mixtures. Enzyme, which remained at the top of the gel without using this detergent, entered the gel easily with the addition of Triton X-100 into the application solution. Incorporation of Triton into the gel matrix resulted in some enzyme band patterns that showed distinct differences from gels containing no Triton.
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PMID:Isoelectric focusing of alkaline phosphatase isoenzymes in polyacrylamide gels. Use of Triton X-100 and improved staining technic. 99 65


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