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:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The 5'-phosphomonoesterase activity of 5'-nucleotidase (EC 3.1.3.5) and alkaline phosphatase (EC 3.1.3.5) participates in the catabolism of purine ribonucleotides to uric acid in humans. Initial velocity studies of 5'-nucleotidase suggest a sequential mechanism of interaction between AMP nad MgCl2, with a Km of 14 and 3 muM, respectively. With product inhibition studies the apparent Ki's for adenosine, inosine, cytidine, and inorganic phosphate were 0.4, 3.0, 5.0, and 42 mM, respectively. A large number of nucleoside mono-, di-, and tri-phosphate compounds were inhibitors of the enzyme. Allopurinol ribonucleotide, ADP, or ATP were competitive inhititors when AMP was the substrate, with a Ki slope of 120 muM. The phosphomonoesterase activity of human placental microsomal alkaline phosphatase had a pH optimum of 10.0 and had only 18% of maximum activity at pH 7.4. Substrates and inhibitors included almost any phosphorylated compound. The Km for AMP was 0.4 mM and the apparent Ki for Pi was 0.6 mM. Activity was increased only 19% by 5 mM MgCl2. These observations suggest that 5'-nucleotidase and alkaline phosphatase may be inhibited by ATP and Pi, respectively, under normal intracellular conditions, and that AMP may be preferentially hydrolyzed by 5'-nucleotidase.
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PMID:Purine catabolism in man: inhibition of 5'-phosphomonesterase activities from placental microsomes. 101 16

A procedure for determination of serum alkaline phosphatase activity (EC 3.1.3.1) in diethanolamine (DEA) buffer with an AutoAnalyzer II apparatus was designed. The buffer used was 1.0 mol/l DEA-HC buffer, pH 9.8 at 37 degree C, containing 0.5 mmol/l of MgCl2 and 10 mmol/l of substrate 4-nitrophenyl-phosphate. The reaction time was about 3 min at 37 degree C. The enzyme activity (U/l) was calculated by determining the amount of 4-nitrophenol formed in reaction. A sampling rate of 70 samples per hour can be used with good linearity up to 1000 U/l. The results obtained by the new continuous-flow system were compared with those measured by the kinetic method according to the Scandinavian recommendation (10). A close correlation between the two methods was observed.
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PMID:A continuous-flow method for the determination of the activity of serum alkaline phosphatase in diethanolamine buffer. 115 23

The rats were given potassium bichromate (K2Cr2O7) in dose of 2 and 5 mg/kg of body weight and magnesium chloride (MgCl2) in dose of 500 mg/kg for a period of 30 days. The two compounds were also given conjointly (K2Cr2O7-5 mg+MgCl2-500 mg). There were carried out histopathological as well as histochemical examinations of acid phosphatase activity, alkaline phosphatase activity and adenosine triphosphatase activity in the liver. With small doses (2 mg) of potassium bichromate no changes have been stated. With larger doses (5 mg) of potassium bichromate an increase of histochemical reaction to acid phosphatase as well as forming of histopathological changes such as parenchymatous degeneration, steatosis of hepatocytes and their necrobiosis have been observed. There has not been found any protective action of magnesium chloride on the cytotoxic activity of potassium bichromate on the liver cell.
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PMID:[Histopathologic and histochemical examination of rat liver after prolonged experimental application of potassium bichromate]. 136 4

Brush-border membranes from rat duodenal and jejunal mucosa were prepared by differential Ca(2+)-precipitation. Kinetical properties and Mg(2+)-stimulation of alkaline phosphatase were studied for the enzyme either bound to these membranes or purified from these membranes by liquid chromatography. With p-nitrophenylphosphate as substrate, the alkaline phosphatase apparent Km was lower in jejunum (90 microM) as compared with duodenum (160 microM), and lower for the purified enzyme (jejunum: 55 microM; duodenum: 97 microM) as compared to the bound one. In the presence of 5 mM MgCl2, the substrate affinity was in all cases decreased. For the bound enzyme Vmax was 10 times greater in duodenum compared to jejunum. 5 mM MgCl2 tripled the Vmax of the duodenal bound enzyme and increased it by 50% for the jejunal one, but a seven-fold increase was recorded for the purified enzyme at both levels of intestine. The apparent affinity for Mg2+ was similar for the bound and the free enzyme, for duodenum and for jejunum (Mg0.5: +/- 40 microM).
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PMID:Bound and purified alkaline phosphatase from rat duodenal and jejunal brush-border membranes: kinetics and magnesium stimulation. 138 Mar 28

The effects of divalent cations and of some inhibitors on the activities of alkaline phosphatase and ATPase were examined in rat jejunal brush-border membranes (BBM) isolated by tha Ca(2+)-(BBMCa) or the Mg(2+)-precipitation method (BBMMg). Similar results were found in BBMCa and BBMMg though generally higher in BBMCa. Alkaline phosphatase activity was stimulated by 5 mM MgCl2 (30% to 44%), but not by 5 mM CaCl2 or 0.1 mM ZnCl2, at pH 9.5 or 7.4. ATPase activity was equally stimulated by 5 mM MgCl2 and by 5 mM CaCI2 (about 150%). Alkaline phosphatase activity was significantly inhibited by 1 mM vanadate, 5 mM diamox, 5.0 mM L-leucine and 1 mM theophylline. In contrast, Ca(2+)-ATPase and Mg(2+)-ATPase activities were not depressed by those alkaline phosphatase inhibitors, but were inhibited by 0.1 mM trifluoperazine (more than 70%). 0.1 mM ZnCl2 also appeared to be inhibitory to Ca(2+)-ATPase and Mg(2+)-ATPase, but not to alkaline phosphatase activity even in the presence of Ca2+ and Mg2+. These results suggest that Ca(2+)-ATPase and Mg(2+)-ATPase activities of the rat jejunal BBM are not merely manifestations of alkaline phosphatase, but rather belong to (a) distinct enzyme(s).
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PMID:Alkaline phosphatase and ATPases in brush-border membranes of rat jejunum: distinct effects of divalent cations and of some inhibitors. 138 82

A cerium-based incubation medium, developed for the light microscopical demonstration of alkaline phosphatase activity, was tried out for the electron microscopical demonstration of this enzyme in kidney and heart muscle of the rat. The medium is very stable and the pH is in the optimum range of the enzyme. The medium consists of 14 mM CeCl3, 11 mM Na-citrate, 4 mM MgCl2, 10 mM p-nitrophenyl phosphate, 0.18 M glycine/NaOH buffer, pH 9.3. Other concentrations of cerium and citrate were tried out as well but 14 mM CeCl3, and 11 mM Na-citrate gave the best results with a small amount of non-specific reaction product in the nucleus that can be largely avoided by postincubation rinsing in cerium-containing buffer. In the kidney reaction product was only present along the microvilli of the proximal tubular epithelial cells. In the glomerulus no reaction product could be found whereas light microscopical cryotome sections contained activity in the glomerulus. Replacement of glutaraldehyde by formaldehyde fixatives resulted in reaction product in glomerular and tubular basement membranes, on podocyte plasma membranes and in tubular basal infoldings. In glutaraldehyde-fixed heart muscle, reaction product was present in the basement membranes and on lateral plasma membranes of endothelial cells of blood capillaries.
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PMID:Electron microscopical demonstration of alkaline phosphatase activity with the cerium-based method in citrate-containing medium at pH 9.3 and the influence of glutaraldehyde fixation. 148 7

The ultrastructural localization of alkaline phosphatase (A1P) activity has been demonstrated in epiphyseal growth cartilage and metaphyseal bone of rats. Epiphyso-metaphyseal specimens were decalcified with EDTA and treated with MgCl2 to regenerate the enzymatic activity before incubation in a medium containing beta-glycerophosphate, MgCl2 and CeCl3. A1P activity was present on the outer surface of the plasmamembrane of maturing and hypertrophic chondrocytes and of osteoblasts. Moreover, the reaction product was present in chondrocyte lacunae, in matrix vesicles, and in cartilage matrix, as well as among uncalcified collagen fibrils of osteoid tissue in bone. The intensity of reaction was the lowest, or completely lacking, where the degree of matrix calcification was the highest. These results suggest that alkaline phosphatase is transported from the cells into the cartilage and bone matrix by its association with matrix vesicles and plasmamembrane components, and that its activity in cartilage and bone matrix is inhibited as it is incorporated in the mineral substance.
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PMID:Extracellular alkaline phosphatase activity in mineralizing matrices of cartilage and bone: ultrastructural localization using a cerium-based method. 161 46

Some investigators have described the presence in Alzheimer's disease brain extracts of several abnormal forms of the microtubule-associated protein tau, based on their unusual mobility in SDS/PAGE. It has been proposed that these abnormal forms of tau may be the result of aberrant tau phosphorylation. In this study we show that tau in extracts of Alzheimer's disease brain can be separated into two fractions based upon its solubility (100,000 g x 1 h supernatant) in non-denaturing conditions (100 mM-Mes, pH 6.5, 0.5 mM-MgCl2, 1 mM-EGTA and 1 M-NaCl). The tau isoforms with decreased mobility in SDS/PAGE are predominantly in an insoluble fraction, whereas the soluble tau is indistinguishable by its mobility in SDS/PAGE from tau in soluble extracts of control brain. Insoluble tau displaying abnormal mobility on SDS/PAGE was only found in Alzheimer and adult Down's syndrome brains and was absent from the brains of age-matched controls and from foetal and infant Down's syndrome brains. There was a good correlation between the presence of insoluble tau in brain extracts and the abundance of neurofibrillary tangles and senile neuritic plaques. The monoclonal antibody Tau. 1 stained insoluble tau on Western blots only after treatment of the nitrocellulose transfers with alkaline phosphatase, implying that this insoluble tau is in a particular state of phosphorylation. We conclude that, in Alzheimer's disease, a fraction of tau has a modified phosphorylation state and a decreased solubility; these modifications may precede formation of the neurofibrillary tangles characteristic of Alzheimer's disease and Down's syndrome in adults.
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PMID:Tau in Alzheimer's disease and Down's syndrome is insoluble and abnormally phosphorylated. 182 35

We have produced a batch of lyophilized alkaline phosphatase (AP) for use as an enzyme reference material. The enzyme was partly purified from pig kidney to a specific activity of 400 U/mg of protein and is essentially free from contaminating enzyme activities. The kinetic properties of the preparation are very close to those of the enzyme present in human serum. The partly purified AP was lyophilized in a matrix containing bovine serum albumin (40 g/L), MgCl2, ZnCl2 and NaCl. The vial-to-vial variability with respect to the catalytic concentration of the final product was 0.008. The predicted annual relative loss of activity was less than 0.01% at -20 degrees C and 0.04% at 4 degrees C. This material was certified using the IFCC proposed method. The certification procedure involved 19 laboratories throughout the world. The certified alkaline phosphatase catalytic concentration in the reconstituted material was 254 U/L with a 0.95 confidence interval of +/- 6 U/L.
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PMID:Certification of an enzyme reference material for alkaline phosphatase (CRM 371). 204 88

A soluble form of an alkaline phosphatase obtained from rat osseous plates was purified 204-fold with a yield of 24.3%. The purified enzyme showed a single protein band of Mr 80,000 on SDS-PAGE and an apparent molecular weight of 163,000 by gel filtration on Sephacryl S-300 suggesting a dimeric structure for the soluble enzyme. The specific activity of the enzyme at pH 9.4 in the presence of 2 mM MgCl2 was 19,027 U/mg and the hydrolysis of p-nitrophenyl phosphate (K0.5 = 92 microM) showed positive cooperativity (n = 1.5). The purified enzyme showed a broad substrate specificity, however, ATP, bis(p-nitrophenyl) phosphate and pyrophosphate were among the less hydrolyzed substrates assayed. Surprisingly the enzyme was not stimulated by cobalt and manganese ions, in contrast with a 20-25% stimulation observed for magnesium and calcium ions. Zinc ions exerted a strong inhibition on p-nitrophenylphosphatase activity of the enzyme. This paper provides a simple experimental procedure for the isolation of a soluble form of alkaline phosphatase which is induced by demineralized bone matrix during endochondral ossification.
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PMID:Alkaline phosphatase from rat osseous plates: purification and biochemical characterization of a soluble form. 206 78


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