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 work described in this report suggests the existence of two biochemically distinguishable forms of the interferon-inducible, double-stranded RNA-dependent protein kinase. Kinase isolated from the cytosolic fraction (S-100) and the ribosome salt wash fraction of interferon-treated cells differed in their chromatographic properties. S-100 kinase eluted from a gel filtration column with M(r) = 140,000-160,000 and was predominantly anionic in nature, whereas ribosomal kinase eluted with M(r) = 66,000 and was predominantly cationic in nature. Purified preparations of S-100 kinase contained the M(r) = 66,000 subunit, P1, as the only polypeptide present in stoichiometric amounts, and thus the S-100 kinase appears to be a dimer of P1 subunits. Dimerization of the S-100 kinase was dependent on the phosphorylation state of the enzyme. Kinase isolated from S-100 was partially phosphorylated. Dephosphorylation of the S-100 kinase by treatment with alkaline phosphatase resulted in a monomeric form of the enzyme with biochemical characteristics similar to that of the ribosome salt wash kinase.
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PMID:Cytosolic double-stranded RNA-dependent protein kinase is likely a dimer of partially phosphorylated Mr = 66,000 subunits. 137 30

The localization of Ca(++)-ATPase activity was investigated in the rat cerebral cortex using an ultracytochemical method. Our cytochemical procedure for the detection of enzyme activity is based on an incubation medium consisting of tricine buffer, ATP-disodium salt as substrate, cerium chloride as capturing agent, CaCl2, and levamisole as a nonspecific alkaline phosphatase inhibitor. Final pH was 7.4. Reaction products showing Ca(++)-ATPase activity were localised on the pre- and postsynaptic plasma membrane in association with synaptic vesicles, postsynaptic dendrites and on the axolemma in myelinated nerve fibres. The verification of the main enzymatic properties of Ca(++)-ATPase localization activity is the subject of the following report.
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PMID:Enzyme ultracytochemical demonstration of Ca(++)-ATPase in the rat cerebral cortex. 144 75

Certain phosphonocarboxylate analogues of phosphate are known to inhibit Na(+)-phosphate (Pi) cotransport in renal brush border membrane (BBM), but previously tested potential inhibitors incorporating structurally versatile aryl functionality were inactive. In this work, a series of novel alpha-halogenated [(phenylphosphinyl)methyl]phosphonates [PhpXYMP: X, Y = H, F (2); F, F (3); H, Cl (6); Cl, Cl (4); H, Br (7); Br, Br (5); and Cl, Br (8)] were prepared via synthesis of the corresponding triethyl esters, acid hydrolysis, and isolation as pyridine salts. The compounds were evaluated as inhibitors of Na(+)-gradient-dependent 32Pi uptake by rat renal cortex BBM vesicles (BBMV) in vitro. The PhpFMP racemate 2 had higher activity (-49% delta inhibition) than other members of the series (-22 to -39% delta inhibition). pKa values of 1.5-2.0, 2.7, and 7.1 were estimated for 2 using a 31P delta vs pH plot, indicating that in the activity assays it exists as both dianion and trianion, with the latter form predominant. PhpFMP had no significant inhibitory effect on Na(+)-gradient-dependent uptake of D-glucose or L-proline in the same BBMV, and did not inhibit BBM alkaline phosphatase. Kinetic analysis showed that PhpFMP acts as a strictly competitive inhibitor of Na(+)-Pi cotransport with Ki = 0.358 +/- 0.021 mM (n = 3). The racemate 2 was resolved as its (-)-quinine salt into enantiopure (+)-2 [Na+ salt, [alpha]25D = +6 degrees (aqueous MeOH)] and a Na+ salt of 2 enriched in (-)-2. The two compounds did not differ significantly as inhibitors of Na(+)-gradient dependent 32Pi uptake by rat renal cortex BBM vesicles (BBMV) in vitro. The results are discussed in terms of structural requirements for inhibition.
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PMID:Alpha-halo [(phenylphosphinyl)methyl]phosphonates as specific inhibitors of Na(+)-gradient-dependent Na(+)-phosphate cotransport across renal brush border membrane. 147 88

A method which allows discrete nucleic acid sequences to be detected with differently colored hybridization signals on the same blot involving only a single hybridization step is described. Nucleic acid probes labeled with digoxigenin, fluorescein, or biotin are hybridized simultaneously to immobilized target nucleic acids. Differential colorimetric detection is carried out in consecutive alkaline phosphatase-based immunoassays with one of three 3-hydroxy-2-naphthoic acid anilide phosphate/diazonium salt combinations as substrate. Each label is visualized by a different color precipitate (green, red, and blue) directly on the membrane. We demonstrate the use of this method in multicolor plasmid mapping, detection of different genomic sequences on a single Southern blot, discrimination of transcription levels in a Northern blot, and colony screening. Advantages and limitations of the method, as well as further applications, are discussed.
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PMID:Multiple nucleic acid labeling and rainbow detection. 148 97

Two commonly used diagnostic tests for Staphylococcus aureus (MRSA) are cultivation of bacteria from clinical samples on mannitol-salt agar plate or on MRSA-screen agar plate with oxacillin. However, the use of PCR and DNA probes is considered more rapid and sensitive, as staphylococcal cells have high resistance to beta-lactam due to the novel penicillin-binding protein, PBP2'. Therefore, three different pairs of DNA primers (P1F-P1R, P2F-P2R and P3F-P3R) complementary with unique regions of the MRSA PBP2' gene were synthesized for use in polymerase chain reactions with DNA of MRSA. Amplified target DNA of 466, 480 and 630 bp could be resolved on ethidium bromide-stained gels, and hybridized with DNA probes conjugated to alkaline phosphatase. When applied to pure cultures on the MRSA screen agar, the DNA probes detected MRSA in 180 of 196 (P1F-P1R), 72 of 83 (P2F-P2R) and 66 of 71 (P3F-P3R) culture-positive specimens (accuracy range, 86.7-93.0%), while 61.6-89.3% of MRSA were detectable in culture-positive specimens streaked on the mannitol-salt agar. When applied directly to clinical samples, this DNA probe assay detected MRSA in culture-positive specimens within an accuracy range of 50.0-79.3%. It was thus clear that comparison of cultivation and DNA hybridization results yields good correlation with respect to detection of MRSA. These results suggest that the DNA probe assay may be useful in complementing existing culture techniques.
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PMID:[Detection of methicillin-resistant Staphylococcus aureus using PCR and non-radioactive DNA probe]. 150 81

To study the phosphorylation of one of the G-box binding factors from Arabidopsis (GBF1), we have obtained large amounts of this protein by expression in Escherichia coli. Bacterial GBF1 was shown to be phosphorylated very efficiently by nuclear extracts from broccoli. The phosphorylation activity was partially purified by chromatography on heparin-Sepharose and DEAE-cellulose and was characterized. It showed the essential features of casein kinase II activity: utilization of GTP in addition to ATP as a phosphate donor, strong inhibition by heparin, preference for acidic protein substrates, salt-induced binding to phosphocellulose, and salt-dependent deaggregation. The very low Km value for GBF1 (220 nM compared to approximately 10 microM for casein) was in the range observed for identified physiological substrates of casein kinase II. Phosphorylation of GBF1 resulted in stimulation of the G-box binding activity and formation of a slower migrating protein-DNA complex. The conditions of this stimulatory reaction fully corresponded to the properties of casein kinase II, in particular its dependence on the known phosphate donors. The DNA binding activity of the endogenous plant GBF was shown to be reduced by treatment with calf alkaline phosphatase; this reduction was diminished by addition of fluoride and phosphate or incubation in the presence of casein kinase II and ATP.
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PMID:DNA binding activity of the Arabidopsis G-box binding factor GBF1 is stimulated by phosphorylation by casein kinase II from broccoli. 152 62

The effects of rifampicin treatment (10 mg.kg-1.day-1) on pruritus and cholestasis were evaluated in 16 patients with primary biliary cirrhosis and pruritus followed up for 2-24 months. Assessment of pruritus severity, liver tests, aminopyrine breath test, and bile acids was done at 2 weeks and every 3 months after the beginning of the study. Two patients (12.5%) were withdrawn after 2 months of treatment because they had hepatitis caused by rifampicin. Four patients were withdrawn after 4 months because of liver transplantation (3 cases) and the development of leg edema associated with administration of rifampicin. The remaining 10 patients received therapy for 14.4 +/- 0.7 months and did not experience side effects. Pruritus improved in all patients and disappeared in 11 patients (79%) after 3 months of treatment. Moreover, all patients followed up for more than 1 year were free of pruritus. The alkaline phosphatase level decreased significantly, and the aminopyrine breath test results increased significantly after 2 weeks of treatment (P less than 0.001) and did not change thereafter. In the 9 patients treated for 15 months, alkaline phosphatase levels decreased to 63% of the basal levels and aminopyrine breath test results increased to 153% of baseline values. Transaminases, gamma-glutamyltransferase, and total bile salt levels decreased significantly after 2 weeks of treatment but returned to baseline after 3 months. No changes in bilirubin and cholesterol levels were observed. It is concluded that long-term rifampicin treatment is effective for relieving pruritus in primary biliary cirrhosis, but liver enzymes should be monitored to detect drug-induced hepatitis.
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PMID:Effects of long-term rifampicin administration in primary biliary cirrhosis. 158 27

Cyanine dye fluorescence and alkaline phosphatase activities have been compared directly by confocal microscopy in a wide variety of cells present in the follicle-associated epithelium of the mouse Peyer's patch to test the hypothesis that antigen-transporting M cells have a low membrane potential. In order to make these comparisons it was first necessary to equilibrate living tissue with the membrane potential sensitive dye DIOC5(3), fix with glutaraldehyde and then incubate the fixed tissue with naphthol AS-BI phosphate, a substrate which is hydrolysed by alkaline phosphatase present in the luminal membrane of these epithelial cells. Naphthol AS-BI produced by this reaction is then coupled to Fast Red TR diazonium salt at the site of hydrolysis. Selecting the 488 nm wavelength of the argon laser source then allows one to measure alkaline phosphatase activities as Fast Red absorbance and membrane potentials by DIOC5(3) fluorescence. Results obtained show a linear correlation between membrane potential and alkaline phosphatase activity. Relative lack of alkaline phosphatase activity, determined in fixed tissue, has been used previously to identify antigen-transporting M cells (Smith et al., 1987). The present work shows that it is now possible to recognize these cells in living tissue by measurement of DIOC5(3) fluorescence. The possible importance of this finding in providing a way to study cell surface-antigen interactions taking place in living tissue is discussed.
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PMID:Confocal microscopical analysis of epithelial cell heterogeneity in mouse Peyer's patches. 160 96

To investigate the effect of lipopolysaccharide (LPS) on phagocytic activity of collagen fibrils by periodontal fibroblasts, we studied rat molar gingival connective tissue and periodontal ligament under light and electron microscopy after topical application of LPS (5 mg/ml in physiological salt solution (PS)) on the gingival sulcus. Phagocytic activity of collagen fibrils by fibroblasts was evaluated by counting the number of collagen-containing vacuoles inside fibroblasts that were present within a defined area (1200 microns2). Values obtained from fibroblasts in the subepithelial connective tissue, the region near the alveolar crest, and the middle region of periodontal tissue were compared. Periodontal ligament fibroblasts showed increased phagocytosis of the collagen fibrils from 3 hours to 1 day after topical LPS application, but no differences were observed in the gingival tissue. The intracytoplasmic vacuoles containing collagen fibrils were of various sizes and shapes, showing positive for acid phosphatase and/or alkaline phosphatase reaction. Collagen phagocytic activity of the fibroblasts in the middle region of the periodontal ligament also increased after PS treatment. However, this was significantly less than that observed in LPS-treated animals (p less than 0.01). This study indicates that LPS may enhance the degradation of collagen by stimulating the phagocytic activity of the periodontal ligament fibroblasts.
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PMID:Enhanced collagen phagocytosis by rat molar periodontal fibroblasts after topical application of lipopolysaccharide--ultrastructural observations and morphometric analysis. 160 30

The absorption characteristics of ethiofos were studied using the rat in situ intestine circulating perfusion technique. Slow absorption kinetics were observed for ethiofos with varying rates of absorption and metabolism/degradation in situ as a function of buffer and absorption enhancers. In most cases less than 10 per cent of the radiolabeled compound is lost from the circulating perfusate in 90 min. In addition, over the same time period greater than 40 per cent of the intact parent compound was lost by degradation. Much of the difference can be accounted for in the formation of the free thiol metabolite. WR-1065, suggesting ester hydrolysis or metabolic activity. Good stability was observed in all perfusate systems ex vivo indicating that the degradation occurs in situ. The disodium salt of ethylenediaminetetraacetic acid (EDTA) was shown to be an effective absorption enhancer of ethiofos. The enhancement of intestinal absorption by EDTA was dose-dependent resulting in a 20-fold increase in blood levels of ethiofos in the portal blood. Follow-up studies in the rhesus monkey confirm this observation. Salicylate and dimethylsulfoxide (DMSO) also resulted in absorption enhancement although to a lesser degree than that seen after EDTA treatment. Addition of several alkaline phosphatase inhibitors did not significantly improve absorption of ethiofos in the rat small intestine. Proposed mechanism(s) for intestinal absorption and absorption enhancement of ethiofos are discussed.
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PMID:Characterization of ethiofos absorption in the rat small intestine. 165 90


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