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

Three nucleosides catalyzing the oxidoreduction of NADH and K3Fe(CN)6 were isolated from Torula yeast RNA and also obtained by a series of steps: SDS-phenol extraction, nuclease P1 digestion, alkaline phosphatase digestion, anion exchange chromatography, and HPLC on an ODS column. Their chemical structures were clearly determined as 5-hydroxyuridine, 8-hydroxyguanosine, and 8-hydroxyadenosine from the results of FAB-MS, 1H and 13C-NMR spectroscopies.
 ...
PMID:Novel minimum ribozymes with oxidoreduction activity: 5-hydroxyuridine, 8-hydroxyguanosine, and 8-hydroxyadenosine isolated from Torula yeast RNA. 184 45

Cultured cardiomyocytes were used to study the turnover and post-translational modification of connexin43 (Cx43), a major gap junction protein in neonatal cardiac myocytes. Immunoprecipitation of [35S]Met-labelled lysates with anti-Cx43 antibodies followed by analysis using SDS/PAGE and fluorography revealed two bands, one at 40 kDa and the other at 42 kDa. Alkaline phosphatase treatment of [35S]Met-labelled Cx43 eliminated the band at 42 kDa, suggesting that it represented a phosphorylated form of the protein. This was confirmed by [32P]P1 incorporation into the 42 kDa band, but not into the band at 40 kDa. In addition, another alkaline phosphatase-sensitive phosphorylated form of Cx43 was identified at 44 kDa. In pulse-chase experiments, the half-life of Cx43 in cardiomyocytes was determined to be 1-2 h. Furthermore, the turnover rate of phosphate groups on Cx43 was found to be experimentally defined by the half-life of the protein. The observation that phosphate groups can remain with the protein throughout its life is consistent with the finding that in isolated adult rat heart gap junction plaques, Cx43 is primarily phosphorylated. We postulate that the rapid turnover of Cx43 and its multiple sites of phosphorylation play important roles in the regulation of cell-cell communication via gap junctions.
 ...
PMID:Turnover and phosphorylation dynamics of connexin43 gap junction protein in cultured cardiac myocytes. 184 32

To investigate the extent to which whole tau proteins, structurally abnormal tau and fragments of tau are incorporated into neurofibrillary tangles in Alzheimer's disease, an immunocytochemical mapping study using a panel of antibodies to several synthetic human tau peptides has been performed. Neurofibrillary tangles were immunolabelled in situ, and paired helical filaments (PHF), the principal structural component of tangles, were immunolabelled after isolation and Pronase treatment. N-Terminal and C-terminal domains of tau were found to be present in tangles in situ. SDS-treated PHF were found to contain most of the C-terminal half of tau and were also labelled by antibodies to ubiquitin. Only some of these PHF were labelled by antisera to tau sequences towards the N-terminus, and this enabled the identification of a region of tau in which proteolytic cleavage may occur. The ultrastructural appearance of the immunolabelling suggested that both the N- and C-terminal domains of tau extend outwards from the axis of PHF. After Pronase treatment. PHF were strongly labelled only by an antiserum to PHF and by the antiserum to the most C-terminal tau synthetic peptide. The latter antiserum also strongly labelled extracellular tangles in situ, whereas these extracellular tangles were poorly labelled by the antisera to the other synthetic peptides. One anti-(tau peptide) serum labelled a population of neurofibrillary tangles in situ only after alkaline phosphatase pretreatment of tissue sections. Our results show that, although peptides along the length of the tau molecule are associated with neurofibrillary tangles in situ, only the C-terminal one-third of the molecule is tightly associated with PHF, since this region of tau is resistant to SDS treatment of PHF. We also report the existence in PHF in situ of a masked tau epitope which is partially unmasked by dephosphorylation. These results are indicative of post-translational changes in tangle-associated tau in degenerating neurons in Alzheimer's disease.
 ...
PMID:Tau in Alzheimer neurofibrillary tangles. N- and C-terminal regions are differentially associated with paired helical filaments and the location of a putative abnormal phosphorylation site. 189 84

The tau-immunoreactive A68 polypeptides found in brains from patients with Alzheimer's disease have been studied by Western blotting using (1) antibodies to synthetic peptides corresponding to sequences that span the complete human tau molecule, and (2) antibodies specific for inserts 1 and 2 found towards the N-terminus of some tau isoforms. The three major A68 polypeptides were labelled by all of the antibodies to sequences common to all tau isoforms, but the faster-migrating A68 polypeptides was not labelled by either of the two antibodies specific for inserts 1 and 2. Treatment with alkaline phosphatase of non-solubilized A68 did not change its electrophoretic mobility on SDS/PAGE under the conditions described here. However, A68 that was solubilized before treating it with alkaline phosphatase was found to move faster on SDS/PAGE than untreated A68, to a position similar to that of normal tau. We also confirmed that A68 preparations contain numerous paired helical filaments (PHF). These PHF were labelled by all anti-tau antibodies, including insert-specific antibodies. Our results further support the notion that PHF contain abnormally phosphorylated tau in an aggregated state, and indicate that these abnormally phosphorylated tau forms are composed of several tau isoforms and that the full length of the tau molecule is present in these polypeptides.
 ...
PMID:A68 proteins in Alzheimer's disease are composed of several tau isoforms in a phosphorylated state which affects their electrophoretic mobilities. 195 78

We have detected the in situ activities of DNA glycosylase, endonuclease, exonuclease, DNA polymerase, and DNA ligase using a novel polyacrylamide activity gel electrophoresis procedure. DNA metabolizing enzymes were resolved through either native or SDS-polyacrylamide gels containing defined 32P-labeled oligonucleotides annealed to M13 DNA. After electrophoresis, these enzymes catalyzed in situ reactions and their [32P]DNA products were resolved from the gel by a second dimension of electrophoresis through a denaturing DNA sequencing gel. Detection of modified (degraded or elongated) oligonucleotide chains was used to locate various enzyme activities. The catalytic and physical properties of Novikoff hepatoma DNA polymerase beta were found to be similar under both in vitro and in situ conditions. With 3'-terminally matched and mismatched [32P]DNA substrates in the same activity gel, DNA polymerase and/or 3' to 5' exonuclease activities of Escherichia coli DNA polymerase I (large fragment), DNA polymerase III (holoenzyme), and exonuclease III were detected and characterized. In addition, use of matched and mismatched DNA primers permitted the uncoupling of mismatch excision and chain extension steps. Activities first detected in nondenaturing activity gels as either multifunctional or multimeric enzymes were also identified in denaturing activity gels, and assignment of activities to specific polypeptides suggested subunit composition. Furthermore, DNA substrates cast within polyacrylamide gels were successfully modified by the exogenous enzymes polynucleotide kinase and alkaline phosphatase before and after in situ detection of E. coli DNA ligase activity, respectively. Several restriction endonucleases and the tripeptide (Lys-Trp-Lys), which acts as an apurinic/apyrimidinic endonuclease, were able to diffuse into gels and modify DNA. This ability to create intermediate substrates within activity gels could prove extremely useful in delineating the steps of DNA replication and repair pathways.
 ...
PMID:Characterization of DNA metabolizing enzymes in situ following polyacrylamide gel electrophoresis. 200 53

The chalcone synthase (EC 2.3.1.74) gene promoter from the bean Phaseolus vulgaris L. contains a silencer element between positions -140 and -326 fro the transcription start site that is functional in electroporated soybean protoplasts. This element contains three binding sites for a bean nuclear factor (SBF-1) with DNA sequence recognition properties that are very similar to those of nuclear factor GT-1. By using a synthetic tetramer of one of the binding sites as probe, we have purified sequence-specific SBF-1 activity approximately 1750-fold from suspension-cell nuclei, by using a combination of ammonium sulfate precipitation, gel filtration, heparin-agarose chromatography, and sequence-specific DNA affinity chromatography. The factor exhibited an apparent molecular weight of 160,000-200,000 on the basis of gel filtration. A subunit molecular weight of approximately 95,000 was determined from SDS/polyacrylamide gel electrophoretic analysis of purified fractions, followed by Southwestern blot analysis (a protein blot probed with oligonucleotide probes), and from UV-cross-linking experiments. The factor lost DNA-binding activity on treatment with alkaline phosphatase. We discuss the properties of SBF-1 in relation to the functionality of GT-1 binding sequences in plant genes.
 ...
PMID:Characterization of a nuclear protein that binds to three elements within the silencer region of a bean chalcone synthase gene promoter. 200 88

tolA mutants of Escherichia coli K-12 release periplasmic proteins into the extracellular medium; they are sensitive to growth inhibitors such as cholic acid and tolerant to group A colicins and filamentous bacteriophage. Suppressor mutants of the tolA-876 allele were isolated by selecting for cholic acid resistant clones that did not release periplasmic ribonuclease I. One class of tolA suppressor strains carried mutations in the staA gene (for suppressor of tolA) located a 41 min. tolA-876 staA strains partially recovered a wild-type phenotype: they exported alkaline phosphatase and beta-lactamase into the periplasm and only released very low amounts of periplasmic proteins; moreover, they were sensitive to E1 and A colicins and more resistant than tolA-876 staA+ strains to various growth inhibitors. Furthermore, tolA-876 staA-2 and tolA+staA-2 mutants were 10- to 2700-times more resistant than staA+ strains to bacteriophages TuIa, TuIb and T4, and TuII whose receptors are major outer membrane proteins OmpF, OmpC and OmpA, respectively. SDS-PAGE analysis suggested that cell envelopes of staA or staA+ strains contained similar amounts of these proteins but characterization of strains carrying ompF (or C or A)-phoA gene fusions showed that mutation stA-2 reduced ompF gene expression by a factor of two. Analysis of double mutants strains carrying mutation staA-2 and a tolA, tolB, excC or excD periplasmic-leaky mutation showed that staA suppression was allele specific which suggested that proteins TolA and StaA might directly interact.
 ...
PMID:Isolation and characterization of extragenic suppressor mutants of the tolA-876 periplasmic-leaky allele in Escherichia coli K-12. 204 Apr 38

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.
 ...
PMID:Alkaline phosphatase from rat osseous plates: purification and biochemical characterization of a soluble form. 206 78

The supra- and suboesophageal ganglia of the American cockroach contain material which catalyses the alkaline hydrolysis (pH 9.5) of 5-bromo-4-chloro-3-indolyl phosphate in the presence of Nitro blue tetrazolium. Histochemical studies on unfixed cryostat sections indicate that this type of alkaline phosphatase is restricted to discrete regions in the cockroach brain. Highest enzyme activity is encountered in the mushroom bodies, central body, antennal glomeruli and specific parts of some distinct neural connections including the optic nerve, antennal nerve, circumoesophageal connectives and nerves leaving the suboesophageal ganglion. Tissue fixation by use of formaldehyde-type fixatives, as well as routine paraffin-embedding, completely destroy all histochemically detectable enzyme activity. Native polyacrylamide gradient electrophoresis suggests that the alkaline phosphatase activity is present as multiple isozymic forms, which show up in the 120-130 kD range of standard proteins. Enzyme activity becomes undetectable after fixation (trichloroacetic acid, formaldehyde containing fixatives) of electrophoretically separated native proteins, as well as after electrophoresis in denaturing conditions (SDS and beta-mercapto-ethanol, boiling). However, the enzyme activity remains virtually unaffected after storage of the sample for prolonged periods at -20 to -80 degrees C.
 ...
PMID:Alkaline phosphatase activity in the brain of the American cockroach Periplaneta americana L. 207 11

Sequences of hepatitis B virus DNA were detected specifically in the sera of infected individuals by a non-radioactive riboprobe nucleic acid hybridization assay. The nucleic acids contained in serum specimens were prepared by an overnight proteinase K-/SDS-digest, phenol/chloroform-extraction and ethanol-precipitation, and immobilized on nylon membranes by the dot-blot technique. Digoxigenin-labelled 1.4 kb RNA-probes were generated by the phage sp 6 RNA-polymerase from a linearized HBV DNA transcription template containing the appropriate promoter. Hybridized probes were detected by alkaline phosphatase-conjugated sheep anti-digoxigenin Fab-fragments, and 5-bromo-4-chloro-3-indolylphosphate (BCIP) and nitroblue tetrazolium salt (NBT) as chromogenic substrates for the subsequent ELISA procedure. With a detection limit of 0.5-1.0 pg HBV DNA and a high specificity, the results were comparable to those obtained by the standard assay employing radioactively labeled DNA-probes.
 ...
PMID:A nonradioactive riboprobe assay for the detection of hepatitis B virus DNA in human sera. 208


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>