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

---

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

Some factors influencing the detection of human cytomegalovirus (HCMV) in urine were investigated employing 2 enzyme-linked immunosorbent assays (ELISAs); one utilised anti-CMV DNA polymerase while the other anti-CMV mouse monoclonals as the detecting antibodies. The use of anti-CMV DNA polymerase was found to be superior in detecting HCMV in both urine and tissue culture fluids than anti-CMV monoclonals. Furthermore, alkaline phosphatase conjugates produced much lower background than did peroxidase conjugates. In reconstruction experiments, the extremes of pH in the urine clearly had an adverse effect on the detection rate of extracellular virus. pH correction of urines to neutrality improved the detection rate considerably. On the other hand, pH correction had little effect on the detection rate of intracellular HCMV in urine, although it was improved when specimens were subjected to repeated cycles of freeze-thawing, ultrasonication, and storage at 4 degrees C. It was concluded that, in addition to the factors investigated which all appear to affect virus detection rate, there may well be additional factors that interfere with CMV detection in the urine by ELISA particularly with intracellular virus.
...
PMID:Factors affecting the detection of cytomegalovirus in urine by sandwich enzyme immunoassays. 302 18

By statistical study on 135 patients with endometrial carcinoma, it is clarified that the most effective prognostic factor of the cancer is the histological grading. Well differentiated type is best prognostic and possesses hormone receptors. Application of cell culture is one of the most suitable choices in the study of hormone and human endometrial carcinoma. Present paper is to show usefulness of in vitro study by taking example of the above theme. 1) Binding ability of endometrial carcinoma cells to estrogen: Being explained by Gurpide et al. by using HEC-1 cells, the ability is under control of cGMP and cAMP ratio. 2) Responses to estrogen: DNA polymerase alfa of Ishikawa cells which possesses both estrogen and progesterone receptors (ER, PR) is stimulated first showing peak at 18 hours and alkaline phosphatase (ALP) is at 72 hours by E(2)10(-8)M, which is antagonized by OH-tamoxifen. PR level is also enhanced at its maximum after 3 day E2 treatment, and is analyzed by immunocytochemistry with PR mono-clonal antibody as well as biochemical assay. Gorski and Greene's theory that steroid receptor is localized in nuclei is confirmed in endometrial carcinoma. Growth of Ishikawa cells is apparently enhanced in the aspects of shortened cell cycle and unlimited saturation density. 3) Responses to progestogen: Nucleic acid syntheses of HEC-1 are immediately suppressed by progesterone (P) 2.5 microg or more. Electron microscopic findings show appearances of Golgi apparatus and lysosomal granules. Growth suppression is observed in the cell lines regardless of PR positivity. ALP activity of PR-negative HEC-50 cells
...
PMID:[Cell culture--its application in the study of hormone and endometrial carcinoma and feed-back to clinical medicine]. 315 Aug 47

The distribution of termination and initiation sites in a 5081-nucleotide minute virus of mice DNA template being copied by a highly purified mouse DNA polymerase alpha-DNA primase complex in the presence of GTP has been examined. The 3'-hydroxyl termini (17 in all) were clustered at six sites that were located 2-14 nucleotides upstream of C2A2C2, C2AC3, or C2A2T2 sequences. When either [alpha-32P]- or [gamma-32P]GTP was included in the DNA polymerase reaction mixtures, nascent DNA became radiolabeled. Analysis of the 32P-labeled material following treatment of the DNA with tobacco acid pyrophosphatase, bacterial alkaline phosphatase, or ribonuclease T1 revealed the presence of oligoribonucleotide chains averaging 5-7 nucleotides long and beginning with 5' GTP residues. Eight presumptive DNA primase initiation sites were located opposite C4 or C5 sequences 3-9 nucleotides upstream of one of the three closely related hexanucleotides C2A2C2, C2AC3, and C2A2T2. RNA-DNA junctions were found 3-10 nucleotides downstream of DNA primase initiation sites. The results indicate that hexanucleotides having the general formula C2A1-2(C2-3/T2), herein referred to as psi, are involved in promoting termination of DNA synthesis and/or de novo initiation of RNA-primed DNA chains by DNA polymerase alpha-primase.
...
PMID:Mouse DNA polymerase alpha-primase terminates and reinitiates DNA synthesis 2-14 nucleotides upstream of C2A1-2(C2-3/T2) sequences on a minute virus of mice DNA template. 385 59

The nascent DNA synthesized by permeable cells of Bacillus subtilis in the presence of 5'-mercurideoxycytidine triphosphate and 2',3'-dideoxyATP has been isolated and characterized. The newly synthesized DNA was isolated free from other cellular nucleic acids by affinity chromatography on thiol-substituted agarose. The number average chain length of the nascent DNA synthesized in one minute at 25 degrees C was 33 nucleotide residues, due to the chain-terminating action of 2',3'-dideoxyATP. Several lines of evidence indicated that at least 90% of the DNA thus isolated carried a terminally phosphorylated RNA moiety at its 5'-end: (1) the nascent DNA was resistant to exonucleolytic degradation by spleen phosphodiesterase unless first hydrolyzed by strong alkali or ribonuclease; (2) the 5'-termini of nascent DNA could not be phosphorylated by polynucleotide kinase unless first treated with alkaline phosphatase or subjected to hydrolysis by strong alkali or ribonuclease; (3) alkaline hydrolysis of nascent DNA labeled with 32P at the 5'-end released unlabeled DNA with a free 5'-terminus and 32P-labeled ribonucleoside 3',5'-bisphosphates; (4) ribonuclease degradation of similarly labeled material produced an unlabeled DNA-containing polynucleotide fraction and 32P-labeled ribo-oligonucleotides; (5) chromatography on dihydroxyboryl cellulose showed that the RNA moiety lacked a 3'-terminal cis-diol grouping (even after treatment with alkaline phosphatase) unless first subjected to the 3'-exonucleolytic action of bacteriophage T4 DNA polymerase. The sequence of the ribonucleotide chains was elucidated by end-group labeling with polynucleotide kinase and digestion with various ribonucleases. The ribonucleotide moiety was primarily three and four residues in length with the predominant sequence (pp)pApG(pC)1-2pDNA. The possibility that it represents a primer for discontinuous DNA synthesis is discussed.
...
PMID:Analysis of the 5'-termini of nascent DNA chains synthesized in permeable cells of Bacillus subtilis. 618 36

gamma-Irradiation of DNA in vitro produces two types of single strand breaks. Both types of strand breaks contain 5'-phosphate DNA termini. Some strand breaks contain 3'-phosphate termini, some contain 3'-phosphoglycolate termini (Henner, W.D., Rodriguez, L.O., Hecht, S. M., and Haseltine, W. A. (1983) J. Biol. Chem. 258, 711-713). We have studied the ability of prokaryotic enzymes of DNA metabolism to act at each of these types of gamma-ray-induced 3' termini in DNA. Neither strand breaks that terminate with 3'-phosphate nor 3'-phosphoglycolate are substrates for direct ligation by T4 DNA ligase. Neither type of gamma-ray-induced 3' terminus can be used as a primer for DNA synthesis by either Escherichia coli DNA polymerase or T4 DNA polymerase. The 3'-phosphatase activity of T4 polynucleotide kinase can convert gamma-ray-induced 3'-phosphate but not 3'-phosphoglycolate termini to 3'-hydroxyl termini that can then serve as primers for DNA polymerase. E. coli alkaline phosphatase is also unable to hydrolyze 3'-phosphoglycolate groups. The 3'-5' exonuclease actions of E. coli DNA polymerase I and T4 DNA polymerase do not degrade DNA strands that have either type of gamma-ray-induced 3' terminus. E. coli exonuclease III can hydrolyze DNA with gamma-ray-induced 3'-phosphate or 3'-phosphoglycolate termini or with DNase I-induced 3'-hydroxyl termini. The initial action of exonuclease III at 3' termini of ionizing radiation-induced DNA fragments is to remove the 3' terminal phosphate or phosphoglycolate to yield a fragment of the same nucleotide length that has a 3'-hydroxyl terminus. These results suggest that repair of ionizing radiation-induced strand breaks may proceed via the sequential action of exonuclease, DNA polymerase, and DNA ligase. The possible role of exonuclease III in repair of gamma-radiation-induced strand breaks is discussed.
...
PMID:Enzyme action at 3' termini of ionizing radiation-induced DNA strand breaks. 636 Oct 28

T4 DNA polymerase copolymerizes the SP isomers of 2'-deoxyadenosine 5'-O-(1-thiotriphosphate) and 5'-O-(2-thiotriphosphate) with dTTP onto a poly(d(A-T) template in the presence of various metal ions. The corresponding RP diastereomers are inactive, independent of the metal ion used. The polymer resulting from the polymerization of the SP diastereomer of 2'-deoxyadenosine 5'-O-(1-thiotriphosphate) and dTTP can be degraded by the 5' leads to 3' exonuclease activity of Escherichia coli DNA polymerase I and alkaline phosphatase (Brody, R. S., and Frey, P. A. (1981) Biochemistry 20, 1245-1252) to d(Tp(S)A). This material has the RP configuration as determined by comparison with the RP and SP diastereomers obtained by chemical synthesis and preparative separation by high performance liquid chromatography. This result indicates inversion of configuration at the alpha-phosphorus in the nucleotidyl transfer reaction and is compatible with the absence of a covalent enzyme intermediate.
...
PMID:A study of the mechanism of T4 DNA polymerase with diastereomeric phosphorothioate analogues of deoxyadenosine triphosphate. 704 12

The bacteriophage T7 induced DNA polymerase, consisting of the phage specified gene 5 protein associated with Escherichia coli thioredoxin, catalyzes the copolymerization of SP-dATP alpha S with dTTP, producing the alternating of polymer poly[dTs-A)] by a mechanism involving inversion of configuration at P alpha. Degradation of poly[d(5s-A)] by the nucleolytic action of E. coli DNA polymerase produced the dinucleotide pdTps-dA, whose configuration at the phosphorothioate diester was assigned as R by comparison of the phosphorus-31 nuclear magnetic resonance chemical shift (55.0 ppm downfield from H3PO4) with that of an authentic sample. Further degradation by alkaline phosphatase to Rp-dTps-dA (55.6 ppm downfield from H3PO4) confirmed the configuration. The stereochemistry provides no evidence of a double displacement mechanism.
...
PMID:Stereochemical course of nucleotidyl transfer catalyzed by bacteriophage T7 induced DNA polymerase. 709 4

The ligation-independent cloning of PCR products (LIC-PCR) is a versatile and highly efficient cloning procedure resulting in recombinant clones only. Recombinants are generated between PCR products and a PCR-amplified vector through defined complementary single-stranded (ss) ends artificially generated with T4 DNA polymerase. This procedure does not require restriction enzymes, alkaline phosphatase, or DNA ligase. The primers used for amplification contain an additional 12-nucleotide sequence at their 5' ends that is complementary in the vector- and insert-specific primers. The (3'-->5') exonuclease activity of T4 DNA polymerase is used in combination with a predetermined dNTP (dGTP for the inserts and dCTP for the vector) to specifically remove 12 nucleotides from each 3' end of the PCR fragments. Because of the complementarity of the ends that are generated, circularization can occur between vector and insert. The recombinant molecules do not require in vitro ligation for efficient bacterial transformation. To make this technique widely applicable, we have simplified the handling of the PCR fragments prior to LIC. The PCR products do not need further purification following the T4 DNA polymerase treatment. Incubation of vector and insert PCR fragments for as little as 5 min is sufficient for a high yield of recombinants. Comparison of the transformation efficiencies using different-length LIC tails revealed that using 12-nucleotide cohesive ends produced four times more transformants than were obtained with the LIC with 10-nucleotide cohesive ends. When the LIC tails were 8 nucleotides long, no transformants were obtained. PCR fragment purification, T4 DNA polymerase treatment, and LIC is complete in < 1 hr.
...
PMID:Minimal length requirement of the single-stranded tails for ligation-independent cloning (LIC) of PCR products. 758 Sep 2

A new method for typing single nucleotide polymorphisms in DNA is described. In this method, specific fragments of genomic DNA containing the polymorphic site(s) are first amplified by the polymerase chain reaction (PCR) using one regular and one phosphorothioate-modified primer. The double-stranded PCR product is rendered single-stranded by treatment with the enzyme T7 gene 6 exonuclease, and captured onto individual wells of a 96 well polystyrene plate by hybridization to an immobilized oligonucleotide primer. This primer is designed to hybridize to the single-stranded target DNA immediately adjacent from the polymorphic site of interest. Using the Klenow fragment of E. coli DNA polymerase I or the modified T7 DNA polymerase (Sequenase), the 3' end of the capture oligonucleotide is extended by one base using a mixture of one biotin-labeled, one fluorescein-labeled, and two unlabeled dideoxynucleoside triphosphates. Antibody conjugates of alkaline phosphatase and horseradish peroxidase are then used to determine the nature of the extended base in an ELISA format. This paper describes biochemical features of this method in detail. A semi-automated version of the method, which we call Genetic Bit Analysis (GBA), is being used on a large scale for the parentage verification of thoroughbred horses using a predetermined set of 26 diallelic polymorphisms in the equine genome.
...
PMID:Genetic Bit Analysis: a solid phase method for typing single nucleotide polymorphisms. 793 43

A nonradioactive method is developed to detect DNA polymerase activity after sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis containing gapped DNA as template. The technique is based on the use of digoxigenin- or biotin-labeled deoxynucleotides during DNA synthesis, and their detection by means of an anti-digoxigenin antibody-alkaline phosphatase conjugate or by a streptavidin-alkaline phosphatase conjugate. The detection of the DNA polymerase catalytic subunit is achieved after incubation of the gels with colorimetric alkaline phosphatase substrates. The technique is able to detect nanogram amounts of Escherichia coli DNA polymerase I and picogram amounts of its Klenow fragment. The results with other DNA polymerases and E. coli extracts suggest that this colorimetric detection system could be used for the analysis of an extended range of DNA polymerase enzymes. The method presented in this report offers an alternative to the already described radioactive techniques for detection of DNA polymerase activity after SDS-polyacrylamide gel electrophoresis.
...
PMID:Colorimetric detection of DNA polymerase activity after sodium dodecyl sulfate polyacrylamide gel electrophoresis. 798 4


<< Previous 1 2 3 4 Next >>

---