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)

Evidence is presented that the gene A and A * proteins of bacteriophage phi X 174 form covalent associations with the 5' ends of the DNA molecules when superhelical phi X replicative form DNA is nicked by a combination of these proteins in vitro. This evidence is: 1, The 5' ends of the DNA molecules nicked by the gene A protein and reacted with bacterial alkaline phosphatase were protected against subsequent phosphorylation by polynucleotide kinase even after treatment of the nicked DNA with SDS and pronase followed by centrifugation on a high-salt neutral sucrose gradient. 2, Iodinated pronase-sensitive material remained attached to the nicked replicative form DNA and could not be removed by exposure to SDS or 2 M NaCl, either by sedimentation through high-salt neutral sucrose gradients, or by CsCl equilibrium centrifugation. 3, Iodinated pronase-sensitive material was detected on DNA that had been nicked during the reaction, but not on unreacted DNA. 4, Electrophoresis of the iodinated pronase-sensitive, DNA-bound material in SDS-polyacrylamide gels after DNAse digestion revealed that it was composed almost entirely polypeptides with electrophoretic mobilities similar to those of the gene A and A * proteins. We speculate that the gene * protein may be essential for normal progeny single-stranded DNA synthesis in vivo.
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PMID:The mechanism of replication of phi X 174. XVIII. Gene A and A* proteins of phi X 174 bind tightly to phi X 174 replicative form DNA. 626 20

Phage phi 29 DNA cannot be phosphorylated with polynucleotide kinase and [gamma-32P]ATP because of the presence of a viral protein covalently linked to the 5' termini. The 5' ends can, however, be made susceptible to phosphorylation by treatment with alkali and alkaline phosphatase. Restriction fragments Hpa II C and Hpa II F, corresponding to the right and left ends of phi 29 DNA, respectively, were labeled at the 5' ends with polynucleotide kinase and [gamma-32P]ATP or at the 3' ends with terminal transferase and [alpha-32P]ATP or [alpha-32P]cordycepin 5'-triphosphate. After a secondary cleavage of the labeled fragments, the sequence of the first 150-180 nucleotides at the termini of phi 29 DNA was determined by the method of Maxam and Gilbert. The ends of phi 29 DNA are flush, and a six-nucleotides-long inverted terminal repetition was found. The functional implications of the sequences determined are discussed.
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PMID:Nucleotide sequence at the termini of the DNA of Bacillus subtilis phage phi 29. 626

Endonuclease V of bacteriophage T4 has been purified to physical homogeneity from T4D-infected Escherichia coli 1100. The enzyme, whose molecular weight is 16,000, possessed two distinct catalytic activities, a pyrimidine dimer-DNA glycosylase and an apurinic/apyrimidinic endonuclease. They acted on UV-irradiated poly(dA) . poly(dT) in a sequential manner; the glycosylase cleaved the N-glycosyl bond between the 5'-pyrimidine of a dimer and the corresponding sugar and then the endonuclease hydrolyzed a phosphodiester bond on the 3'-side of the apyrimidinic site. The 5'-termini thus generated were phosphorylated by T4 polynucleotide kinase only after they had been subjected to direct photoreversal and then treated with alkaline phosphatase. By using two phage mutants, uvs-5 and uvs-13, it was shown that occurrence of an amber mutation in the denV gene caused a simultaneous loss of the two activities. Suppression of the mutation of uvs-5 rendered both activities partially active. When the mutation of uvs-13 was suppressed, a mutant form of enzyme that possessed only a glycosylase activity was produced. This suggests that there are two distinct domains in a single enzyme, each of which corresponds to one of the activities.
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PMID:Purification and characterization of normal and mutant forms of T4 endonuclease V. 627 6

The adenovirus-specific DNA-binding protein (DBP) has been shown to inhibit the hydrolysis of single-stranded DNA by a DNase isolated from KB cells, (Nass, K., and Frenkel, G.D. (1980). J. Virol. 35, 314-319). The specificity of the inhibition has now been investigated. The DBP inhibits the hydrolysis of single-stranded DNA by several different DNases (DNase II, KB DNase, S1 nuclease) under a variety of reaction conditions, but it has no effect on DNase I-catalyzed hydrolysis of single-stranded DNA. The DBP also inhibits the rate of hydrolysis of double-stranded DNA by KB DNase and DNase II, but has no effect on DNase I-catalyzed hydrolysis of this substrate. The DBP also inhibits the dephosphorylation of 5'-phosphoryl-terminated DNA by bacterial alkaline phosphatase but stimulates the phosphorylation of 5'-hydroxyl-terminated DNA by polynucleotide kinase.
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PMID:DNase inhibition by the adenovirus DNA-binding protein exhibits specificity for the enzyme but not for the secondary structure of the DNA. 630 53

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.
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PMID:Enzyme action at 3' termini of ionizing radiation-induced DNA strand breaks. 636 Oct 28

The structures of the 5' termini of the protamine mRNAs (PmRNAs) have been investigated by inhibiting their translation in wheat-germ extracts in the presence of 7-methyl guanosine 5'-phosphate (m7-GMP), an analogue of 'cap' structure in mRNAs. Second, the cap structures on PmRNAs were examined by labelling the RNA at the 5' end with T4 polynucleotide kinase and [gamma-32P]ATP before and after removal of these structures with tobacco acid pyrophosphatase and alkaline phosphatase. The results indicate that cap structures of the PmRNAs are heterogeneous. It appears that the mRNAs coding for protamine components CI and CIII have at least a cap 1 structure while the mRNAs coding for CII do not appear to be capped or methylated.
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PMID:Studies on the heterogeneity of the 5' ends of the protamine mRNAs from rainbow trout testis. 728 75

8-Methoxypsoralen is a bifunctional furocoumarin used in human photochemotherapy. It can form two kinds of DNA adduct, monoadducts and interstrand cross-links. These adducts have been detected by 32P-postlabelling using hydrolysis with DNase1, alkaline phosphatase and snake venom phosphodiesterase, and longer labelling than usual, with more T4 polynucleotide kinase. Using preliminary two-dimensional chromatography (D1, D2) followed by transfer of adducts to separate PEI cellulose sheets for further development (D3, D4), we observed three spots corresponding to the adducts sought. Two experiments (dose-effects and shift of radioactivity) have confirmed the origin of the three spots.
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PMID:Characterization by 32P-postlabelling of 8-methoxypsoralen adducts. 822 76

In mammalian cells DNA synthesis is more complicated than in prokaryotes and less well understood. Here we incubated intact mammalian cells (polyamine auxotrophic Chinese hamster ovary cells and primary human fibroblasts) with [32P]orthophosphate and found that, besides high molecular weight DNA, a species of low molecular weight DNA, approximately 450 bp in size, became efficiently labeled. The short DNA was labeled first, and in pulse-chase experiments the labeling was transient. The isolated small DNA fragments (RNase A-treated) were phosphorylated by T4 polynucleotide kinase specific for polynucleotides with 5'-OH ends. A polynucleotide kinase phosphorylating these DNA pieces was also detected in nuclear extracts of the cells. Treatment with alkaline phosphatase removed most of the 32P label incorporated into the small DNA in vivo. Labeling with deoxyribonucleosides did not reveal these fragments. We hypothesize that the low molecular weight DNA represents Okazaki fragments and that the mammalian DNA replication machinery includes a polynucleotide kinase phosphorylating the 5'-termini of Okazaki fragments. This would imply a novel step in DNA synthesis. We also show that depriving cells of polyamines reversibly blocks synthesis of high molecular weight DNA and leads to accumulation of the short DNA pieces, suggesting a role for polyamines in joining the Okazaki fragments.
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PMID:Phosphorylation of Okazaki-like DNA fragments in mammalian cells and role of polyamines in the processing of this DNA. 860 90

A combination of T4 polynucleotide kinase, Escherichia coli alkaline phosphatase, yeast Saccharomyces cerevisiae capping enzyme consisting of alpha (RNA guanylyltransferase) and beta (RNA 5'-triphosphatase) subunits. and its alpha subunit without RNA 5'-phosphatase activity was used to establish a simple enzymatic method for determination of RNA species with 5'-hydroxyl, 5'-monophosphate, 5'-diphosphate or 5'-triphosphate termini. Using this method, we found that viral genome RNA (vRNA) segments of both A-type and C-type influenza viruses carry tri- or diphosphates at their 5' termini. The conclusion was based on the observations that: (i) 5' phosphorylation of vRNAs by T4 polynucleotide kinase takes place only after phosphatase treatment; and (ii) capping of vRNAs can be observed with both the intact yeast capping enzyme and its alpha subunit alone devoid of RNA 5'-triphosphatase activity; but (iii) the level of capping is higher for the alphabeta holoenzyme than the alpha subunit though the relative level varies depending on RNA preparations. The results support the de novo initiation for the RNA replication although transcription of influenza vRNAs is initiated by host cell capped RNAs as primers.
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PMID:Identification of the 5' terminal structure of influenza virus genome RNA by a newly developed enzymatic method. 972 72

Single- and double-strand breaks bearing 3'-phosphoglycolate termini are among the most frequent lesions formed in DNA by ionizing radiation and other oxidative mutagens. In order to obtain homogeneous preparations of defined 3'-phosphoglycolate substrates for repair studies, 5'-(32)P-end-labeled partial duplex DNAs were treated with bleomycin, and individual cleavage products were isolated from polyacrylamide gels. The fragments were then treated with alkaline phosphatase and further purified by reverse-phase HPLC. Electrospray ionization Fourier transform ion cyclotron resonance (ESI-FTICR) mass spectrometry of the purified oligomers produced molecular ions of the expected masses, with no detectable contaminants. Gas-phase sequencing by tandem mass spectrometry of these single species yielded the expected sequence ions and confirmed the presence of phosphoglycolate on the 3'-terminal fragments only. The fragments could be relabeled with polynucleotide kinase to yield highly purified, high-specific-activity substrates for repair studies.
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PMID:Homogeneous preparations of 3'-phosphoglycolate-terminated oligodeoxynucleotides from bleomycin-treated DNA as verified by electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry. 1116 22


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