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:2.7.7.7 (DNA polymerase)
17,007 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A novel method for detecting possible DNA repair enzymes on sodium dodecyl sulfate-polyacrylamide gels by blotting them onto a damaged DNA-fixed membrane is presented. To prepare the membrane, highly polymerized calf thymus DNA immobilized on a nylon membrane is damaged chemically. Enzymes, either homogeneous or crude, that are possibly involved in the priming step of DNA repair are fractionated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and are renatured to active form by incubating the gel in an appropriate buffer. The renatured enzyme is then blotted onto the damaged DNA-fixed membrane, a process during which incision and/or excision are introduced to the damaged DNA by the enzymes. The incision and/or excision provide priming sites for repair DNA synthesis in the subsequent step in which the membrane is incubated with DNA polymerase in the presence of alpha-32P-labeled substrate. The site of substrate incorporation on the membrane reflecting the molecular weight of the repair enzyme is finally visualized by autoradiography. The present technique is established using Escherichia coli exonuclease III and a DNA-fixed membrane treated with bleomycin or acid-depurinated. By application of this method, a priming factor (an exonuclease) involved in the initiation of bleomycin-induced DNA repair is detected in the extract of mouse ascites sarcoma cells, and thus the molecular weight of the enzyme is estimated. Some apurinic/apyrimidinic endonucleases of mammals are also detected by the present procedure.
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PMID:Detection of possible DNA repair enzymes on sodium dodecyl sulfate-polyacrylamide gels by protein blotting to damaged DNA-fixed membranes. 171 Aug 77

A new class of compounds, 9-[(2RS)-3-fluoro-2-phosphonylmethoxypropyl] [(RS)-FPMP] derivatives of purines, is described that has selective activity against a broad spectrum of retroviruses [including human immunodeficiency virus type 1 (HIV-1) and type 2 (HIV-2)] but not other RNA or DNA viruses. This activity spectrum is completely different from that of the parental compounds, 9-[(2S)-3-hydroxy-2-phosphonylmethoxypropyl] [(S)-HPMP] derivatives of purines, which are active against a broad range of DNA viruses. The racemic (RS)-FPMP derivatives of adenine and 2,6-diaminopurine, termed (RS)-FPMPA and (RS)-FPMPDAP, respectively, are markedly more selective as in vitro antiretroviral agents than their 9-(2-phosphonylmethoxyethyl) (PME) counterparts, PMEA and PMEDAP. Also, (RS)-FPMPA and (RS)-FPMPDAP have a substantially higher therapeutic index in mice in inhibiting Moloney murine sarcoma virus-induced tumor formation and associated death and are markedly less inhibitory to human bone marrow cells than PMEA and PMEDAP. The diphosphate derivative of (RS)-FPMPA [(RS)-FPMPApp] is a potent and selective inhibitor of HIV-1 reverse transcriptase but not of HSV-1 DNA polymerase or DNA polymerase alpha. (RS)-FPMPApp, akin to PMEA diphosphate (PMEApp), acts as a DNA chain terminator. The DNA chain-terminating properties of PMEApp and (RS)-FPMPApp seem to be a prerequisite for acyclic nucleoside phosphonates to exhibit antiretrovirus (i.e., anti-HIV) activity.
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PMID:9-[(2RS)-3-fluoro-2-phosphonylmethoxypropyl] derivatives of purines: a class of highly selective antiretroviral agents in vitro and in vivo. 171 Dec 14

1. DNA damage by peplomycin, an antitumor antibiotic, and its repair by cellular enzymes were studied using pUC18 plasmid DNA. The DNA damage and repair were measured by monitoring the conformational changes of pUC18 DNA. 2. Peplomycin-induced DNA damage was enhanced by addition of ferrous ion and inhibited by deferoxamine, a specific iron chelator, suggesting iron-requirement for the DNA damage. 3. DNA damage by peplomycin was inhibited by superoxide dismutase in both native and heat-inactivated forms, possibly due to non-enzymatic interaction. 4. Peplomycin-induced, single-strand breaks in pUC18 DNA was repaired by incubating with a priming factor (an exonuclease purified from mouse ascites sarcoma cells), DNA polymerase beta, four deoxynucleoside triphosphates, T4 DNA ligase and ATP. The average repair patch size was estimated to be approximately four nucleotide length.
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PMID:DNA damage by peplomycin and its repair in an in vitro system. 171 61

DNA repair synthesis induced in permeable mouse ascites sarcoma cells by peplomycin, an antitumor antibiotic, was studied. Mouse ascites sarcoma (SR-C3H/He) cells were permeabilized with a low concentration of Triton X-100 in an isotonic condition. Permeable cells were treated with an appropriate concentration of peplomycin to introduce single-strand breaks in permeable cell DNA. DNA repair synthesis in peplomycin-treated permeable cells was measured by incubating the cells with four deoxynucleoside triphosphates in an appropriate buffer system. The DNA repair synthesis was enhanced by ATP and NaCl at near physiological concentrations. More than 90% of DNA synthesis in the present system depended on the peplomycin-treatment. The repair nature of the DNA synthesis was confirmed by a BrdUMP density shift technique. The repair patches were largely completed and ligated in the presence of ATP. Analyses using selective inhibitors for DNA polymerases showed that both DNA polymerase Beta and aphidicolin-sensitive DNA polymerases (DNA polymerase alpha and/or delta) were involved in the repair DNA synthesis.
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PMID:Peplomycin-induced DNA repair synthesis in permeable mouse ascites sarcoma cells. 171 30

A mouse repair enzyme having priming activity on bleomycin-damaged DNA for DNA polymerase was purified to apparent homogeneity and characterized. The enzyme extracted from permeabilized mouse ascites sarcoma (SR-C3H/He) cells with 0.2 M potassium phosphate buffer (pH 7.5) was purified by successive chromatographies on phosphocellulose, DEAE-cellulose, phosphocellulose (a second time), Sephadex G-100, single-stranded DNA cellulose and hydroxyapatite. The purified enzyme has an Mr of 34,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Enzymatical studies indicated that it is a multifunctional enzyme having exonuclease, apurinic/apyrimidinic endonuclease and phosphatase activities, similar to Escherichia coli exonuclease III. This enzyme is tentatively designated as APEX nuclease for apurinic/apyrimidinic endonuclease and exonuclease activities. The amino acid composition, amino-terminal amino acid sequence and an internal amino acid sequence of APEX nuclease are determined.
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PMID:A mouse DNA repair enzyme (APEX nuclease) having exonuclease and apurinic/apyrimidinic endonuclease activities: purification and characterization. 171 53

Repair of X-ray-induced single-strand breaks of DNA was studied in vitro using an exonuclease purified from mouse ascites sarcoma (SR-C3H/He) cells. X-ray-dose-dependent unscheduled DNA synthesis was primed by the exonuclease. Repair of X-ray-induced single-strand breaks in pUC19 plasmid DNA was demonstrated by agarose gel electrophoresis after incubating the damaged DNA with the exonuclease, DNA polymerase (Klenow fragment of DNA polymerase I or DNA polymerase beta purified from SR-C3H/He cells), four deoxynucleoside triphosphates, ATP and DNA ligase (T4 DNA ligase or DNA ligase I purified from calf thymus). The present results suggested that the exonuclease is involved in the initiation of repair of X-ray-induced single-strand breaks in removing 3' ends of X-ray-damaged DNA.
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PMID:Repair of X-ray-induced single-strand breaks by a cell-free system. 237 79

Enzymological properties of the RNA-directed DNA polymerase associated with the Suncus murinus mammary tumor virus (Sm-MTV) was investigated and its antigenic relatedness to other retroviral DNA polymerases was examined. The enzyme exhibited higher activity in the presence of Mg2+ than in the presence of Mn2+ with endogenous RNA as well as with almost all of the synthetic template X primers tested. Mg2+ was also effective with poly(2'-O-methylcytidylate) X oligodeoxyguanylate which was known to be specific for Mn2+. To examine the immunological relatedness of this enzyme with other retroviral DNA polymerases, remaining Sm-MTV DNA polymerase activity was measured after treatment of this enzyme with various antisera prepared against each of the reverse transcriptases of Mason-Pfizer monkey virus (MPMV), murine mammary tumor virus (MuMTV), simian sarcoma virus-simian sarcoma associated virus (SSV/SSAV), and Rauscher murine leukemia virus (RLV). No inhibition of the Sm-MTV enzyme activity was observed when treated with the latter three antisera with which the DNA polymerase activities of the corresponding retroviruses were fully inhibited. Only the antiserum against MPMV-enzyme, however, was found to slightly inhibit the Sm-MTV enzyme activity. These results indicate that Sm-MTV DNA polymerase has similar enzymological properties to those of MPMV and MuMTV and shares some common antigenic determinant group(s) with MPMV DNA polymerase.
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PMID:Biochemical and immunological characterization of Suncus murinus mammary tumor virus DNA polymerase. 241 16

2',3'-dideoxy-2',3'-dehydrothymidine 5'-triphosphate (dddTTP) shows termination substrate properties in the DNA synthesis catalyzed by E. coli DNA polymerase I KF, rat liver DNA polymerase beta, reverse transcriptases of avian myeloblastosis virus and Raus sarcoma virus and calf thymus terminal deoxynucleotidyl transferase. This implies that the mononucleotide residue of dddTTP incorporates into 3'-termini of newly synthesized DNA chains. However, dddTTP has no influence on the DNA synthesis catalyzed by calf thymus DNA polymerase alpha. In the case of some DNA polymerases dddTTP was one order of magnitude more effective in comparison with the other known termination substrates.
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PMID:Properties of 2',3'-dideoxy-2',3'-dehydrothymidine 5'-triphosphate in terminating DNA synthesis catalyzed by several different DNA polymerases. 243 82

Effects of ATP and some other nucleotides (AMP, ADP, CTP, GTP, UTP and dATP) on reparative DNA synthesis and repair patch ligation in bleomycin-pretreated permeable mouse sarcoma cells were studied. Reparative DNA synthesis was significantly stimulated by 2.5 mM ATP, ADP or dATP. The stimulation was observed on both DNA polymerase alpha- and beta-dependent reparative DNA synthesis. ATP concentration required for repair patch ligation was much lower than that required for the stimulation of reparative DNA synthesis. An apparent Km value for ATP of the repair patch ligation was about 40 microM. ADP supported repair patch ligation after being converted into ATP by adenylate kinase in permeable cells.
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PMID:Effects of ATP and other nucleotides on DNA repair synthesis in bleomycin-pretreated permeable mouse sarcoma cells. 244 62

To investigate factors involved in excision repair DNA synthesis, a soluble extract was prepared from permeable mouse sarcoma (SR-C3H/He) cells by homogenization and ultracentrifugation. DNA synthesis measured by using native calf thymus DNA as the template-primer and the extract as the polymerase source showed low activity. The DNA synthesis was enhanced more than ten-fold by the addition of an appropriate concentration of bleomycin, a radiomimetic DNA-damaging drug. Using selective inhibitors of DNA polymerases, it was shown that the DNA polymerase involved in the bleomycin-induced DNA synthesis was DNA polymerase beta. In addition to DNA polymerase beta, an exonuclease which converts bleomycin-damaged DNA into suitable template-primers for repair DNA synthesis appeared to be present in the permeable cell extract.
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PMID:Bleomycin-induced DNA synthesis in a cell-free system using a permeable mouse sarcoma cell extract. 244 66


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