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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)

DNA polymerase gamma and mitochondrial DNA polymerase were isolated from brain nuclei and synaptosomes respectively. The presence of a single DNA polymerase in synaptosomal mitochondria was established by chromatography on DEAE-cellulose, phosphocellulose and DNA-cellulose, as well as by sedimentation analysis and isoelectric focusing. A great similarity between the purified nuclear DNA polymerase gamma and the mitochondrial enzyme was found by the following criteria: chromatographic behaviour in three column systems; essentially complete inhibition by N-ethyl-maleimide (2 mM); optimal requirements of Mn2+ (0.1 mM), Mg2+ (5 mM) and pH (8.0); template preferences, poly(A) - (dT)20-25 larger than activated DNA larger than poly(dA) - (dT)12-18; lack of activity on single-stranded polynucleotides and (dT)12-primed mRNA; molecular weight (180000), sedimentation (9.2 S) and isoelectric point (pI 5.4). We therefore conclude that brain nuclear DNA polymerase gamma and synaptosomal mitochondrial DNA polymerase are closely related and may even be identical.
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PMID:Identity of DNA polymerase gamma from synaptosomal mitochondria and rat-brain nuclei. 59 67

Four DNA polymerases from the marine diatom Cylindrotheca fusiformis, polymerases A, B, C and D, were further differentiated by their subcellular localization, presence of deoxyribonuclease activity, apparent heterogeneity and molecular weights. Polymerases A, B and D occur in significant amounts in the soluble fraction, suggesting that they were originally localized in the nuclei, whereas polymerase C predominates in the chloroplasts. A mitochondrial DNA polymerase was also isolated and characterized by ion-exchange chromatography. Polymerase D has an associated nuclease activity which prefers denatured DNA and Mg2+, and has a pH optimum higher than that for polymerase activity. Co-elution from a DEAE-Sephadex column and co-sedimentation in glycerol density gradients of deoxyribonuclease and polymerase D activity suggest a molecular association. Polymerases A, B and C are devoid of nuclease activity. Glycerol-gradient-sedimentation analysis showed that all DNA polymerase fractions are heterogeneous at low ionic strengths, with the appearance of a single homogeneous activity of 0.5M-KCl. Estimated molecular weights of 100000, 82000 and 120000 for polymerases A, B and C respectively were obtained from sedimentation analysis and gel filtration. Polymerase D was estimated to have a molecular weight of about 100000 as determined by sedimentation analysis alone.
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PMID:The deoxyribonucleic acid polymerases from the diatom Cylindrotheca fusiformis. Subcellular distribution, exonuclease activity and heterogeneity of the enzymes. 60 24

DNA-dependent DNA polymerase has been extracted from the soluble cytoplasmic fraction of regenerating rat liver and purified using phosphocellulose and DEAE-cellulose chromatography. Glycerol gradient analysis showed that the enzyme was predominantly DNA polymerase alpha, having a sedimentation coefficient of 10.5 S at low ionic strength and of 6--8 S at higher salt concentrations. The fidelity of purified enzyme was assessed using the co-polymer poly(dA-dT).poly(dA-dT) as a template for DNA synthesis. For both the aggregated (10.5 S) and disaggregated (6--8 S) forms, fidelities in the range of 1 wrong base in 100,000--150,000 complementary bases were obtained.
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PMID:Molecular size and fidelity of DNA polymerase alpha from the regenerating liver of the rat. 62 56

A low molecular weight DNA polymerase which sediments at 3.3 S on sucrose gradients has been purified from total cell homogenates of rapidly dividing embryos of the sea urchin Strongylocentrotus purpuratus. In the presence of 2 mM N-ethylmaleimide, it is the major polymerase activity in whole cell homogenates when assayed with an oligo(dT)10.poly(dA)200 template; a template which it uses about 200 times more efficiently than activated DNA. The requirement for N-ethylmaleimide exists only in crude cell fractions where it acts to inhibit a template digesting nuclease activity. The polymerase is highly stable if maintained in the presence of 20% glycerol, is completely dependent on added template, and shows no end addition activity. The physical and enzymatic properties of this enzyme clearly distinguish it from the DNA polymerase previously described by Loeb (Loeb, L. A. (1969) J. Biol. Chem. 244, 1672-1681) which sediments as a high moeluclar weight (5.6 to 6.6 S) enzyme and prefers the activated DNA template. In addition, these two DNA polymerase enzymes show distinctive chromatographic properties using DEAE-cellulose and phosphocellulose columns as well as their sensitivity to N-ethylmaleimide. The properties of the low molecular weight polymerase indicate close similarity to the beta-polymerase isolated from mammalian cells. These low molecular weight enzymes are both sensitive to phosphate salt and able to utilize the artificial ribohomopolymer template oligo(dT)10.poly(rA)200. A quantitative analysis of the low molecular weight DNA polymerase during early embryonic development indicates that the activity of this enzyme increases at least 2-fold immediately following fertilization and again during early blastula stage (hatching). Such quantitative changes in a beta enzyme activity are in contrast to findings with the alpha-polymerase which remains constant during early development.
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PMID:A low molecular weight DNA polymerase beta in the sea urchin Strongylocentrotus purpurantus. Partial purification, properties, and changes in development. 71 48

Three ribonucleotidyl transferase types have been described in the sea urchin: riboadenylate trnasferase, the DNA dependent RNA polymerases, and a DNA polymerase associated ribonucleotidyl transferase (Biochemistry 15:3106-3113, 1976). In the present work this latter ribonucleotidyl transferase was found to purify with DNA polymerase alpha through phosphocellulose, DEAE-Sephadex and DNA cellulose and to cosediment at 6.5 S. This ribonucleotidyl transferase was active with Mn+2, but not Mg+2, on calf thymus DNA and poly(dC). Other synthetic templates elicited DNA polymerase alpha but no ribonucleotidyl transferase activity. From alkaline hydrolysates of the poly(dC) directed GTP polymerization, we found Goh and Gp in a ratio of 1:16 indicating an average chain length of 17 residues after a 20 min reaction. Co-polymerization of GTP (5 micrometer) and dGTP (10 micrometer) yielded a non-random distribution of the ribonucleotide in the deoxyribonucleotide. The properties of this urchin ribonucleotidyl transferase are unlike any previously described eukaryotic transferase and the data is discussed with reference to the known properties of E. coli DNA polymerase I and the primase.
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PMID:Ribonucleotidyl transferase in preparations of partially purified DNA polymerase alpha of the sea urchin. 72 5

The enzyme which catalyses template independent synthesis of polydeoxynucleotides from deoxynucleoside diphosphates was separated from E. coli DNA polymerase I by DEAE-cellulose chromatography followed by ultrafiltration through the M-50 Amicon filter. The ultrafiltration data indicate that the molecular weight of the enzyme is not higher than 50,000. The enzyme is not able to use deoxynucleoside triphosphates, ribonucleoside di- or triphosphates as substrates for the polymerization. The reaction of template independent polymerization proceeds with a lag period varying from 2 to 20 hours (for different preparations of enzyme) and is activated by Mg2+ (the optimal concentration 1-2 . 10(-3) M). The pH optimum of the reaction is at 8.5. The optimal concentration of deoxyribonucleoside diphosphates is 10(-3) M, and its increase strongly inhibits polymerization. The enzyme was supposed to be called deoxynucleoside diphosphate: olygonucleotide deoxynucleotidyltransferase (catalyzing polymerization without template). The presence of the enzyme in the preparations of E. coli DNA-polymerase I can explain the ability of the latter to catalyze the untemplated synthesis of poly dG : poly dC.
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PMID:[Separation of the enzyme catalyzing polymerization of deoxyribonucleoside diphosphates from preparations of E. coli DNA-polymerase I]. 80 82

A cellular factor which makes T7 DNA irradiated with gamma-rays a better primer for Micrococcus DNA polymerase was partially purified by DEAE and phosphocellulose column chromatography and named "primer activating enzyme". Sucrose density gradient sedimentation analysis was carried out to examine actions of one major active fraction that appeared by phosphocellulose chromatography. It was shown that this factor introduced new nicks in T7 DNA in addition to those introduced directly by gamma-ray irradiation. This enzyme fraction also had an endonucleolytic activity towards DNA containing apurinic sites induced by heat treatment and had capacity to enhance the priming activity of heat- or methyl methansulfonate-treated DNA but affected very little that of ultraviolet-irradiated DNA. This enzyme had no effect on T7 DNA when it was not treated with the DNA-damaging agents. From these results we concluded that this enzyme may be analogous to the endonuclease II or apurinic site-specific enconuclease of Escherichia coli.
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PMID:Studies on DNA repair in Bacillus subtilis. II. Partial purification and mode of action of an enzyme enhancing the priming activity of gamma-irradiated DNA. 80 55

The subcellular location and growth stage dependence of the DNA polymerases of Euglena gracilis strain Z and of a bleached derivative of the strain have been studied by fractionation of the enzymes from extracts of whole cells and subcellular fractions on DEAE-cellulose. A new method for the rapid isolation of nuclei was employed. Of the major enzymes, pol A has a predominantly nuclear location and pol B a predominantly cytoplasmic location. Pol A is 4-fold and pol B 15-fold more active in exponentially-growing cells than in stationary-phase cells, pol B representing 90% of the combined activities in exponential-phase cells. The activity of the mitochondrial DNA polymerase increases about 3-fold as the cells enter stationary phase while that of the chloroplast DNA polymerase is greater in exponential-phase cells. The chloroplast enzyme persists in cells which have been reversibly bleached. The results are compared to those of similar experiments involving primitive and higher eucaryotes.
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PMID:Subcellular location and growth stage dependence of the DNA polymerases of Euglena gracilis. 81 Jun 22

A simple and reproducible procedure is described which allows the fast and almost quantitative removal of DNA polymerases I and II from DNA polymerase III, in crude extracts of polA+ strains of Bacillus subtilis. The procedure entails streptomycin sulfate and ammonium sulfate fractionations; subsequent analysis of the partially purified preparation by G-200 chromatography, DEAE cellulose chromatography and density gradient sedimentation, shows that the ammonium sulfate fraction contains less than 5% of the total activity as DNA polymerase I and less than 2% as DNA polymerase II. The purification procedure, up to the ammonium sulfate step, was utilized for the analysis of the level of DNA polymerase III in several B. subtilis mutants, with results comparable to those obtained from the corresponding polA- strains following more cumbersome purification procedures. The M.W. of the purified form is of 227.000, somewhat greater than the published values. The early fractions of the purification have revealed the existence of a form with a M.W. of 426.000; the nature of this form, which has been observed in several instances and which is very unstable and short-lived, is under investigation.
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PMID:A simplified procedure for the analysis of DNA polymerase III levels in Bacillus subtilis strains. 82 85

A DNA-dependent DNA polymerase from rat liver mitochondria was partially purified and characterized. Mitochondrial DNA polymerase has been found to be quite different from other DNA-dependent DNA polymerases alpha and beta present in the rat liver in the following points: elution patterns in a DEAE-cellulose column chromatography, sedimentation coefficients determined by the glycerol gradient centrifugation in the presence of high salt, and sensitivities to N-ethylmaleimide, ethidium bromide and KCl.
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PMID:Mitochondrial DNA polymerase from rat liver. 85 77


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