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)

Poly(dT) products which were synthesized depending on (rA)n . (dT)12-18 as a template . primer by mammalian DNA polymerases beta and gamma were analyzed by alkaline sucrose gradient centrifugation. The size of the population of poly(dT) chains synthesized by DNA polymerase beta increased slowly and consistently during incubation up to at least 30 min. On the other hand, the product size with DNA polymerase gamma reached the final size (7 s) within 5 min and the number of products increased during further incubation. Comparison of product number per enzyme molecule suggests that DNA polymerase beta acts on multiple primers in a distributive fashion while DNA polymerase gamma completes poly(dT) chains of large size in a one-by-one fashion.
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PMID:Difference in the mechanisms of poly(dT) synthesis by DNA polymerases beta and gamma. 45 49

DNA polymerase was purified 1000-fold from the cytoplasm of microplasmodia of the myxomycete Physarum polycephalum. The activity was found in two forms exhibiting molecular weights of 204000 and 116000 respectively. Both forms eluted together from DNA-cellulose and DEAE-Sephadex columns. The Stokes radii were 6.5 and 5.5 nm. The sedimentation coefficients were 7.6 and 5.2 S. The frictional ratios of 1.69 suggest a highly hydrated and/or an asymmetric structure of the molecule. The enzyme-catalyzed reaction was sensitive to N-ethylmaleimide (60% inhibition by 1 mM). Unlike DNA polymerase alpha from mammalian cells the Physarum enzyme was stimulated by 30 mM NaCl. Activated DNA was the preferred template. Poly(A) . (DT)12 was not accepted. The Km value for deoxynucleoside triphosphates was 3 micron, for activated DNA 50 microgram/ml and for Mg2+ at the optimum [k+] of 150 mM about 0.6 mM.
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PMID:Deoxyribonucleic acid polymerase from Physarum polycephalum. Properties of the major cytoplasmic activity in exponentially growing microplasmodia. 56 99

Avian erythroid cells were separated into five developmental stages by sedimentation on discontinuous isotonic albumin gradients. Solubilized enzyme activities from whole cells were partially purified and characterized by ion exchange and ion filtration chromatography and velocity sedimenttation analysis. Three nucleotide polymerase types were investigated: (a) DNA-dependent RNA polymerases; (b) RNA-dependent terminal ribonucleotidyltransferases, and (c) DNA-dependent DNA polymerases. The two characteristic forms of eucaryotic DNA-dependent RNA polymerases, polymerase I (nucleolar) and polymerase II (nucleoplasmic), were identified. Polymerase III was only marginally detectable even in the earliest developmental populations. At least two species of RNA-dependent terminal ribosyltransferases were present. One apparently was the poly(A) polymerase observed in other systems. The other terminal transferase was present in two chromatographic forms, required an RNA primer, and used UTP and/or CTP as particularly efficient substrates. Three DNA polymerase activities were resolved, two of which were characteristic of the alpha and beta DNA polymerases described in other eucaryotic systems. The third polymerase was not the gamma polymerase but a separate entity. Poly(dC)-dependent RNA polymerase activity, associated with the alpha polymerase, was relatively enriched in the third DNA polymerase species. The activity levels of the nucleotide polymerases were monitored as a function of red cell maturation. Characteristic declining patterns of activity were obtained for each enzyme which correlate well with the synthetic rates of their in vivo products where these are known. These results correlate well with the synthetic rates of their in vivo products where these are known. These results are consistent with the postulate that the general transcriptive and replicative control processes operating during development may involve changes in the level of the requisite polymerases.
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PMID:Nucleotide polymerases in the developing avian erythrocyte. 83 21

A study of the inhibition of mouse cellular DNA polymerases by poly-nucleotides and their vinyl analogs is presented. Poly(dT)-directed poly(dA) synthesis by representatives of all three classes of cellular DNA polymerase could be completely inhibited by poly(9-vinyladenine), although higher concentrations were required in the case of the gamma class enzyme. Studies on the mechanism of the inhibition using the alpha class DNA polymerase and different templates showed that the enzyme activity was inhibited in all cases where base-pairing between the vinyl polymer and the template occurred; poly(9-vinyladenine) did not interfere with the replication of templates to which it does not bind. The inhibition occurred shortly after addition of poly(9-vinyladenine) to ongoing reactions, yet the enzyme was not displaced from the template - primer complex.
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PMID:Template specific inhibitor of mammalian DNA polymerases. 94 16

We have compared the relative inhibitory activity of poly (A) with its analogues poly N6-isopentenyl adenylic acid (poly(i6 A)) and poly N6-benzyl adenylic acid (poly(bzl6A)), and of poly (U) with its analogue poly 2'-fluoro-2'-deoxyuridylic acid (poly(dUfl)), against DNA polymerase, alpha, beta and gamma and terminal deoxynucleotidyl transferase from human cells and two oncorna virus DNA polymerases. Although poly (A) and its analogues were equally inhibitory against endogenous RNA-directed DNA polymerases of murine and feline leukemia viruses, the analogues in contrast to poly (A) were strongly inhibitory against all four cellular enzymes. Poly (dUfl), on the other hand, was up to 100-fold more potent than poly (U) against both viral and cellular enzymes. Since poly (U) at 100 mug/ml and poly (dUfl) at 1 mug/ml had no effect on terminal deoxynucleotidyl transferase while inhibiting other enzymes by 80--100 per cent these polymers could be useful in the characterization and assay of terminal deoxynucleotidyl transferase. In addition, the polymers such as poly (igA) and poly (bzl5A) which were strongly inhibitory to all cellular enzymes, could be useful in cancer chemotherapy if taken up preferentially by the malignant calls due to their high pinocytic activity. The results also demonstrate potential for large variation in inhibitory activity of polyribonucleotides as related to their chemical composition.
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PMID:Modified nucleotide polymers as inhibitors of DNA polymerases. 119 5

Poly(L-malate) is an unusual polyanion found in nuclei of plasmodia of Physarum polycephalum. We have investigated, by enzymatic and fluorimetric methods, whether poly(L-malate) and structurally related polyanions can interact with DNA-polymerase-alpha-primase complex and with histones of P. polycephalum. Poly(L-malate) is found to inhibit the activities of the DNA-polymerase-alpha-primase complex and to bind to histones. The mode of inhibition is competitive with regard to DNA in elongation and noncompetitive in the priming of DNA synthesis. Spermidine, spermine, and histones from P. polycephalum and from calf thymus bind to poly(L-malate) and antagonize the inhibition. The polyanions poly(vinyl sulfate), poly(acrylate), poly(L-malate), poly(D,L-malate), poly(L-aspartate), poly(L-glutamate) have been examined for their potency to inhibit the DNA polymerase. The degree of inhibition is found to depend on the distance between neighboring charges, given by the number of atoms (N) interspaced between them. Poly(L-malate) (N = 5) and poly(D,L-malate) (N = 5) are the most efficient inhibitors, followed by poly(L-aspartate) (N = 6), poly(acrylate) (N = 3), poly(L-glutamate) (N = 8), poly(vinyl sulfate) (N = 3). It is proposed that poly(L-malate) interacts with DNA-polymerase-alpha-primase of P. polycephalum. According to its physical and biochemical properties, poly(L-malate) may alternatively function as a molecular chaperone in nucleosome assembly in the S phase and as both an inhibitor and a stock-piling agent of DNA-polymerase-alpha-primase in the G2 phase and M phase of the plasmodial cell cycle.
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PMID:Specific inhibition of Physarum polycephalum DNA-polymerase-alpha-primase by poly(L-malate) and related polyanions. 137 54

The Km and Vmax values for a set of primers: d(pT)n (pC) (pT)m (n = 3-9, m = 0-7) and d(pT)4 (pCpG)k (pT)4 (k = 1-5) have been estimated. Poly(dA) was used as a template. The number of complementary bases from the 3' end to a noncomplementary ones was shown to determine the efficiency of interaction of d(pT)n (pC) (pT)m with the Klenow fragment. Oligonucleotides d(pT)4 (pCpG)k (pT)4, in solution forming duplexes containing hairpin-like elements, show a higher affinity to the enzyme than control d(pT)4, d(pT)8 and d(pT)n (pC) (pT)m primers. For example, the Km value (1.1 nM) for d(pT)4 (pCpG)5 (pT)4 is about 14,000 and 200 times lower than those for d(pT)4 and d(pT)8, respectively. Possible reasons for such an abnormally high affinity of the above primers are discussed.
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PMID:The affinity of the Klenow fragment of E. coli DNA-polymerase 1 to primers containing bases noncomplementary to the template and hairpin-like elements. 154 84

Purpuromycin, an antibiotic active against both fungi and bacteria, shows different modes of action against these two kinds of micro-organisms; in Candida albicans it inhibits RNA synthesis, whereas in Bacillus subtilis protein synthesis is primarily affected, with DNA and RNA synthesis blocked at higher concentrations of the drug. In bacterial cell-free protein-synthesis systems, purpuromycin did not inhibit synthesis from endogenous mRNA (elongation of peptides initiated within the intact cell) but inhibited MS2-phase RNA-dependent protein synthesis (which requires initiation) by 50% at 0.1 mg/l. Poly(U)-directed polyphenylalanine synthesis was 50% inhibited by 20 mg of purpuromycin/l when added to a complete system; however, when purpuromycin was preincubated with ribosomes dissociated into 30 S and 50 S subunits, the concentration for 50% inhibition fell to 0.1 mg/l. By contrast, in a C. albicans cell-free system poly(U)-directed polyphenylalanine synthesis was partially inhibited only at 200 mg/l. Purpuromycin also inhibited polynucleotide synthesis in vitro in reactions using Escherichia coli or wheat-germ RNA polymerases or E. coli DNA polymerase I. We suggest that in bacteria the primary target of purpuromycin is on ribosomes and that its action precedes the elongation step of protein synthesis. The effect on nucleic acid synthesis in both fungi and bacteria may be due to interaction of purpuromycin with DNA.
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PMID:Mechanism of action of purpuromycin. 162 8

A crude in vitro system which initiates chloroplast DNA synthesis near the D-loop site mapped by electron microscopy [Wu, M., Lou, J. K., Chang, D. Y., Chang, C. H., & Nie, Z. Q. (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 6761-6765] consists of soluble proteins and proteins extracted from purified thylakoid membrane. In this paper, a DNA polymerase activity was purified to near homogeneity from the soluble protein fraction of this in vitro system by sequential chromatographic separations on heparin-agarose, DEAE-cellulose, and single-stranded DNA-agarose columns and sedimentation in a glycerol gradient. In the glycerol gradient, the enzyme activity sedimented at a position corresponding to a 110-kDa protein. Electrophoretic analysis of the highly purified fraction on SDS-polyacrylamide gel revealed a major polypeptide band with an apparent molecular mass of approximately 116 kDa. In situ DNA polymerase activity assay shows that the DNA polymerization function is associated with the 116-kDa band and an 80-kDa band which could be a subunit of the enzyme. Polymerization activity is inhibited by N-ethylmaleimide, ethidium bromide, and dideoxycytosine triphosphate and is relatively resistant to aphidicolin. Poly(dA).(dT)10 and gapped double-stranded DNA are preferred templates. The purified enzyme contains no exonuclease activity and can initiate DNA replication in a supercoiled plasmid DNA template containing the chloroplast DNA replication origin.
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PMID:Purification and characterization of a gamma-like DNA polymerase from Chlamydomonas reinhardtii. 198 80

The DNA and protein sequences of single-stranded DNA binding proteins (SSBs) encoded by the plP71a, plP231a, and R64 conjugative plasmids have been determined and compared to Escherichia coli SSB and the SSB encoded by F-plasmid. Although the amino acid sequences of all of these proteins are highly conserved within the NH2-terminal two-thirds of the protein, they diverge in the COOH-terminal third region. A number of amino acid residues which have previously been implicated as being either directly or indirectly involved in DNA binding are conserved in all of these SSBs. These residues include Trp-40, Trp-54, Trp-88, His-55, and Phe-60. On the basis of these sequence comparisons and DNA binding studies, a role for Tyr-70 in DNA binding is suggested for the first time. Although the COOH-terminal third of these proteins diverges more than their NH2-terminal regions, the COOH-terminal five amino acid residues of all five of these proteins are identical. In addition, all of these proteins share the characteristic property of having a protease resistant, NH2-terminal core and an acidic COOH-terminal region. Despite the high degree of sequence homology among the plasmid SSB proteins, the F-plasmid SSB appears unique in that it was the only SSB tested that neither bound well to poly(dA) nor was able to stimulate DNA polymerase III holoenzyme elongation rates. Poly [d(A-T)] melting studies suggest that at least three of the plasmid encoded SSBs are better helix-destabilizing proteins than is the E. coli SSB protein.
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PMID:Single-stranded DNA binding proteins (SSBs) from prokaryotic transmissible plasmids. 200 32


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