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)

With gene-43 (DNA polymerase)-ts-mutants of T4 phage, L98 (mutator) and CB121 (antimutator), and the T4 wild type, double labelling of DNA was carried out with (H3)-bromodeoxyuridine (BUdR) and (C14)-thymidine (TdR). Experiments on (C14) TdR for DNA synthesis measurement in the presence of BUdR offered evidence of the ability of the CB121 mutant to excise BUdR from the DNA. This effect took place only at increased temperature. As distinct from DNA synthesis of the host, all T4 phages used preferred TdR rather than BUdR for net synthesis.
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PMID:Excision of bromodeoxyuridine from T4-DNA by an antimutator polymerase of T4 phage. 23 71

Expression of the Epstein-Barr virus (EBV) DNA polymerase (EBVpol) open reading frame (BALF5) by in vitro transcription-translation yielded a 116-kDa primary translation product. Enzymatic DNA polymerase activity of the in vitro translated polypeptide required the presence of the 47-kDa BMRF1 (EA-D) gene product. Antiserum raised to the BALF5 gene product expressed in Escherichia coli specifically precipitated a 116-kDa polypeptide in extracts of latently infected lymphoblastoid cells induced for EBV replication. Immunofluorescence microscopy revealed colocalization of the EBVpol and EA-D(BMRF1) to discrete foci within the nuclei of induced cells; however, the blockade of viral DNA synthesis resulted in diffuse nuclear staining patterns for both antigens. Bromodeoxyuridine staining of these discrete foci colocalizing with EBVpol suggests that they are sites of early viral DNA synthesis. These observations suggest that EA-D(BMRF1) may be an accessory protein of the EBV DNA polymerase which colocalizes in vivo with EBVpol to sites of viral DNA replication and cooperates in vitro to form an active EBVpol holoenzyme.
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PMID:Cooperation of EBV DNA polymerase and EA-D(BMRF1) in vitro and colocalization in nuclei of infected cells. 165 95

The proliferative capacity of brain-tumor cells was analyzed in vitro and in situ using monoclonal antibody (MAb) against deoxyribonucleic acid (DNA) polymerase alpha. For the in vitro studies, two cultured human glioma cell lines were investigated using MAb against DNA polymerase alpha, the MAb Ki-67, a serum against proliferating cell nuclear antigen (PCNA/cyclin), bromodeoxyuridine (BUdR), and an anti-BUdR MAb. During exponential growth of the cells, the percentage of polymerase alpha-positive cells (the "polymerase alpha score") ranged from 72.0% to 77.1%, the Ki-67-positive cells (the "Ki-67 score") ranged from 43.4% to 59.4%, the PCNA/cyclin-positive cells from 30.9% to 41.4%, and the BUdR labeling index from 28.6% to 39.3%. For the in situ studies, tissue from 60 human brain tumors and from two normal human brains was investigated and the polymerase alpha scores and Ki-67 scores were compared. In normal brain tissue, no immunostaining was found by either method. In brain tumors, both the polymerase alpha scores and the Ki-67 scores correlated with the histological grade of malignancy. Polymerase alpha scores were generally higher than Ki-67 scores in the same specimen, especially in malignant brain tumors. These findings suggest that immunostaining of DNA polymerase alpha is a convenient and important new method by which to estimate the cellular proliferation rate of brain tumors. Polymerase alpha scores may be closer to the growth fraction of the individual tumor than the MAb Ki-67 or other scores.
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PMID:Immunohistochemical demonstration of DNA polymerase alpha in human brain-tumor cells. 196 2

Mutations which allow tolerance to 5-bromo-2'-deoxyuridine (BUdR) in a thymidine (TdR)-requiring strain of Bacillus subtilis have been examined. Differences in sensitivity to BUdR existed between isogenic strains harbouring the mutations. Those mutations originally isolated as BUdR-tolerant also bestowed tolerance to 5-bromouracil and vice versa. The strain exhibiting the greatest tolerance to BUdR maintained a normal rate of replication in the presence of BUdR whereas the parent strain did not, but the tolerant strain incorporated less analogue into DNA than the parent strain. The basis of the tolerance mutation appeared to lie at the point of uptake of the analogue into the cell as the tolerant mutant preferentially took up TdR over BUdR into whole cells. DNA polymerase activity measured in vitro did not distinguish between TdR and BUdR in either the parent or the mutant strain and although TdR kinase activity showed a preference for TdR over BUdR as a substrate, the extent of discrimination was similar in both strains.
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PMID:Tolerance to bromodeoxyuridine in a thymidine-requiring strain of Bacillus subtilis. 242 35

A tumor-derived factor that inhibits cellular DNA synthesis was identified. The factor was extractable from a small-cell lung carcinoma cell line grown in either chemically defined medium or nu/nu mice and inhibited tritiated thymidine ([3H]dThd) incorporation by tumor cell lines of autologous, allogeneic, and xenogeneic origins. The viability of nonproliferating cells from normal tissue was not affected. Tumor extract inhibitory activity was trypsin labile but was resistant to other proteases, neuraminidase, lipase, DNase, RNase, glucosidase, extremes of pH-temperature, and reducing conditions. Inhibitory activity was reversibly bound to helix pomatia lectin but not to lentil, wheat germ, or concanavalin A lectins. Purification by size-exclusion high-performance liquid chromatography yielded a bioactive unimodal 12-kilodalton (kd) peak. The bioactive 12-kd moiety could be eluted from sodium dodecyl sulfate-polyacrylamide gels. Redosing of populations of the T-lymphoblastoid cell line CEM achieved an early (24 hr) sustained depression of pulse [3H]dThd incorporation and ultimately led to decreased population density of factor-treated populations. DNA histogram analysis demonstrated no change in cell cycle phase distribution after factor treatment. 5-Bromo-2'-deoxyuridine (BrdUrd) vs. propidium iodide with the two-parameter Fluorescence-Activated Cell Sorter analysis showed relative inhibition of non-S-phase BrdUrd uptake at 24 hours. A cell-free DNA polymerase assay demonstrated significant inhibition of non-alpha-polymerase-associated DNA synthesis in factor-treated cells. These studies suggest that this tumor-derived inhibitor of DNA synthesis represents a class of cellular products involved in the autoregulation of growth by regulation of DNA synthetic activity.
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PMID:Inhibition of DNA synthesis by a small-cell lung carcinoma-derived protein. 302 Mar 1

5-Bromo-2'-deoxyuridine triphosphate (Br-dUTP) and dTTP are used interchangeably for DNA synthesis in vitro by the Klenow fragment of Escherichia coli DNA polymerase I. When DNA containing Br-dUMP instead of dTMP at a few preselected sites is transfected into competent bacteria, no mutation occurs, indicating that in vivo E. coli DNA polymerase always places a dAMP residue in front of any unrepaired Br-dUMP residue. On the other hand, in vitro Br-dUTP can also replace dCTP, but only with difficulty: when dCTP is absent, Br-dUMP can be forced in front of a dGMP residue, but the Klenow polymerase pauses before and after addition of Br-dUMP. Transfection into E. coli of the substituted DNA leads to the expected G----A transitions. These mutations can easily be targeted by using a suitable primer and the correctly chosen mix of deoxynucleoside triphosphates containing Br-dUTP. When Br-dUMP has been placed in front of a dGMP residue, the mutation yield is not 100%, showing a partial repair of the transfected DNA before it is replicated. Advantage can be taken of this partial repair to prepare a set of different mutations within a target region in a single experiment.
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PMID:Pairing properties of bromouracil and repair of bromouracil-containing DNA. Possible utilization of bromodeoxyuridine triphosphate for site-directed mutagenesis. 317 34

Comparison was made of the ability of calf thymus DNA polymerases alpha and beta to replicate the following templates: native E. coli CR-34 DNA (T-DNA), calf thymus DNA activated by DNase I (act.DNA), BU-DNA (from E. coli CR-34 cells cultured on BUdR-containing medium) with damages resulting from incomplete excision repair, as well as thermally denatured act.DNA and BU-DNA (s.s.act.DNA and s.s.BU-DNA). 3H-TTP incorporation during extensive replication of act.DNA was similar for both enzymes, being, as expected, 40 times higher than for T-DNA. Likewise, the differences in the yield of the s.s.act.DNA or s.s.BU-DNA replication between both enzymes were negligible. In contrast, damaged native DNA was 6 - 30 times more extensively replicated by DNA polymerase beta than alpha. We propose that this is due to the greater ability of DNA polymerase beta compared with alpha to replicate single-stranded gaps, the presence of which is more likely in damaged BU-DNA than in T-DNA and act.DNA.
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PMID:Evidence implying DNA polymerase beta function in excision repair. 625 46

A polA1 mutation in the DNA polymerase I gene of E. coli results in a drastic reduction of the frequency of mutagenesis induced by 5-bromo-2'-deoxyuridine (BUdR). Comparisons of the effect of a polA1 mutation on mutagenesis induced by methyl methane sulfonate (MMS), ultraviolet irradiation (UV) and 2-aminopurine (2-AP) demonstrated that a similar effect of a polA1 mutation is observed with MMS. This effect is much less marked with UV-and-2-AP-induced mutagenesis. It follows that DNA polymerase I plays a key role in the process of mutagenesis induced by BU and MMS. Bearing in mind that mutagenesis provoked by UV, MMS and BU involves participation of the accompanying induced error-prone system, the sources of the differences in requirement for DNA polymerase I are critically examined.
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PMID:Mutagenesis induced by 5-bromouracil and methyl methane sulfonate: role of DNA polymerase I. 637 42

Proliferative activity of 28 human brain tumors was estimated by simultaneous measurement of DNA polymerase alpha, Ki-67 and bromodeoxyuridine (BUdR) labeling indices on microscopic tissue preparations stained immunologically with monoclonal antibodies using a peroxidase technique. All the antigens were exclusively found in the nucleus. The labeling index of BUdR was lower than those of the other indicators. The values of the DNA polymerase alpha labeling index were almost the same as those of the Ki-67 labeling index. Simultaneous measurement of these parameters may provide more useful information on tumor cell growth kinetics than that of a single parameter.
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PMID:Measurement of labeling index of DNA polymerase alpha in human brain tumors. Comparative study with labeling indices of BUdR in vitro and Ki-67. 832 65

In order to evaluate at the ultrastructural level the three dimensional arrangement of the dispersed chromatin during the intephase, the immunogold detection of Bromodeoxyuridine (BrdU), of the DNA polymerase alpha and of the proliferating cell nuclear antigen (PCNA) was performed on human HL60 leukemia cells and nuclear matrices extracted from the same cellular model. The Field Emission In lens Scanning Electron Microscopy analysis of the ultrathin cryosectioned cells revealed the presence of a chromatin three dimensional network where the different constituents appeared repetitively assembled. Also the nuclear matrix showed a repetitive structure, on which the deprivation of the DNA corresponded to the selective loss of particular class sized fibers. The single or multiple combined immunolocalization of different structures involved in the DNA replication, where BrdU, DNA polymerase alpha and PCNA represent, respectively, the substratum, the polymerizing enzyme and a regulator of the reaction, allowed the understanding of its reciprocal spatial relationship on the dispersed interphasic chromatin and the role of the nuclear matrix in the replicative process.
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PMID:Chromatin fibers organization within the nucleus. 1173 1


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