Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.6 (RNA polymerase)
 34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Nuclei were isolated from bloodstream forms of Trypanosoma brucei by nitrogen cavitation and sedimentation through percoll density gradients. Transcription studies with these nuclei in vitro demonstrated features not seen with other eukaryotes: RNA synthesis was much greater in the presence of Mn2+ than with Mg2+ and was sensitive to high concentrations (10-100 micrograms/ml) of alpha-amanitin at all salt concentrations tested (25-300 mM ammonium sulphate). RNA polymerase extracted from nuclei by sonication at high ionic strength chromatographed as a single peak, sensitive to high alpha-amanitin concentrations, on DEAE-sephadex under conditions which resolved the classic three RNA polymerase forms when rat liver nuclear extracts were used.
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PMID:Unusual RNA polymerase content of Trypanosoma brucei nuclei. 241 50

Protein extracts from the protozoan ciliate Paramecium tetraurelia revealed high levels of RNA-dependent DNA polymerase activity (reverse transcriptase). Stable and constant during the somatic phase of the cell cycle, the reverse transcriptase activity quickly diminished following the completion of the sexual phases of the cell cycle: conjugation and autogamy. The Paramecium reverse transcriptase presented a number of common features with retroviral polymerases: ability to copy synthetic templates such as poly(rCm).oligo(dG) as well as mRNA; sensitivity to various reverse transcriptase inhibitors such as HPA 23, suramin, phosphonoformate and ethidium bromide; insensitivity to the action of other DNA and RNA polymerase inhibitors and, finally, the requirement for divalent cations before the enzyme can function: either magnesium or manganese. Although the reverse transcriptase activity was not proven to be independent from one of the DNA polymerases in paramecia, its high activity predicts a role in the paramecia cell cycle. From what we are able to conceive today two possible roles could be envisaged. Participation in the anlage macronucleus formation: micronuclear sequences are first transcripted and, after rearrangements of the RNA molecules, these are retrotranscribed into the macronuclear DNA molecules or association with retrotransposons that participate in the movement of certain macronuclear sequences into the germ-line micronucleus.
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PMID:RNA-dependent DNA polymerase activity in Paramecium tetraurelia: what for? 243 48

Incubation of purified wheat-germ RNA polymerase II with poly[d(A-T)] template, Mn2+, U-A dinucleoside monophosphate primer and UTP substrate resulted in catalytic formation of the trinucleoside diphosphate U-A-U, in accordance with the results of previous studies. Both Sarkosyl and heparin inhibited completely and immediately (within less than 1 min) U-A-U synthesis, if either of these compounds was added to the assays during the progress of the reaction. This behaviour is in marked contrast to that reported for single-step addition reactions catalysed by Escherichia coli RNA polymerase on the same template [Sylvester & Cashel (1980) Biochemistry 19, 1069-1074]. However, treatment of the transcription complexes with Sarkosyl or heparin for periods sufficient to abolish U-A-U formation completely did not suppress completely the ability of such complexes to elongate RNA chains. Hence, the effect of Sarkosyl or heparin on the rate of U-A-U synthesis was predominantly due to change in the rate (or in the mechanism) of trinucleotide product release by the transcription complexes. Furthermore, once U-A-U synthesis has begun on the poly[d(A-T)] template, the transcription complexes became resistant to the action of a competitor DNA such as poly[d(G-C)]. The results are consistent with a model where at least a sizeable fraction of the enzyme molecules remains associated with the DNA template upon formation of a single phosphodiester bond.
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PMID:Effect of Sarkosyl and heparin on single-step addition reactions catalysed by wheat-germ RNA polymerase II--poly[d(A-T)]transcription complexes. 247 3

Specific roles of manganese (Mn) and magnesium (Mg) on the activities of DNA-dependent RNA polymerases I and II isolated from rabbit bone marrow erythroid cell nuclei were investigated. Three main polymerases were separated from the cell nuclei. When RNA polymerase I and Mg were added to the RNA synthesis assay mixture containing erythroid cell DNA as template, RNA transcription activity was highest, but when Mg was replaced with Mn, denatured calf thymus DNA formed a better template than erythroid cell DNA. In contrast, nucleoplasmic DNA from erythroid cell and liver DNA were the best templates to stimulate RNA transcription when RNA polymerase II and Mn were added to the assay mixture. However, if Mn was replaced with Mg, RNA synthesis activity was drastically reduced when the template was nucleoplasmic DNA of erythroid cell. RNA polymerase I and Mg synthesized GC rich RNA, whereas RNA polymerase II and Mn synthesized AU rich RNA. Sedimentation analysis showed that the molecular weights of the RNA produced by polymerase I were larger when the enzyme was activated with Mg than with Mn, whereas those of the RNA produced by polymerase II were larger with Mn than with Mg. Furthermore, RNA produced by polymerase I and Mg using chromatin as a template hybridized better with nucleolar DNA than with nucleoplasmic DNA, whereas that produced by polymerase II and Mn hybridized better with nucleoplasmic DNA than with nucleolar DNA. These results suggest that RNA synthesis is dependent on the activity of specific RNA polymerases and the presence of specific divalent cations and templates, and that the cofactor and template for RNA polymerase I are, respectively, Mg and the nucleolar DNA of cell nuclei, whereas those for RNA polymerase II are Mn and nucleoplasmic DNA.
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PMID:Specific role of manganese and magnesium on RNA synthesis in rabbit bone marrow erythroid cell nuclei. 248 49

To analyze transcription in Typanosoma brucei, we have characterized the trypanosomal RNA polymerases. Here we present our results, which allow a discrimination between the different classes of RNA polymerases in nuclear run-on experiments by polymerase inhibitors and Mn2+ dependence. We also describe the separation of trypanosomal RNA polymerases by chromatography, demonstrating that T. brucei contains RNA polymerases I-III. The outcome of our experiments suggests that the VSG genes of T. brucei are not transcribed by RNA polymerase I, as previously reported, but by RNA polymerase II. We propose that an additional factor modifies RNA polymerase II, resulting in the alpha-amanitin-resistant transcription of VSG genes. Our data also suggest that the mini-exon genes, which encode the 5' end of each trypanosomal mRNA, are probably transcribed by RNA polymerase III.
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PMID:Characterization of the RNA polymerases of Trypanosoma brucei: trypanosomal mRNAs are composed of transcripts derived from both RNA polymerase II and III. 258 3

RNA-polymerases of mollicutes differ considerably from certain species of bacteria in the temperature optimum of the activity manifestation. The activity of mollicute enzymes is considerably higher in the presence of manganese ions than in the presence of magnesium ions. They differ from Escherichia coli transcriptase in this character but are similar to the RNA-polymerase of lactic bacteria, hypothetic ancestors of acholeplasm. Sensitivity of RNA-polymerases to metals may be one of arguments explaining phylogeny of mycoplasms. It is established that for studying mollicute transcriptase the reacting mixture for examining the enzymic activity besides the major components should have the following parameters: pH 8.0; the MnCl concentration--8-10 mM; for form II of the A. laidlawii subsp. granulum 118 enzyme--4.5 mM; ammonium sulphate concentration--40 mM; for form II of st. 118-20 mM; the reaction should be conducted at the temperature of 27 degrees C for 30 min.
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PMID:[Optimization of physico-chemical parameters of reaction media for determining the activity of RNA-polymerases of mollicutes]. 266 9

We have purified from nuclear extracts of Drosophila Kc cells a 36-kDa protein, DmS-II, which has an effect on the elongation properties of RNA polymerase II. DmS-II stimulates RNA polymerase II during the transcription of double-stranded DNA templates when the nonphysiological divalent cation manganese is present. In the presence of physiological mono- and divalent cations, the factor reduces the tendency of RNA polymerase II to pause at specific sites along a dC-tailed template including the major pause encountered after 14 nucleotides have been incorporated. Based on its size and chromatographic properties, as well as its ability to stimulate RNA polymerase II activity in the presence of manganese, the protein seems to be analogous to a factor S-II purified from mouse cells (Sekimizu, K., Kobayashi, N., Mizuno, D., and Natori, S. (1976) Biochemistry 15, 5064-5070). We have used a completely defined system and show that the properties of DmS-II are intrinsic to the factor and not mediated through other factors.
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PMID:Properties of a Drosophila RNA polymerase II elongation factor. 272 10

The putative structural gene encoding the vaccinia virus type I DNA topoisomerase (EC 5.99.1.2) was expressed in Escherichia coli under the control of a bacteriophage T7 promoter. Provision of T7 RNA polymerase resulted in the accumulation to high level of a Mr = 33,000 type I topoisomerase with the properties of the vaccinia enzyme. A simple purification scheme yielded approximately 8 mg of recombinant vaccinia topoisomerase from 400 ml of bacteria. DNA unwinding by the enzyme was stimulated by magnesium, manganese, calcium, cobalt, and spermidine, but inhibited by copper and zinc. Like eukaryotic cellular type I topoisomerases, but unlike the prokaryotic counterpart, the recombinant topoisomerase relaxed positively and negatively supercoiled DNA. The viral topoisomerase I was, however, resistant to the effects of camptothecin, a drug that specifically inhibits cellular type I topoisomerases.
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PMID:Characterization of vaccinia virus DNA topoisomerase I expressed in Escherichia coli. 284 43

An RNA polymerase activity capable of initiating transcription at both the heavy strand rRNA promoter and the light strand promoter of human mitochondrial DNA has been partially purified from HeLa cell mitochondria and characterized in its requirements and products. The ratio of the two transcription initiating activities varied considerably from preparation to preparation. The human mtRNA polymerase partially purified by DEAE-cellulose and heparin-agarose chromatography exhibits a great sensitivity to ionic strength and to Mn2+, characteristics which clearly differentiate this enzyme from bacterial and eukaryotic nuclear RNA polymerases, and in contrast resemble the behavior of the yeast mtRNA polymerase. The human mtRNA polymerase exhibits a requirement for ATP which is 15- to 20-fold higher than that for the other NTPs, a low optimum template DNA concentration, and a marked susceptibility to inhibition by non-mitochondrial DNA.
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PMID:Characterization of an RNA polymerase activity from HeLa cell mitochondria, which initiates transcription at the heavy strand rRNA promoter and the light strand promoter in human mitochondrial DNA. 298 80

The kinetics of interaction of PPi and its diphosphonic analog, methylenediphosphonic acid (MDPA), with nucleoside triphosphates, DNA and Mg2+ binding sites of DNA-dependent RNA polymerase II from calf thymus was investigated. The values of apparent Km in the NTP polymerization reaction for ATP and CTP equal to 2.7 X 10(-4) and 1.8 X 10(-4) M, respectively, were determined. It was shown that MDPA and PPi competitively inhibited the RNA polymerase reaction with respect to nucleoside triphosphate. The inhibition constants (Ki) of ATP and CTP incorporation for MDPA were 2.2 X 10(-4) and 3.3 X 10(-4) M, respectively, while those of the nucleoside triphosphate incorporation for PPi were equal to 1.4 X 10(-4) and 2.0 X 10(-4) M, respectively. MDPA and PPi were incompetitive inhibitors of template (DNA) and Mn2+. A possible mechanism of inhibition of the RNA polymerase reaction by MDPA is proposed.
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PMID:[Kinetics of the interaction of methylene diphosphonic acid and inorganic pyrophosphate with DNA-dependent RNA-polymerase from calf thymus]. 298 49


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