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)

A procedure was developed for isolating nuclei from either the conidial or germinated conidial growth phase of Neurospora crassa. A frozen conidial suspension was lysed by passage through a French pressure cell, and the nuclei were freed from the broken cells by repeated homogenization in an Omni-Mixer. Pure nuclei were obtained from the crude nuclear fraction by density banding in a Ludox gradient. The final nuclear yield was 20 to 30%. The nuclei had a deoxyribonucleic acid (DNA):ribonucleic acid (RNA):protein ratio of 1:3.5:7 and were active in RNA synthesis. The nuclei, stained with the DNA stain 4,6-diamidino-2-phenylindole, appeared under fluorescence microscopy as bright blue spheres, 1 micron in diameter, essentially free from cytoplasmic attachments. Chromatin extracted from the nuclei in a 70 to 75% yield by dissociation with 2 M sodium chloride and 5 M urea had a DNA:RNA:protein ratio of 1:1.05:1.7. Chromatin reconstituted from this preparation exhibited a level of RNA polymerase template activity lower than that of pure Neurospora DNA, but the maximum level of reconstitution obtained was only 10%. Fractionation of Neurospora chromatin on hydroxylapatite separated the histones from the chromatin acidic proteins. The normal complement of histone proteins was present in both the reconstituted and dissociated chromatin preparations. The acidic protein fraction exhibited a variety of bands on sodium dodecyl sulfate gel electrophoresis ranging in molecular weight from 15,000 to 70,000. The gel pattern was much more complex for total dissociated chromatin than for reconstituted chromatin.
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PMID:Isolation and characterization of nuclei from Neurospora crassa. 16 36

1. The interaction of aflatoxin B(1) with different polynucleotides was studied spectrophotometrically. Equations were derived that enable the degree of binding to be determined without first determining the extinction coefficient of the bound form. 2. The interaction with calf thymus DNA obeys first-order relationships with an association constant of 0.40mm(-1), but there is some evidence for a secondary binding process from results obtained at 390nm. 3. The spectral shifts decreased in the order polyadenylic acid+polyuridylic acid>DNA>polyadenylic acid>polyadenylic acid+polyinosinic acid. Polycytidylic acid, polyuridylic acid, polyinosinic acid (both single- and triple-stranded), AMP, CMP, GMP and UMP did not interact with aflatoxin. It was concluded that there is a requirement for the amino group of adenine (or possibly guanine) for binding of aflatoxin to polynucleotides to occur. 4. Binding is reversed by increasing ionic strength, and by Mn(2+) and Mg(2+) in the concentration range studied (0-5mm). The effect of the Mn(2+) or Mg(2+) was far greater than would be expected on the basis of their ionic strength. With both the bivalent cations and sodium chloride the reversal is greatest with double-stranded polynucleotides. 5. Inhibition in vitro of the DNA-dependent RNA polymerase of Escherichia coli by aflatoxin B(1) was detected only in the absence of Mg(2+) and at concentrations of Mn(2+) below the optimum for RNA synthesis in vitro. 6. The degree of inhibition (maximally 30%) was dependent on the concentration of Mn(2+) and decreased during incubation.
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PMID:The interaction of aflatoxin B1 with polynucleotides and its effect on ribonucleic acid polymerase. 489 17

1. Crude extracts of the extreme halophile Halobacterium cutirubrum contain separable DNA-dependent and RNA-dependent RNA polymerases. 2. The RNA-dependent enzyme has been purified about 2800-fold. 3. It requires RNA, preferably of high molecular weight, and all four ribonucleoside triphosphates to incorporate (14)C-labelled nucleoside triphosphate into an acid-insoluble, ribonuclease-sensitive product. 4. Both the stability and activity of the RNA polymerase are relatively insensitive to changes in potassium chloride or sodium chloride concentration, but incorporation is stimulated by both Mg(2+) and Mn(2+). 5. The molecular weight of the enzyme is about 17000-18000.
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PMID:Nucleic acid enzymology of extremely halophilic bacteria. Halobacterium cutirubrum ribonucleic acid-dependent ribonucleic acid polymerase. 511 74

1. Conditions have been established for the estimation of molecular weights of proteins by analytical gel filtration and sucrose-density-gradient centrifugation in 2.5m-potassium chloride-1m-sodium chloride; Halobacterium cutirubrum polynucleotide phosphorylase, DNA-dependent RNA polymerase and RNA-dependent RNA polymerase have been studied by these methods. 2. The RNA-dependent polymerase has also been studied by density-gradient centrifugation in the absence of salt. 3. All three proteins are of unusually low molecular weight compared with similar enzymes from non-halophilic bacteria.
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PMID:Nucleic acid enzymology of extremely halophilic bacteria. Gel-filtration and density-gradient-centrifugation studies of the molecular weights of Halobacterium cutirubrum polynucleotide phosphorylase and deoxyribonucleic acid- and ribonucleic acid-dependent ribonucleic acid polymerases. 511 75

The influenza virus A/duck/Alberta/48/76 with the antigen formula H7N3 (16) and Hav1 Nav2 (WHO nomenclature from 1971) (15), respectively, as well as a nonpathogenic virus of the subtype Hav1 were purified to a high degree by ultracentrifugation in continuous sucrose gradients (15-40% w/w and 20-60% w/w, respectively). The activity of the RNA polymerase of this virus preparation was determined by incorporating 3H-UMP in acid insoluble material following preincubation of the virus with the nonionic detergens Nonidet P-40 for 15 min at 32 degrees C. The influence of different concentrations was investigated of dinucleotid, NaCl, MgCl2, Nonidet P-40 and different incubation temperatures. Optimal incorporation rates were found at following conditions: 0.2 mM dinucleotid ApG, 150 mM sodium chloride and 8 mM magnesium chloride by concentration of ions, 0.25-0.5% detergens Nonidet P-40 as well as a temperature of incubation of 32 degrees C. The data for optimal polymerase activity for the avian influenza virus A/duck/Alberta/48/76 are generally not different from the conditions described for the Fowl-Plague-Virus and for human strains.
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PMID:[Characterization of RNA polymerase activity of highly purified preparations of influenza virus A/duck/Alberta/48/76]. 637 62

Hantaan virus, the prototype virus of hemorrhagic fever with renal syndrome, was examined for nucleic acid characteristics which would support its previously proposed inclusion in the virus family Bunyaviridae. Nucleocapsid RNA from Hantaan virions and a control bunyavirus were examined for ribonuclease A (RNase A) sensitivity. Both viruses exhibited a similar accessibility of RNA within nucleocapsids to digestion by RNase A. Complete digestion of the RNA of both viruses was affected with high concentrations of ribonuclease. Evidence for negative strand RNA polarity was obtained by an in vitro transcriptase assay. RNA dependent RNA polymerase activity was associated with Hantaan virions. Polymerase activity required manganese and nucleoside triphosphates and was enhanced by magnesium, 2-mercaptoethanol, and sodium chloride. Oligonucleotide map analysis of the large (L), medium (M), and small (S) genome segments of Hantaan virus demonstrated that each RNA species was unique with respect to each other and was different from host cell ribosomal RNA. A common 3' terminal sequence of the three genome segments was determined to be 3' AUCAUCAUCUG. This sequence is different from those reported for viruses within the four recognized genera of the Bunyaviridae. Because all other data were consistent with nucleic acid characteristics of the Bunyaviridae, we propose a separate genus within the Bunyaviridae with Hantaan as its prototype virs.
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PMID:Analysis of Hantaan virus RNA: evidence for a new genus of bunyaviridae. 641 60

Hepatitis A virus (HAV) strains found in selected South African (SA) surface waters were characterised to establish what HAV types are circulating in the environment, thus reflecting circulation in the surrounding communities. Surface water samples used for irrigation or domestic purposes, and water samples from the outflow of wastewater plants were collected from six provinces. Viruses were recovered from the samples using a glass wool adsorption-elution method and then further concentrated using polyethylene glycol/sodium chloride precipitation. After automated nucleic acid extraction, samples were analysed for HAV by real-time reverse-transcriptase polymerase chain reaction. HAV strains were genotyped by nucleotide sequence analysis of the capsid gene VP1 and the VP1/P2B junction. HAVs were detected in 76% (16/21) of the surface water samples and in 37% (19/51) of the samples from the wastewater plants. Strains were characterised from 32 of the 35 samples and classified within genotype IB. The presence of genotype IB in the water sources confirms human faecal contamination. Hence, these faecally-contaminated water sources may be a potential transmission route of HAV infection and a potential source of contamination of irrigated fresh produce in SA.
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PMID:Molecular characterisation of hepatitis A virus strains from water sources in South Africa. 2462 38