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Deoxyribonucleic acid-dependent ribonucleic acid polymerases from normal and polyoma-transformed BHK-21/C13 cells. 下载免费PDF全文
DNA-dependent RNA polymerase (EC 2.7.7.6) ACTIVITIES FROM NORMAL BHK-21/C13 cells and from BHK-21/C13 cells transformed by polyoma virus (PYY cells) were solubilized and fractionated on columns of DEAE-Sephadex. Various properties of the A and B enzymes from the two types of cell were compared. 1. The yields of polymerase relative to the DNA content of the nuclear preparations are similar for both cell types. 2. The ionic-strength optima of polymerases A and B are 12.5 mM and 100mM with respect to (NH4)2SO4 for both cell types. 3. The Mn2+/Mg2+ activity ratio (measured at the respective optimum for each cation) for polymerase A from BHK-21/C13 cells was 1.48 and for the polymerase A from PYY cells was 0.55. The corresponding ratios for polymerase B were 10.11 for BHK-21/C13 cells and 22.75 for PYY cells. 4. Minor differences in the ability of the A polymerases to transcribe native and denatured DNA templates were observed; such differences were not apparent when the B polymerases were compared. 5. All the polymerases were inhibited completely by actinomycin D and by rifampicin AF/013, but not markedly so by rifampicin. Alpha-amanitin inhibited polymerase B but not polymerase A. 相似文献
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Deoxyribonucleic acid dependent ribonucleic acid polymerases from two T4 phage-infected systems 总被引:17,自引:0,他引:17
A Stevens 《Biochemistry》1974,13(3):493-503
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Deoxyribonucleic acid-dependent ribonucleic acid polymerase activity in rat liver after protein restriction. 下载免费PDF全文
Rats were fed for 6 days on a diet containing either 3 or 20% high-quality protein. Nuclei were isolated from liver and DNA-dependent RNA polymerases (EC 2.7.7.6) extracted with 1 M-(NH4)2SO4. The proteins were then precipitated with 3.5 M-(NH4)2SO4 and after dialysis applied to a DEAE-Sephadex column. The column was developed with a gradient of (NH4)2SO4. Polymerase I separated well from alpha-amanitin-sensitive polymerase II. The enzyme activities were compared between the two dietary groups. Rats that had received 3% protein showed a lower polymerase I activity per g wet wt. of liver, per mg of DNA and per mg of protein. Polymerase II was lower in activity per g wet wt. of liver and per mg of DNA, but was higher per mg of protein. Polyacrylamide-gel electrophoretograms showed a higher proportion of contaminating proteins in polymerase II fractions isolated from 20%-protein-fed rats. The data explain the lower activity obtained per mg of protein in these rats. It is concluded that a decrease in dietary protein content from 20 to 3% induces a fall in content and specific activity of RNA polymerase I and II in liver. 相似文献
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Effect of an exotoxin from Bacillus thuringiensis on Deoxyribonucleic acid-dependent ribonucleic acid polymerase in nuclei from adult Sarcophaga bullata. Unusual behaviour of eukaryotic polymerases to inhibitors 下载免费PDF全文
The DNA-dependent RNA polymerase activities in nuclei isolated from adult Sarcophaga bullata are unusual in their responses to metal ions, ionic strength and inhibitors. There is an activity that is sensitive both to rifamycin and to alpha-amanitin. The activity is less sensitive to Bacillus thuringiensis exotoxin than is larval polymerase, and low concentration of exotoxin provoke a slight stimulation. 相似文献
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Synthesis of different species of nucleolar ribonucleic acid 总被引:1,自引:0,他引:1
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Deoxyribonucleic acid-dependent ribonucleic acid polymerase activity in purified trachoma elementary bodies: effect of sodium chloride on ribonucleic acid transcription 总被引:17,自引:7,他引:10 下载免费PDF全文
Highly purified trachoma elementary bodies (T'ang strain), incubated in the presence of the four nucleoside triphosphates [Mg(2+), Mn(2+), 2-mercaptoethanol, tris(hydroxymethyl)aminomethane buffer (pH 7.5)] were found to incorporate (3)H-uridine triphosphate (UTP) into ribonucleic acid (RNA) molecules. Eighty-seven per cent of the labeled molecules were sensitive to ribonuclease treatment. In vitro RNA synthesis was almost completely inhibited by actinomycin D. Rifampin was also inhibitory, but allowed some initial RNA synthesis before complete inhibition occurred. When the reaction mixture lacked Mn(2+), trachoma elementary bodies synthesized, for a limited period, high-molecular-weight RNA species (23 to 24S, 16 to 17S, and 10 to 11S). Addition of 0.2 m NaCl to the same reaction mixture stimulated and prolonged (3)H-UTP incorporation into the same radioactive RNA species. Addition of 0.001 m Mn(2+) instead of NaCl also stimulated (3)H-UTP incorporation but prevented the synthesis of the high-molecular-weight RNA species. 相似文献
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Purification and properties of the ribonucleic acid-dependent ribonucleic acid polymerase from Halobacterium cutirubrum 总被引:4,自引:1,他引:3
1. The RNA-dependent RNA polymerase from Halobacterium cutirubrum was purified to electrophoretic homogeneity. 2. It requires a single-stranded molecule of RNA or polyribonucleotide as template. 3. Nearest-neighbour analyses of the products formed on random poly(A,U) or alternating poly(A-U) templates and base analysis of the product of synthesis directed by wheat-germ RNA prove that the template is copied accurately. 4. The enzyme initiates new chains with purine ribonucleoside triphosphates. 5. Sucrose-density-gradient analysis of the product indicates that it has a size distribution similar to that of the template. 6. Preliminary amino acid analysis of the RNA-dependent polymerase shows that it contains much less serine than either of the subunits of H. cutirubrum DNA-dependent RNA polymerase. 7. The RNA-dependent enzyme is unable to substitute for either subunit of the DNA-dependent polymerase, and both the latter are devoid of RNA-dependent activity. 相似文献
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Robert F. Weaver 《Archives of biochemistry and biophysics》1976,172(2):470-475
RNA Polymerases I and II have been purified to homogeneity from the slime mold Physarum polycephalum. When subjected to ultracentrifugation in separate sucrose gradients, or together, the two enzymes migrate with S values estimated at 14.5 S. Polyacrylamide gel electrophoresis under denaturing conditions reveals that the two enzymes have distinct but similar subunit structures. The probable subunit structure of polymerase II is as follows: 175,0001; 140,0001; 24,0002; 17,0001. 相似文献
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Ribonucleic acid (RNA)-dependent RNA polymerase activity was demonstrated in the microsomal and ribosomal fraction from the spleen cells of immunized mice. The enzyme activity was solubilized by Triton X-100 from the fraction and partially purified by Biogel A 1.5 m column chromatography. The RNA-dependent RNA polymerase activity was eluted in a single peak from the column. High activity was demonstrated with an RNA polymerase activity was eluted in a single peak from the column. High activity was demonstrated with an RAN preparation (iotaRNA) as template made from the spleens of immunized mice but very low activity was found with an RNA preparation made from the spleens of normal mice. Incorporation of 3H-UTP markedly decreased in the presence of RNase but not in the presence of DNase. DNA preparations made from the spleens of immunized mice were inactive as template for this enzyme. The iotaRNA preparation was fractionated by sucrose density gradient centrifugation. A fraction corresponding to 12-13 S was most active as a template. It was followed by a fraction corresponding to 6-7 S. Sucrose gradient analysis of the 3H-UTP-labeled product was attempted. Some properties of this enzyme are described. 相似文献
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Rat liver nuclei were fractionated into chromatin and nucleolar fractions. Chromatin DNA, which does not form hybrids with rRNA, was, nevertheless, able to hybridize with 32P-labelled total nucleolar RNA. The optimal temperature for this hybridization was 55 degrees C when the reaction was carried out in 2 X SSC (0.3 MnaCl + 0.3 M-sodium citrate). The hybrids formed were specific, as judged by analysis of thermal elution profiles. The low Tm (73 degreesC) observed could be explained by the low amount of DNA in the filters. The lenth of the hybridized sequences was extimated as 54 mucleotide pairs. Contamination to nucleolar RNA by nucleoplasmic RNA was ruled out by showing the former was able to form more hybrids than the latter. Competition experiments showed that hybridization of nucleolar RNA, although not competed with by rRNA, suffered pronounced competition from total microsomal RNA, even though the levels of competition obtained did not equal thsoe with cold nucleolar RNA as competitor. 相似文献
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G Quagliarotti E Hidvegi J Wikman H Busch 《The Journal of biological chemistry》1970,245(8):1962-1969
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Nucleic acid enzymology of extremely halophilic bacteria. Halobacterium cutirubrum ribonucleic acid-dependent ribonucleic acid polymerase 总被引:8,自引:6,他引:2
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. 相似文献