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:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The synthesis of ribosomal precursor RNA in Novikoff hepatoma (N1S1) cells is very sensitive to cordycepin (3'-dA). The synthesis of hnRNA, however, is resistant to inhibition concentrations of 3'-dA that completely block the synthesis of 45S ribosomal RNA precursor. We have examined the RNA polymerases present in these cultured cells with regard to their sensitivity to cordycepin 5'-triphosphate (3'-dATP) in an effort to explain the differential inhibition of RNA synthesis observed in vivo. RNA polymerases I and II were characterized on the basis of their chromatographic behavior on DEAE-Sephadex, as well as the response of their enzymatic activities to ionic strength, the divalent metal ions Mn2+ and Mg2+, and the toxin alpha-amanitin. For both enzymes the inhibition of in vitro RNA synthesis by 3'-dATP was competitive for ATP. The km values for ATP and the K1 values for 3'-dATP for the two enzymes were quite similar. RNA polymerase II, the enzyme presumed responsible for hnRNA synthesis, was actually slightly more sensitive to 3'-dATP than RNA polymerase I, the enzyme presumed responsible for ribosomal precursor RNA synthesis. Similar data were obtained when the RNA polymerases were assayed in isolated nuclei. These results indicate that the differential inhibition of RNA synthesis caused by 3'-dA in vivo cannot be simply explained by differential sensitivity of RNA polymerases I and II to 3'-dATP.
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PMID:The sensitivity of RNA polymerases I and II from Novikoff hepatoma (N1S1) cells to 3'-deoxyadenosine 5'-triphosphate. 17 30

The interaction of Mn2+, substrates and initiators with RNA polymerase have been studied by kinetic and magnetic resonance methods. As determined by electron paramagnetic resonance, Mn2+ binds to RNA polymerase at one tight binding site with a dissociation constant less than 10 muM and at 6 +/- 1 weak binding sites with dissociation constants 100-fold greater. The binding of Mn2+ to RNA polymerase at both types of sites causes an order of magnitude enhancement of the paramagnetic effect of Mn2+ on the longitudinal relaxation rate of water protons, indicating the presence of residual water ligands on the enzyme-bound Mn2+. A kinetic analysis of the Mn2+-activated enzyme with poly(dT) as template indicates the substrate to be MnATP under steady-state conditions in the presence or absence of the initiator ApA. ATP and UTP interact with the tightly bound Mn2+ to form ternary complexes with approximately 50% greater enhancement factors. The dissociation constant of MnATP from the tight Mn2+ site as determined by longitudinal proton relaxation rate (PRR) titration (4.7 muM) is similar to the KM of MnATP in the ApA-initiated RNA polymerase reaction (10 +/- 3 muM) but not in the ATP-initiated reaction (160 +/- 30 muM). Similarly, the dissociation constant of the substrate MnUTP from the tight Mn2+ site (90 muM) is in agreement with the KM of MnUTP (101 +/- 13 muM) when poly[d(A-T)]-poly[d(A-T)] is used as template, indicating the tight Mn2+ site to be the catalytic site for RNA chain elongation. Manganese adenylyl imidodiphosphate (MnAMP-PNP) has been found to be a substrate for RNA polymerase. It has the same affinity as MnATP for the tight site but, unlike the results obtained with MnATP, the enhancement is decreased by 43% in the enzyme Mn-AMP-PNP complex. These results suggest that the enzyme-bound Mn2+ interacts with the leaving pyrophosphate group. The initiators ApA and ApU and the inhibitor rifamycin interact with the enzyme-Mn2+ complex producing small (15-20%) decreases in the enhancement. The dissociation constant of ApA estimated from PRR data (less than or equal to 1.5 muM) agrees with that determined kinetically (1.0 +/- 0.5 muM) as the concentration of ApA required to produce half-maximal change in the KM of MnATP. In the presence of the initiation specific reagents ApA, ApU, or rifamycin, the affinity of the enzyme-Mn complex for ATP or UTP shows little change. However, ATP and UTP no longer increase the enhancement factor of the tightly bound Mn2+ but decrease it by 30-55%, indicating a change in the environment of the Mn2+-substrate complex on the enzyme when the initiation site is either occupied or blocked. Although the role of the six weak Mn2+ binding sites is not clear, the presence of a single tightly bound Mn2+ at the catalytic site for chain elongation which interacts with the substrate reinforces the number of active sites as one per molecule of holoenzyme and provides a paramagnetic reference point for further structural studies.
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PMID:Magnetic resonance and kinetic studies of the role of the divalent cation activator of RNA polymerase from Escherichia coli. 18 95

A template-dependent RNA polymerase has been isolated from poliovirus-infected cells by assaying for the ability of the enzyme to copy poly(A) complexed to an oligo(U) primer. The polymerase was solubilized with detergent, and RNA was removed by precipitation with 2 M LiCl. The solubilized polymerase required both poly(A) and oligo(U) for activity and was stimulated by Mg2+ but was inhibited by Mn2+. Poly(A)-oligo(U)-dependent poly(U) polymerase was not found in extracts of HeLa cells until about 2 hr after poliovirus infection, and then there was a linear increase in activity until about 5 hr. Analysis of the polymerase by glycerol gradient centrifugation showed that the majority of the activity sedimented at about 4 S, indicating that it was no longer complexed with high-molecular-weight RNA or cellular membranes. This poly(A)-oligo(U)-dependent polymerase activity could represent an important component of the poliovirus RNA-dependent RNA polymerase.
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PMID:Poliovirus-specific primer-dependent RNA polymerase able to copy poly(A). 19 96

Noradrenaline added to perfused rabbit heart previously perfused with labelled precursors causes, after 2.5 and 5.0 min, a general increase of specific radioactivity or RNA in subcellular fractions, but no augmentation of acetylation of F2a2 and F2a1 histone fractions and no stimulation of DNA-dependent RNA polymerase activities. Synthesis of spermidine and spermine is enhanced at 10.0 min of treatment, when there is also a fall in specific radioactivity of RNA. The cytoplasmic Mn2+-stimulated polyadenylate polymerase activity is strongly enhanced 30s to 2.5 min after injection of noradrenaline or of dibutyryl cyclic AMP. Both the cyclic nucleotide and noradrenaline have no influence in vitro on the polyadenylate polymerase reaction.
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PMID:Modifications of major aspects of myocardial ribonucleic acid metabolism as a response to noradrenaline. Behaviour of polyadenylate polymerase and ribonucleic acid polymerase, acetylation of histones and rate of synthesis of polyamines. 20 85

Two reactions of bacteriophage-Qbeta RNA polymerase with synthetic templates were characterized and used to study the effects of substrate, metal and template on inhibition by Pi and PPi. Analysis of the poly(C)-dependent reaction yielded results on kinetics, GTP-dependence, preference for Mn2+ over Mg2+, and Michaelis constants for template similar to those in the literature. New data are provided for the poly(U2,C)-dependent reaction. Our results suggest that GTP and Mn2+ can form relatively stable complexes with the polymerase and that such complexes change the interaction of the enzyme with the inhibitors, Pi and PPi.
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PMID:Substrate, metal and template effects on inhibition of bacteriophage-qbeta ribonucleic acid polymerase by ortho- and pyro-phosphate. 20 14

Examinations were made on substances that enhance or inhibit the induction of hepatoma in rats previously fed 3'-methyl-4-(dimethylamino)azobenzene (3'-Me-DAB) for a brief period. The substances tested were stilbene, 4-nitrostilbene, 4,4'-dihydroxystilbene, diethylstilbestrol, 17beta-estradiol, and methyltestosterone. Male Donryu rats were fed 0.5 g of 3'-Me-DAB by being maintained on a diet containing 0.06% 3'-Me-DAB, and then they were fed 0.25 or 0.5 g of a test substance with the basal diet. Comparison of the development and yield of hepatomas indicated that 4-nitrostilbene and methyltestosterone had an activity of enhancing 3'-Me-DAB carcinogenesis, whereas diethylstilbestrol and 17beta-estradiol had an activity to retard it. Other substances showed no such activities. The enhancement by 4-nitrostilbene and inhibition by diethylstilbestrol of 3'-Me-DAB carcinogenesis was correlated with their effect on liver nucleic acid metabolism. Feeding of 4-nitrostilbene caused a selective inhibition of Mn2+-(NH4)2SO4-activated RNA polymerase activity of liver nuclei and reduced liver RNA content. The deleterious alteration of liver RNA metabolism was followed by the enhancement in the incorporation of ip-injected 3H-thymidine into DNA of liver nuclei. On the other hand, feeding of diethylstilbestrol increased tissue RNA content without effect on RNA polymerase activity of liver nuclei, and had an activity of increasing the incorporation of 3H-thymidine into DNA. The possible implication of these results with regard to the enhancement and inhibition of hepatocarcinogenesis is discussed.
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PMID:Enhancing and inhibitory effects of some stilbene and steroid compounds on induction of hepatoma in rats fed 3'-methyl-4-(dimethylamino)azobenzene. 20 6

Isolated nuclei from the liver of rats maintained under daily scheduled conditions of light (12 hrs) and food (8 hrs) showed a peak of Mn2+-dependent poly (A) polymerase activity 6 hrs after the beginning of the feeding period, as shown by DNA dependent RNA polymerase B. In fasting animals the peak of poly (A) polymerase occurs at 18.00.
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PMID:[Circadian rhythm of poly A polymerase in rat liver nuclei]. 23 1

Mn2+-dependent poly(A) polymerase activity tested in isolated rat liver nuclei is unchanged both in the absence and presence of exogenous poly(A) 30 min. after administration of a dose of alpha-amanitin that inhibits DNA-dependent RNA polymerase B. Longer times of treatment cause poly(A) polymerase to drop to 50% in the absence of poly(A) and to increase almost twice in its presence.
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PMID:[Effect of administration of alpha-amanitine on Mn2+ - dependent poly-A-polymerase in rat liver nuclei]. 23 4

A high molecular weight membrane-bound DNA polymerase from the mouse myeloma, MOPC-104E, has been purified extensively, and characterized with regard to physical and reaction properties. This enzyme, which is readily distinguishable from other myeloma enzymes that are analogous to the recognized forms of cellular DNA polymerase, is ddesignated DNA polymerase III. DNA polymerase III activity in whole homogenates from MOPC-104E was solubilized and then prurifed using a series of ion-exchange chromatographic procedures followed by DNA-cellulose chromatography and glycerol gradient centrifugation; the enzyme activity as measured with poly(rA)-(dT)12-18 as template-primer and Mn2+ as divalent cation, was purified as much as 18,000-fold. In the final stages of the pruification, DNA polymerase III possessed no detectable RNA polymerase activity, nucleoside diphosphokinase activity, or nucease activity toward DNA or single- and double-stranded RNA...
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PMID:On the DNA polymerase III of mouse myeloma: partial purification and characterization. 23 42

Exposure of sulfhydryl groups as indicated by titration kinetics is decreased under conditions where RNA polymerase exists as a dimer or higher aggregate (low salt), in the presence of Mn2+, or when bound to d(A-T). Incubation of phenylmercurisulfonate with RNA polymerase above pH 9.0 results in loss of d(A-T) binding ability. Poly(U) binding is more sensitive to sulfhydryl modification and is lost as pH's above 8.0. The presence of 4 mM Mn2+ has an obvious effect in stabilizing the polymerase-poly(U) complex when incubated with 10 muM phenylmercurisulfonate + 1 M urea. Incubation of the enzyme with the mercurial and urea results in disaggregation to subprotomeric forms and release of the alpha subunit. Similar treatment in the presence of 4 mM MnSO4 stabilizes the protomeric structure of the enzyme. During chain elongation the enzyme exists as a ternary d(A-T)n-enzyme-r(U-A)n complex in which the bound d(A-T)n is refractory to the destabilizing effect of the mercurial; however, further phosphodiester bond formation is inhibited. The results are defined in terms of a role which reflects the involvement of polymerase sulfhydryl groups in the various conformations necessary for subunit-subunit interaction, tight template binding and catalytic activity.
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PMID:On the role of sulfhydryl groups in the structure and function of the Azotobacter vinelandii RNA polymerase. 24 Apr 5


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