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

---

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

We have analyzed the kinetics of DNA synthesis catalyzed by reverse transcriptase from human immunodeficiency virus 1 (HIV-1). Reverse transcriptase, overproduced in Escherichia coli and purified to homogeneity, has polymerase and RNase H activity. Reverse transcriptase forms a stable complex with poly(rA).oligo(dT) primer-templates in the absence of Mg2+ and dTTP with an equilibrium dissociation constant of 3 nM. Synthesis from these preformed complexes can be initiated, and restricted to a single processive cycle, by the simultaneous addition of Mg2+, dTTP, and excess competitor RNA. Preformed complexes decay with a maximal half-life of 2-3 min. Synthesis on poly(rA) templates is processive with an incorporation rate of 10-15 nucleotides/s at 37 degrees C. Processivity varies widely with the template used, increasing from a few to greater than 300 nucleotides in the order: poly(dA) less than double-stranded DNA less than single-stranded DNA less than single-stranded RNA less than poly(rA). On double-stranded DNA reverse transcriptase catalyzes limited strand-displacement synthesis of up to 50 nucleotides. On RNA-DNA hybrids significant DNA synthesis is observed only after degradation of the RNA strand by the RNase H activity of reverse transcriptase. Intermolecular strand switching occurs with poly(rA) templates. At low ionic strength reverse transcriptase can use multiple templates with a single primer, leading to products of greater than template length. Reverse transcriptase and primer do not have to dissociate during the exchange of template strands, thus allowing processive DNA synthesis across template borders.
...
PMID:Human immunodeficiency virus 1 reverse transcriptase. Template binding, processivity, strand displacement synthesis, and template switching. 246 38

Reverse transcriptase from the simian immunodeficiency virus (SIV) was found to have kinetic behavior similar to that of enzyme from the human immunodeficiency virus (HIV). Michaelis constants for the substrates TTP and dGTP and inhibition constants for the inhibitors 3'-azido-3'-deoxythymidine 5'-triphosphate, 2',3'-dideoxythymidine 5'-triphosphate, and 2'-3'-dideoxyguanosine 5'-triphosphate were obtained for SIV reverse transcriptase and were found to be similar to the corresponding values for HIV reverse transcriptase. Thus, the interaction of SIV reverse transcriptase with nucleotide analogs appears to be indistinguishable from that of the HIV enzyme, suggesting that SIV/simian acquired immunodeficiency syndrome (SAIDS) is a potentially good model of AIDS.
...
PMID:Kinetics and inhibition of reverse transcriptase from human and simian immunodeficiency viruses. 246 88

Racemic carbocyclic analogues of dTTP [(+/-)-C-dTTP] and its ribo counterpart, 5-methyl-UTP [(+/-)-C-m5UTP] were synthesized and examined, in comparison with dTTP and UTP (and m5UTP), as potential substrates of E. coli DNA and RNA polymerases, respectively. Unexpectedly, only a very low (terminal) incorporation of C-dTMP into DNAs of different structure was observed, C-dTTP did not serve as a substrate for chain elongation by the Klenow DNA polymerase. Inhibition of DNA replication was, however, observed in the presence of (+/-)-C-dTTP. The UTP analogue, (+/-)-C-m5UTP proved neither a substrate nor an inhibitor of the RNA polymerase enzyme.
...
PMID:Carbocyclic analogues of dTTP and UTP: properties in polymerase enzyme-catalyzed reactions. 332 Sep 75

Anticapsin, the terminal epoxyaminoacid moiety of tetaine, inhibits irreversibly growth of HeLa S3 cells. The antibiotic decreases to a similar extent incorporation of 3H-labelled precursors into nucleic acids and protein in intact cells: inhibition of protein synthesis prevails on prolonged incubation. Also incorporation of [3H]dTTP and [3H]UTP is inhibited in the presence of anticapsin into permeabilized cells. These effects, however, are not due to the interference with DNA or RNA polymerases since anticapsin only slightly suppresses RNA polymerase activity and has no effect on DNA polymerase in the cell-free systems. The results indicate that the mechanism of antiproliferative action of anticapsin in HeLa S3 cells differs from that of tetaine and imply that inhibition of protein synthesis might be the primary effect of anticapsin.
...
PMID:Pleiotropic effect of anticapsin on HeLa S3 cells. 345 99

A photoactive nucleotide analogue of dUTP, 5-azido-2'-deoxyuridine 5'-triphosphate (5-N3dUTP), was synthesized from dUMP in five steps. The key reaction in the synthesis of 5-N3dUTP is the nitration of dUMP in 98% yield in 5 min at 25 degrees C using an excess of nitrosonium tetrafluoroborate in anhydrous dimethylformamide. Reduction of the resulting 5-nitro compound with zinc and 20 mM HCl gave 5-aminodeoxyuridine monophosphate (5-NH2dUMP). Diazotization of 5-NH2dUMP with HNO2 followed by the addition of NaN3 to the acidic diazonium salt solution gave a photoactive nucleotide derivative in 80-90% yield. The monophosphate product was identified as 5-N3dUMP by proton NMR, UV, IR, and chromatographic analysis as well as by the mode of synthesis and its photosensitivity. After formation of 5-N3dUTP through a chemical coupling of pyrophosphate to 5-N3dUMP, the triphosphate form of the nucleotide was found to support DNA synthesis by Escherichia coli DNA polymerase I at a rate indistinguishable from that supported by dTTP. When UMP was used as the starting compound, 5-N3UTP was formed in an analogous fashion with similar yields and produced a photoactive nucleotide which is a substrate for E. coli RNA polymerase. To prepare [gamma-32P]-5-N3dUTP for use as an active-site-directed photoaffinity labeling reagent, a simple method of preparing gamma-32P-labeled pyrimidine nucleotides was developed. [gamma-32P]-5-N3dUTP is an effective photoaffinity labeling reagent for DNA polymerase I and was found to bind to the active site with a 2-fold higher affinity than dTTP.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Synthesis and biological properties of 5-azido-2'-deoxyuridine 5'-triphosphate, a photoactive nucleotide suitable for making light-sensitive DNA. 354 18

Activity of RNA polymerases I, II and III (distinguished using alpha-amanitin) and activity of DNA polymerases alpha and beta (distinguished using N-ethylmaleimide) were assayed for varying intervals and at varying substrate (UTP or dTTP) concentrations in the purified nuclear fraction from corneal epithelium of carbamylcholine-treated and control eyes of rabbits with resurfacing acid burn defects. Incorporation was linear with time for all enzymes up to 30 min. In 10 min assays at varying substrate concentrations, all polymerases from carbamylcholine-treated eyes had significantly elevated Vmax compared to corresponding control enzymes. The drug also increased apparent affinity of RNA polymerase II for UTP and apparent affinity of DNA polymerases alpha and beta for dTTP. Results are discussed in relation to potential mechanisms by which effects of carbamylcholine on polymerase activity may be mediated.
...
PMID:Activity of DNA and RNA polymerases in resurfacing rabbit corneal epithelium. 358 70

Plasmid DNA containing the adenovirus type 2 genes for VA RNA was linearized at a site distal to the gene, end labeled with a biotin-nucleotide analog of TTP, and incubated with avidin to form an avidin-biotinylated DNA complex. HeLa cell S100 extracts containing crude RNA polymerase III and transcription factors (TFs) IIIB and IIIC were programmed with the avidin-biotin-VA DNA to allow stable complex formation (A.B. Lassar, P.L. Martin, and R.G. Roeder, Science 222:740-748, 1983). Chromatography of the programmed extract over a biotin-cellulose affinity resin resulted in the selective, and virtually quantitative, retention of one of two stable preinitiation complexes, either VA-IIIC or VA-IIIC-IIIB, depending on the length of template incubation in the S100 extract. After washing the resin with 0.10 M and 0.25 M KCl to remove RNA polymerase III and nonspecifically bound proteins, respectively, TFIIIC was eluted from the VA-IIIC complex by the addition of 1.5 M KCl. The VA-IIIC-IIIB complex exhibited a higher salt stability. Most of TFIIIB and some TFIIIC were released by the addition of 1.5 M KCl; however, the majority of TFIIIC activity was recovered only after a subsequent 3.0 M KCl elution. The specific activity of the TFIIIC in the 3.0 M KCl fraction was 770-fold higher than that in the S100 extract, while the protein content of the 1.5 and 3.0 M KCl fractions was reduced 7,500- and 100,000-fold, respectively.
...
PMID:Rapid enrichment of HeLa transcription factors IIIB and IIIC by using affinity chromatography based on avidin-biotin interactions. 378 24

DNA and RNA polymerase activities in the purified nuclear fraction from cultured rabbit corneal epithelial cells were assayed over a range of substrate (labeled dTTP or UTP) concentrations using calf thymus DNA as template. Effects of carbamylcholine on polymerase activities were evaluated over a range of drug concentrations including those saturating muscarinic receptors. Carbamylcholine significantly (p less than 0.001) enhanced activity of both polymerases, both in nuclei incubated with the drug during assay and in nuclei from carbamylcholine-treated cells. Drug effects were blocked by atropine. Regression analysis of Hill plots for variation of polymerase activity with carbamylcholine concentration indicated half-maximal activity of both polymerases at approximately 1 microM carbamylcholine. Mechanisms by which carbamylcholine may alter polymerase activities are discussed in relation to effects of the drug on nuclear enzymes of cyclic nucleotide metabolism and on cyclic nucleotide-dependent protein phosphorylation.
...
PMID:Cyclic nucleotides in muscarinic regulation of DNA and RNA polymerase activity in cultured corneal epithelial cells of the rabbit. 406 34

Various 5-substituted 1-beta-D-xylofuranosyluracil 5'-triphosphates (hydrogen, methyl-, ethyl-, n-propyl, n-butyl, fluoro-, chloro-, bromo-, and iodo derivatives) and some of the 3'-deoxyribofuranosyl nucleotides (3'-deoxy UTP and 3'-deoxy TTP) were synthesized chemically and their inhibitory effects on DNA-dependent RNA polymerases I and II of the cherry salmon (Oncorhynchus masou) were studied systematically. These 3'-modified UTP analogues could not be utilized as substrates in place of UTP, but they did inhibit the incorporation of UMP into RNA in vitro. In contrast, 2'-modified UTP analogues, such as 2'-dTTP and Ara TTP, were neither substrates nor inhibitors. Kinetic analysis showed that the inhibition by these compounds was essentially competitive with substrate UTP. The K1 values of RNA polymerase I for the analogues were smaller (2-6 microM) than the Km value for UTP (8 microM), but those for xylo-EtUTP, xylo-PrUTP, and xylo-BuUTP were larger (about 20 microM) than the Km for UTP. In contrast to these alkyl groups with steric and electron-donating effects, halogen groups have electron-withdrawing effects on the uracil nucleus. Therefore, it was concluded that the inhibitory activity of these analogues on RNA polymerase I was not affected by the inductive effects of substituent groups at the 5-position of uracil nucleus but by their steric effects. On the other hand, all of the K1 values of RNA polymerase II for UTP analogues were smaller (0.4-3 microM) than the Km value for UTP (4 microM). In this case, neither steric effect nor an inductive effect of substituents on UTP analogues influenced the inhibitory activity towards RNA polymerase II.
...
PMID:Differential inhibitory effects of 5-substituted 1-beta-D-xylofuranosyluracil 5'-triphosphates and related nucleotides on DNA-dependent RNA polymerases I and II from the cherry salmon (Oncorhynchus masou). 406 48

Nuclei and hypotonically leached extracts of nuclei prepared from tomato golden mosaic virus (TGMV)-infected Nicotiana benthamiana leaves have been used in in vitro DNA and RNA polymerisation reactions. The synthesis of virus-specific DNA was resistant to aphidicolin, sensitive to N-ethylmaleimide and dideoxy TTP, and stimulated by KC1 and ATP. Variably virion (+) and complementary (-) strand DNA of both the A and B genomic components were synthesised. Virus-specific RNA was synthesised in reactions which were initiated prior to nuclei isolation and leaching. From inhibitor studies and salt requirements RNA synthesis appeared to be catalysed by a DNA-dependent RNA polymerase type II enzyme. Both components of the TGMV genome were transcribed in a bidirectional fashion with a prevalence in some experiments of transcripts derived from DNA component A.
...
PMID:DNA and RNA polymerase activities of nuclei and hypotonic extracts of nuclei isolated from tomato golden mosaic virus infected tobacco leaves. 406 99


<< Previous 1 2 3 4 5 Next >>

---