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)

Bleomycin (BLM) exclusively affects thymidine-containing compounds such as DNA and polydeoxyribonucleotides by releasing free thymine and leaving aldehyde functions. Molecular morphology and base sequence of the DNA strongly influence BLM activity. High BLM concentrations, besides modifying DNA into oligothyminic or athyminic nucleic acids, cause strand scissions. Enzymatic DNA and RNA synthesis is strongly influenced by BLM. The inhibition in DNA-dependent DNA polymerase and DNA-dependent RNA polymerase assays is of the non-competitive type. Protein biosynthesis in in vitro systems is not affected by BLM even at high concentrations. BLM turns out to be a strong inhibitor of DNase I and of DNase II; the inhibition is of the competitive type. The enzymatic activities of nucleases using RNA as substrate (RNase A, RNase B, Rnase T1, venom phosphodiesterase I and spleen phosphodiesterase II) are not influenced by this antibiotic. The antibiotic reduces cell proliferation (L5178y mouse lymphoma cells) in vitro in low concentrations by cytostasis and at higher concentrations by cytotoxicity. In BLM-treated L5178y cells, DNA synthesis is strongly reduced, while RNA and protein synthesis are not affected. In vivo, using growing quail oviducts, cell proliferation and cytodifferentiation are markedly inhibited after BLM treatment. This is attributed to the observed inhibition of DNA synthesis. RNA and protein synthesis as well as gene expression are not influenced by BLM under the conditions used. The selective inhibition of DNA synthesis in vivo may be caused by the following mechanisms: (1) competition of BLM with RNA; (2) blocking of the accessibility of DNA in chromatin to BLM, and (3) dependence from the repair processes. BLM inhibits growth of sarcomas, induced by oncogenic RNA viruses in vivo; well-developed tumours show regression after BLM treatment. Transformation of chick embryo fibroblasts by oncogenic RNA viruses in vitro and growth of these viruses is blocked by BLM; the most sensitive period for BLM inhibition is the time during the first period (integration of viral genome into cellular genome?) after infection.
...
PMID:Effect of bleomycin on DNA, RNA, protein, chromatin and on cell transformation by oncogenic RNA viruses. 6 69

A nucleoprotein complex that is an intermediate in viral transcription has been isolated from simian virus 40 (SV40)-infected BSC-1 cells after lysing infected nuclei with Sarkosyl. It contain DNA, DNA-dependent RNA polymerase II, and nascent RNA chains. RNA chain elongation continues for several hours in vitro and is dependent on exogenous ribonucleoside triphosphates. The complex sediments in neutral sucrose gradients with a main peak at about 24 to 26S. When the nascent RNA on the complex is treated with RNase A, a fraction of the RNA remains resistant to RNase and is hydrogen bonded to the DNA template. The pulse-labeled RNase-resistant RNA can be chased into RNase-sensitive RNA, indicating that it is located at the 3' terminus of the RNA chain. The rate of RNA displacement from the DNA template is consistent with an average rate of RNA chain elongation of 15 to 30 nucleotides per min. At least 70% of the RNA synthesized in this in vitro system is SV40 specific. Hybridization with the separated strands of SV40 DNA and with fragments of SV40 DNA generated with endonucleases HindII + III indicates that this RNA is complementary to all regions of the "late" SV40 DNA strand. Studies of SV40 RNA synthesis in this partially purified preparation at early and late times after infection should provide a way of locating promoter sites for transcription and identifying the form of SV40 DNA that serves as a template for late transcription.
...
PMID:Properties of simian virus 40 transcriptional intermediates isolated from nuclei of permissive cells. 19 3

The synthesis of ovalbumin mRNA sequences was studied in isolated nuclei from hen oviduct. Two different methods of analysis were used to distinguish in vitro synthesized from preexisting mRNA sequences: (i) Mercurated ribonucleotides were used for in vitro RNA synthesis, and the newly synthesized RNA was purified by chromatography on sulfhydryl-agarose and hybridized to radioactive ovalbumin cDNA. (ii) [3H]UTP was used to label the in vitro synthesized RNA. Hybridization to unlabeled mercurated cDNA, RNase A digestion, and subsequent purification of the hybrids on SH-agarose allowed the quantitation of newly synthesized ovalbumin mRNA sequences. Approximately 0.1% of the newly synthesized RNA was identified as ovalbumin RNA by both methods. The synthesis of ovalbumin RNA progressed during the incubation of nuclei and was sensitive to actinomycin D and low concentrations of alpha-amanitin. The preferential in vitro transcription of the ovalbumin gene (1000-fold over random transcription of the chicken genome) by RNA polymerase B (nucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) suggests that the specificity of in vitro RNA synthesis is retained in isolated nuclei.
...
PMID:Preferential transcription of the ovalbumin gene in isolated hen oviduct nuclei by RNA polymerase B. 27 31

The diastereomers of uridyl-(3'-5')adenyl-O,O-phosphorothioate [Up(S)A] have been separated by high-performance liquid chromatography. Their identification as RP and SP follows from the RNase A digestion of these products. It was then shown, by the same method, that the R isomer is hydrolyzed by snake venom phosphodiesterase (PDEase) approximately 500 times faster than the S isomer. Similarly, the stereoisomer of adenosine 5'-O-(1-thiotriphosphate) (ATPalphaS), until now arbitrarily designated as isomer B, is hydrolyzed ca 400 times faster by PDEase than is isomer A. From these results it is concluded that the R isomers of Up(S)A and ATPalphaS, isomers B, have the same absolute configuration. It then follows that isomer A of ATPalphaS, the preferred of the two isomers as substrate for DNA-dependent RNA polymerase, has the S configuration. The implications for the stereochemistry of action of the latter enzyme are discussed.
...
PMID:Absolute configuration of the diastereomers of adenosine 5'-O-(1-thiotriphosphate): consequences for the stereochemistry of polymerization by DNA-dependent RNA polymerase from Escherichia coli. 36 98

RNA polymerases pause conspicuously at certain positions on a DNA template. At the well-studied pause sites in the attenuation control regions that precede the trp and his operons, both formation of secondary structure in the nascent transcript and the DNA sequence immediately downstream contribute to pausing. The mechanisms of these effects are unknown. We report here studies on the structure of the RNA and DNA strands in purified trp and his paused transcription complexes in comparison to ten elongation complexes halted by nucleoside triphosphate deprivation. A 14 to 22 nucleotide region of the DNA strands was accessible to modification by KMnO4 or diethylpyrocarbonate in both the paused and halted transcription complexes. However, the region in front of the nucleotide-addition site was reactive only in some halted complexes. In both types of complexes, approximately eight nucleotides on the template strand immediately preceding the 3' end were protected from modification. We also examined the sensitivity of the nascent transcript to RNase A and found that the 3'-proximal eight nucleotide region could be cleaved without complete loss of the potential for elongation. However, a model RNA:DNA hybrid designed to mimic a hybrid in the transcription complex could also be cleaved under similar conditions. Together, the results suggest that the 3'-proximal eight nucleotides of transcript may pair with the DNA template and that this structure is not disrupted by hairpin formation at a pause site. Rather, pausing may result from distinct interactions between RNA polymerase and both the pause RNA hairpin and the downstream DNA sequence.
...
PMID:Structure of RNA and DNA chains in paused transcription complexes containing Escherichia coli RNA polymerase. 128 87

Diagnostic testing for hepatitis C virus (HCV) infection currently is based on the presence of anti-HCV antibodies or a positive HCV RNA polymerase chain reaction (PCR) test. Although HCV RNA PCR is a sensitive and specific technique, widespread application is limited. Moreover, HCV RNA PCR is subject to false-positive reactions through contamination and is inherently difficult to standardize and quantitate. To overcome limitations of HCV RNA PCR, we produced both cDNA and riboprobes from a 241 nucleotide sequence of the 5' untranslated region of the HCV genome for slot hybridization. Hybridization was absent using normal human serum, horse serum, or hepatic cellular RNA from noninfected liver. Hybridization occurred predominantly with positive-stranded HCV RNA and was abolished by pretreatment with RNase A. Slot hybridization was performed on serum samples from 60 patients with chronic HCV infection and a positive HCV RNA PCR and 20 patients with liver diseases unrelated to HCV who had a negative HCV RNA PCR. Slot hybridization with cDNA and riboprobes showed concordance with HCV RNA PCR of 95 and 98.3%, respectively. There were no false-positive reactions in controls. The sensitivity of riboprobe hybridization was comparable to that of one stage HCV RNA PCR using 5' untranslated region primers. Riboprobe hybridization with the HCV H strain standard was positive in the dilution corresponding to 10(-6) chimpanzee infectious doses50/ml. The density of the hybridization signals correlated significantly with the mass of an RNA standard extracted from the liver of a patient with HCV infection. The relative quantities of HCV RNA in the sera of selected patients varied and were not correlated with the duration of disease or the histopathological stage. The highest relative quantities were associated with concurrent immunosuppression. We conclude that slot hybridization is a sensitive, specific alternative to HCV RNA PCR that can be directly quantitated using appropriate HCV RNA standards.
...
PMID:Direct detection of circulating hepatitis C virus RNA using probes from the 5' untranslated region. 131 28

The Saccharomyces cerevisiae transcription factors (TF) IIIB and IIIC assemble onto their respective DNA-binding sites on the SUP4 tRNA(Tyr) gene at 0 degrees C. RNA polymerase III specifically associates at 0 degrees C with this TFIIIC-TFIIIB-DNA complex to form a stable "closed" promoter complex in which the DNA surrounding the transcriptional start retains its duplex form. Promoter "opening" is a temperature-dependent and readily reversible process that involves up to 22 unwound base-pairs of DNA, and can be followed by analyzing the hyperreactivity of thymine to KMnO4 oxidation. This promoter opening increases progressively from 10 degrees C to 40 degrees C, with at least two regions within the transcription bubble appearing to melt independently. In contrast, the temperature dependence of forming an initiated transcription complex containing a 17 nucleotide nascent RNA chain displays a sharp transition between 10 degrees C and 15 degrees C. When RNA polymerase initiates transcription under conditions that limit the nascent RNA chain to less than six nucleotides, there is no displacement of the transcription bubble. These transcription complexes are distinguishable from "open" promoter complexes in their maintenance of the transcription bubble at 0 degrees C, and from transcription complexes with more extended (17 nucleotide) RNA chains in their sensitivity to disruption by heparin. In light of recent results by others that demonstrate a requirement for an RNA transcription factor in a Bombyx mori-based in vitro RNA polymerase III transcription system, we have searched for a comparable component in the S. cerevisiae-derived system. We show that if an RNA component is required in the yeast-derived system, it is not susceptible to inactivation by massive amounts of micrococcal nuclease, RNase A, or RNase T1.
...
PMID:Formation of open and elongating transcription complexes by RNA polymerase III. 161 62

This report describes a novel assay involving the polymerase chain reaction (PCR) and RNase protection for the rapid and sensitive detection of malignant lymphoid cells by nucleotide sequences within their individual rearranged gamma T-cell receptor (TCRG) genes. In this assay, clonal rearrangements are amplified from the DNA of diagnostic tumor specimens using a consensus V segment primer and a consensus J segment primer to which the promoter for T7 RNA polymerase has been appended. The PCR product from this amplification is transcribed into a radiolabeled RNA probe. Test RNA transcribed from the opposite DNA strand is synthesized by similar methods from TCRG genes of a subsequent biopsy specimen. The test RNA is hybridized with the probe, and mismatched nucleotide sequences in the RNA hybrids are digested by RNase A. Detection of fully protected probe by means of polyacrylamide gel electrophoresis and autoradiography indicates the presence of malignant cells in the test specimen. Dilution experiments with DNA of cell lines from acute lymphoblastic leukemias (ALLs) show that detection of one tumor cell among 10(5) normal bone marrow cells is usually possible. Residual disease was also successfully detected in several cases of ALL during clinical remission, including detection in one case at the 10(-5) level. The procedure described here may provide a simplified and rapid method for the sensitive diagnosis and monitoring of lymphoid malignancies. This procedure should be applicable to most antigen receptor genes, and unlike most comparable methods, requires neither analysis of nucleotide sequence nor synthesis of tumor-specific oligonucleotide probes or primers.
...
PMID:Sensitive detection of clonal antigen receptor gene rearrangements for the diagnosis and monitoring of lymphoid neoplasms by a polymerase chain reaction-mediated ribonuclease protection assay. 165 9

New fluorescent derivatives of dinucleoside monophosphates, (5'-AmNS)UpA/ApU/GpU/CpA, with a fluorophore, 1-aminonaphthalene-5-sulfonic acid (AmNS), attached to the first nucleotide of the dinucleoside monophosphates via a 5'-secondary amine linkage were synthesized in good yield. The chemical structure of (5'-AmNS)ApU was proved by the phosphodiesterase digestion followed by Whatman No. 3MM paper chromatographic and spectroscopic analysis of the digested products. The ability of these analogs to be incorporated into the 5' terminus of RNA chain forming fluorescent oligonucleotides by Escherichia coli RNA polymerase was studied in the presence of a synthetic DNA template. The enzymatic reaction of (5'-AmNS)UpA and [3H]UTP in the presence of poly(dA-dT) yielded (5'-AmNS)UpAp[3H]U in greater than 30% yield with the Km values of 5 and 2.5 microM and Vmax values of 17 and 25 nmol/min/mg of enzyme for (5'-AmNS)UpA and UpA, respectively. The structure of this fluorescent trinucleotide was identified by RNase A digestion and paper chromatographic analysis of the digested products. (5'-AmNS)UpA or (5'-AmNS)ApU exhibits two absorption maxima around 270 and 340-350 nm and a fluorescent emission maximum at 445 nm when excited at 340 nm. These spectral characteristics permit their use as energy donors for the transfer of energy to the intrinsic cobalt of the cobalt-substituted RNA polymerases. Upon hydrolysis of the phosphodiester bond of these analogs by venom phosphodiesterase, the absorption at 340 and 270 nm increased by 5 and 20%, respectively, while their fluorescence at 445 nm was enhanced by 25%. Thus, these analogs can be used for studying the dynamics of initiation and elongation reactions catalyzed by DNA-dependent RNA polymerases by absorption and fluorescence spectroscopies.
...
PMID:Synthesis and characterization of fluorescent dinucleotide substrate for the DNA-dependent RNA polymerase from Escherichia coli. 244 Aug 71

Dinucleotides (3'-5')-ApU and UpA and their 3'-O-phosphonylmethyl and 5'-O-phosphonylmethyl analogues were studied as substrates in the primed abortive synthesis catalysed by Escherichia coli DNA-dependent RNA polymerase on poly[d(A-T)] template. All phosphonate analogues of dinucleotides containing the anomalous sugar-phosphate backbone are substrates for the holoenzyme as verified by RNase A and RNase T2 digestion of the trinucleotide analogues obtained. The finding that phosphonate dinucleotides act as primers for transcription indicates that steric requirements at the initiation site are not as specific as previously supposed. Analysis of kinetic constants of ordered bibi reaction Kia, KmA, KmB and Vmax suggests that the instability of short RNA-DNA hybrids contributes to the abortive release of trinucleotides formed.
...
PMID:Phosphonate analogues of dinucleotides as substrates for DNA-dependent RNA polymerase from Escherichia coli in primed abortive initiation reaction. 248 57


1 2 3 4 Next >>

---