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)

Localized mutagenes of Salmonella typhimurium followed by a [3H]uridine enrichment procedure yielded a temperature-sensitive strain with a mutation in the rpo region of the chromosome. Ribonucleic acid (RNA) polymerase (EC 2.7.7.6; nucleoside triphosphate: RNA nucleotidyltransferase) purified from this mutant was considerably less active at the nonpermissive temperature than wild-type enzyme. Furthermore, the enzyme from this mutant, unlike RNA polymerase of previously isolated temperature-sensitive mutants, was as thermostable as wild-type enzyme when preincubated at 50 degrees C. Subunit reconstitution experiments have shown that the temperature sensitivity is caused by an alteration in the beta' subunit of the enzyme.
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PMID:Temperature-sensitive ribonucleic acid polymerase mutant of Salmonella typhimurium with a defect in the beta' subunit. 78 38

The phosphodiester bond formation by DNA-dependent RNA-polymerase (RNA nucleotidyltransferase, nucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) can in principle result in retention, inversion, or racemization of configuration at the alpha-phosphorus of the nucleoside 5'-triphosphate being polymerized. As a first step in elucidating the stereochemistry of this reaction, one diastereomer (A) of adenosine 5'-O-(1-thiotriphosphate) (ATPalphaS) was polymerized with UTP in the presence of poly(dA-dT)-poly(dA-dT). The resulting polymer was enzymatically cleaved to uridine 2',3'-cyclic phosphorothioate which was determined to be the endo-isomer by comparison with an authentic sample. This shows that no reacemization had occurred and that isomer A of ATPalphaS gives a phosphorothioate diester bond with the R-configuration. Whether this represents inversion of retention of configuration awaits elucidation of the absolute configuration of isomer A for ATPalphaS.
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PMID:Stereochemistry of polymerization by DNA-dependent RNA-polymerase from Escherichia coli: an investigation with a diastereomeric ATP-analogue. 78 80

1. EDTA inhibited incorporation of [3H]uridine into RNA of lymphocytes, but did not decrease uptake into the cold-acid-soluble fraction of the cells. The inhibition by EDTA was largely reversible by simultaneous addition of Zn2+. 2. Low concentrations pf actinomycin D (3 ng/ml) added at the time of stimulation of the cells inhibited [3H]uridine incorporation into RNA, but concentrations of 50-100 ng/ml were required to produce the same degree of inhibition if addition of actinomycin D was delayed until just before the incorporation was measured. This difference in sensitivity did not reg within the cells. 3. When added immediately before phytohaemagglutinin, actinomycin D (3 ng/ml) and EDTA produced similar time-courses of inhibition of uridine incorporation. 4. Uridine incorporation at 32h was inhibited when actinomycin D (3 ng/ml) or EDTA was added just before stimulation of the cells, but was only slightly affected when they were added at 32h. At intermediate times the incorporation of uridine remained sensitive to addition of EDTA for longer than it was sensitive to actinomycin D. 5. Polyacrylamide-gel separation of RNA synthesized in EDTA-treated cultures in the presence or absence of added Zn2+ showed that lower availability of Zn2+ resulted in a decreased rate of transfer of radioactivity from 32S to 28S rRNA and decreased survival of 28S rRNA relative to 18S rRNA. 6. Close similarities have been shown to exist between the effects of EDTA and low concentrations of actinomycin D. Not all the effects of EDTA could be explained by postulating that Zn2+ was a constituent of RNA polymerase I, nor were the effects of actinomycin D readily explained by previously suggested mechanisms of action of this antibiotic.
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PMID:Comparison of the effects of zinc deprivation and actinomycin D on ribonucleic acid synthesis by stimulated lymphocytes. 81 Jan 42

A comparative radioautographic study of the RNA precursors incorporation on polytene chromosomes of Drosophila in vivo in the cells of salivary glands, and in vitro during incubation of E.coli RNA polymerase on slides with fixed chromosomes was performed.--The pattern of in vivo 3H-uridine incorporation on different sections of the chromosomes drastically differed from the in vitro 3H-UTP incorporation which seems to be much more related to DNA content of the individual small sections. In both cases puffing of the loci resulted in the increase of RNA synthesis but in vitro only 2-3 fold and in vivo much more. Hence, RNA synthesis in vitro was unspecific and did not reflect the in vivo RNA synthesis.--On the other hand, E.coli RNA polymerase completely mimics in vitro the dosage compensation phenomenon making twice as much RNA on one X-chromosome of males (1X2A) as on each of X-chromosomes of diploid (2X2A) and triploid (3X3A) females and super-females (3X2A), and the intermediate amount of RNA on each of X-chromosomes of intersexes (2X3A). It is suggested that the differences in the in vitro template activity of X-chromosomes of cells with different X:A ratio are due to different extent of condensation of their deoxyribonucleoprotein (DNP). Yet, both male and each of female X-chromosomes bind the same amount of thymus histone FI labelled with fluorochrome which indicates that they contain the same amount of "open" regions with exposed chromosomal DNA accessible to external proteins.--On the basis of these observations a hypothesis is put forward which suggests that RNA transcription in animal chromosomes is regulated at two levels by different mechanisms; the first one controls the extent of condensation of DNP of genetic loci and determines their competence to the second mechanism which involves the action of gene-specific activator proteins. According to this hypothesis the phenomenon of dosage compensation of sex-linked genes is due to decondensation of DNP of male X-chromosome which renders its loci twice as responsive to activators as compared to the same loci in females.
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PMID:Comparison of in vivo and in vitro RNA synthesis on polytene chromosomes of Drosophila. 81 77

A transcriptionally active chromosome has been isolated in highly purified form from choroplasts of Euglena gracilis, It contains chloroplast DNA, DNA-dependent RNA polymerase, and other proteins. Transcription occurs at low levels of endogenous DNA, and is indifferent to high levels of exogenous DNA. RNA chain elongation continues for several hours in vitro, and RNA chain initiation, determined by [gamma-32P]ATP incorporation, is continuous for at least 1 h in vitro. Maximal rates for RNA synthesis require only a divalent cation and the four ribonucleoside triphosphates. Apparent Km values for adenosine triphosphate, cytidine triphosphate, guanosine triphosphate, and uridine triphosphate are 4.0, 0.6, 2.5, and 2.3 muM, respectively. As would be expected for a DNA-dependent RNA polymerase, RNA synthesis is inhibited by actinomycin D. However, rifampicin and streptolydigin, inhibitors of procaryotic RNA synthesis, and alpha-amanitin, an inhibitor of eucaryotic nuclear RNA polymerases II and III, do not inhibt the RNA synthesis reaction. Heparin, which is a potent inhibitor of the initiation of RNA synthesis by a nontemplate bound RNA polymerase, also does not inhibit RNA synthesis. Isolation of transcriptionally active chromosomes should prove to be a useful method to study the mechanism of selective RNA transcription of eucaryotic chromosomes.
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PMID:Isolation of a transcriptionally active chromosome from chloroplasts of Euglena gracilis. 82 16

Total RNA polymerase activity, as well as the proportion of alpha-amantin-sensitive and resistant during activity, have been measured in the posterior silk glands of the silkworm as a function of growth the fifth larval instar. During the first 5 days, termed the growth phase, the total enzyme activity and particularly the portion that is alpha-amantin-resistant increases to reach a peak value and thereafter declines during the secretory phase, Much of the enzyme remains firmly bound and insoluble. Heparin only only does not inhibit this insoluble and probably chromatin-bound activity which would indicate lack of initiation, but it enhances the activity. A large proportion of newly transcribed RNA is released from the transcription complex. The synthesis of RNA has been studied both qualitatively and quantitatively during the same period. RNA synthesis becomes important on the second day of the fifth instar, as does the RNA polymerase, and stays at a high level for several more days. The results from these studied as well as those with incorporation of 32P indicate interference of varying precursor pools in quantitatively measured RNA synthesis. However, RNA content as well as RNA synthesis in vitro show a close correlation with RNA polymerase activity. The labeled RNAs extracted at different days of the fifth instar have been fractioned on sucrose gradients; this demonstrated that the predominant product of RNA synthesis, as followed by [3H]uridine incorporation at short time intervals, is 45-S preribosomal RNA and 4-5 S RNA. The 45-S RNA is transformed to 19-S and 30-S ribosomal RNA as time progresses or after a chase with unlabeled and/or actinomycin D. There also exists a component heavier than 45S which is fairly rapidly labeled to a small extent.
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PMID:RNA polysomes and RNA synthesis in the silk glands of the silkworm Bombyx mori;. 83 21

1. Purified form A RNA polymerase and the endogenous, nuclear form A RNA polymerase are shown to incorporate 5-mercuri-uridine 5'-triphosphate (Hg-UTP) into RNA in vitro. The Km for both Hg-UTP and UTP are in the region of 10 muM for the purified enzyme. 2. The RNA products formed in nucleoli by endogenous RNA polymerase A have similar base compositions (G + C-rich) whether UTP or Hg-UTP is provided as the substrate in vitro. 3. Sulphydryl-Sepharose chromatography of RNA synthesised in vitro by nucleoli allows separation of this material from the endogenous RNA, when the former is synthesised in the presence of Hg-UTP. 4. In-vitro-synthesised nucleolar RNA hybridises with cot profiles similar to 28-S ribosomal RNA, when made with either Hg-UTP or UTP. 5. Hybridisation studies using DNA excess suggest that little competition occurs between the in vitro transcripts and the endogenous nucleolar RNA. 6. Size analysis of in vitro transcripts show that although some degradation occurs during isolation, purification and hybridisation of the RNA species, most of the RNA remains larger than 5 S throughout.
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PMID:The use of mercurated nucleoside triphosphate as a probe in transcription studies in vitro. 95 55

The 3' terminal nucleosides of RNA transcribed in vitro by E. coli RNA polymerase from T7 DNA and UV irradiated TN DNA were determined. The 3' terminal nucleoside of naturally terminated (t1 termination site) RNA cytidine. In the case of RNA terminated at UV lesions, it is cytidine in 0 per cent of the molecules and adenosine in the remaining 30 per cent. Cytidine trialcohols are labile in high concentrations of KOH and at high temperature and appear to convert to uridine.
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PMID:End group of naturally terminated and UV lesion terminated T7 in vitro RNA. 100 22

Administration of phenobarbital in rats is followed by an in vitro stimulation of hepatic ribosomal protein synthesis. The increase of polyribosomes which is also observed is paralleled by a phenobarbital-dependent enhancement of uridine incorporation into ribonucleic acids. Sedimentation of either ribosomes or ribosomal RNA extracted from ribosomal subunits through sucrose gradients reveals stimulation by phenobarbital of 3H-uridine incorporation into messenger RNA as well as into ribosomal RNA. These experiments together with the findings of stimulation of RNA polymerase activity and blockade of the effects of phenobarbital by the administration of Actinomycin D indicate a primary action of phenobarbital on RNA synthesis.
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PMID:Stimulation of hepatic RNA synthesis by phenobarbital. 100 24

Mercurated uridine triphosphate has been used to label transcripts of chicken reticulocyte chromatin made with Escherichia coli RNA polymerase. The mercury-labeled RNA product can be completely separated from endogenous RNA sequences in the chromatin by passage through a sulfhydryl Sepharose column. Globin cDNA hybridization to the transcript shows that only 2.6 x 10-5 of the transcript is globin RNA. In contrast to this result, erythrocte chromatin transcript contains less than one tenth as many globin RNA sequences.
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PMID:In vitro transcription of chromatin in the presence of a mercurated nucleotide. 106 24


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