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)

We have synthesized N2-[4-(2,2,6,6-tetramethyl-1-piperidinyloxy)]actinomycin D And the related 1,2-diaminoethane and 1,3-diaminopropane derivatives and evaluated their biological properties. Binding studies with the spin-labeled actinomycin D analogues and DNA were carried out by using circular dichroism, electron spin resonance, and thermal denaturation. These studies have suggested that the derivatives bind to DNA and that their DNA-binding modes are similar but not identical. Spin-labeled actinomycin D derivatives were less potent in inhibiting Escherichia coli DNA-dependent RNA polymerase reaction than actinomycin D and were less toxic to L1210 cells in vitro than the parent compound. Spin-labeled actinomycin D derivatives were more common than the parent compounds against P-388 leukemia cells in vitro with little or no toxicity.
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PMID:Synthesis and biological properties of N2-substituted spin-labeled analogues of actinomycin D. 22 5

Nucleotides containing the fluorophore 1-aminonaphthalene-5-sulfonate attached to the gamma phosphoryl group via a phosphoamidate bond are excellent substrates for Escherichia coli DNA-dependent RNA polymerase. Cleavage of the alpha-beta-phosphoryl bond produces significant changes in both absorption and fluorescence spectra. These alterations provide a sensitive and precise means for continuous monitoring of transcription. Under appropriate conditions one can detect the utilization of less than 1 nmol of nucleotide. Since the spectroscopic techniques measure nucleotide utilization they can be used in conjunction with measurements of incorporation of radiolabeled precursor such as [3H]UTP into acid-insoluble material to determine whether significant amounts of acid-soluble oligonucleotides are formed.
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PMID:Spectroscopic techniques for study of phosphodiester bond formation by Escherichia coli RNA polymerase. 22 88

RNA polymerase I was isolated from parsley cells grown in suspension culture and from soybean hypocotyls. Kinetic studies of the enzyme revealed that RNA polymerase I is an allosteric regulated enzyme. The enzyme activity was influenced by nucleoside triphosphates (NTP) and divalent cations. NTP exceeding a 1:1 ratio of these two components acted as allosteric inhibitors, contrary to free divalent cations, which had promotive effects on the RNA polymerase I. Furthermore, isolated nuclei from parsley exhibited a powerful nucleoside triphosphatase (NTPase) activity. Contrary to RNA polymerase I, this enzyme was stimulated by NTP exceeding the 1:1 ratio of NTP and divalent cations. Free divalent cations had an inhibitory effect. Assuming that a causal connection of these two processes does exist, a possible role of this NTPase would be the control of NTP pools in relation to divalent cations and thus regulating RNA synthesis.
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PMID:RNA polymerase I from higher plants. Evidence for allosteric regulation and interaction with a nuclear phosphatase activity controlled NTP pool. 23 67

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

We have analyzed the transcription of a cloned silkworm tRNA2Ala gene in germinal vesicle extracts of X. laevis oocytes. The primary transcript was sequenced; it is 98 nucleotides long, beginning with a 5' triphosphate nucleotide and ending in a 3' oligouridine stretch. After transcription for long periods of time, enzymes in the frog extract also process the tRNA2Ala precursor to remove extra 5' and 3' nucleotides and to add a CCA end. The twenty-two extra nucleotides at the 3' end of this precursor are recovered as an intact fragment, implicating a new site of endoribonuclease cleavage in eucaryotic tRNA processing. This enzyme activity has also been demonstrated by reincubation of isolated pre-tRNA2Ala with a germinal vesicle extract. The products of in vitro cleavage are the same as those seen in the transcription reactions. The tRNA2Ala precursor molecules are made faithfully in the system with as few as 6 bp of Bombyx morti DNA upstream of the transcription initiation site of the tRNA2Ala gene. This result narrows down the minimal amount of DNA adjacent to the 5' end of a eucaryotic tRNA gene needed to support proper initiation by RNA polymerase III.
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PMID:Transcription of a cloned Bombyx mori tRNA2Ala gene: nucleotide sequence of the tRNA precursor and its processing in vitro. 26 Jun 97

Genomic Xenopus borealis oocyte-specific 5S DNA (Xbo) contains clusters of 5S rRNA genes. The number of genes varies among clusters, and the distance between genes within a cluster is about 80 nucleotides. The spacer DNA between gene clusters is AT-rich and heterogeneous in length due in part to variable numbers of a tandemly repeated 21 nucleotide sequence. A cloned fragment of Xbo 5S DNA (Xbo1) containing three 5S rRNA genes has been sequenced. The sequences of Xbo1 genes 1 and 2 are very similar to the dominant 5S RNA sequence, whereas 15 of the 120 residues in the third gene are different. The sequence of gene 3 is as different from the dominant gene sequence as the X. laevis pseudogene is from the 5S RNA gene. Sequence analysis of genomic DNA shows that gene 3 is an abundant component of the multigene family. All three genes are transcribed when added to an extract of X. laevis oocyte nuclei, and a fragment of Xbo1 lacking the AT-rich spacer DNA and the 5' end of the first gene supports transcription of genes 2 and 3 in this in vitro system. Thus the 80 nucleotides preceding each 5S gene are sufficient for promoter function. Nucleic acid sequences preceding several eucaryotic genes that are transcribed by RNA polymerase III were analyzed and the following common features were found: a purine-rich region; at least one direct repeat; the absence of dyad symmetry; transcription beginning with a purine; a pyrimidine residue immediately preceding the first nucleotide of the gene; and the oligonucleotides AAAAG, AGAAG and GAC, located approximately 15, 25 and 35 nucleotides, respectively, before the start of transcription. The 10 base pair (bp) spacing between the homologous oligonucleotides is that expected for a recognition signal on one face of a DNA double helix. The extensive sequence differences between most of the spacers that precedes these genes make the three conserved oligonucleotides more striking. Parts of the 5' flanking regions of the three Xbo1 gene (-12 to -40), which include the conserved oligonucleotides, are identical. In contrast, 7 of the first 11 nucleotides that precede the third 5S RNA gene in Xbo1 differ from those that precede the first gene. The sequences following the X. borealis oocyte and somatic 5S genes are identical in 12 of the first 14 residues and contain two or more T clusters, as does the corresponding region of X. laevis oocyte 5S DNA. The 3' sequences of the Xenopus 5S rRNA genes and several other eucaryotic genes contain features in common with procaryotic transcription termination sites. The 3' end of the gene is GC-rich and contains a dyad symmetry. Termination occurs in an AT-rich region containing one or more T clusters on the noncoding strand.
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PMID:Nucleotide sequence of Xenopus borealis oocyte 5S DNA: comparison of sequences that flank several related eucaryotic genes. 26 40

Chromatin isolated from immature oocytes was found to contain an endogenous RNA polymerase activity (RNA nucleotidyltransferase; nucleoside triphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) that synthesizes predominately 5S RNA. However, the levels of total RNA synthesis and 5S RNA synthesis in chromatin were each stimulated 10- to 50-fold by an exogenous RNA polymerase III purified from X. laevis oocytes. The 5S genes in chromatin were transcribed by the exogenous enzyme in a highly selective (3000-fold above random) and predominately asymmetric fashion. A significant fraction of 5S RNA sequences were also found in a discrete transcript, approximately 5S in size. Total RNA synthesis was significantly stimulated when chromatin was transcribed by oocyte RNA polymerase I, murine RNA polymerase II, and low levels of Escherichia coli RNA polymerase. However, these enzymes did not significantly stimulate 5S RNA synthesis above the endogenous levels. Both homologous oocyte RNA polymerase I and III and E. coli RNA polymerase transcribed the 5S genes in deproteinized DNA to approximately the same extent (severalfold above random) and both the sense and anti-sense strands of the gene were transcribed. It appears, therefore, that both chromatin-associated components and a purified RNA polymerase III are necessary and sufficient for the selective and accurate transcription of the 5S RNA genes in vitro.
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PMID:Selective and accurate transcription of the Xenopus laevis 5S RNA genes in isolated chromatin by purified RNA polymerase III. 26 93

RNA synthesis in isolated HeLa cell nuclei prepared from cells pretreated with 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) is inhibited in a time-dependent manner. After 40-min pretreatment of cells with 60 muM DRB in the presence of actinomycin D (0.04 mug/ml), the rate of RNA synthesis in isolated nuclei, measured by [(3)H]UTP incorporation, is decreased by 63%. The DRB-resistant one-third of heterogeneous nuclear is distributed over the entire size range of heterogeneous nuclear RNA with some enrichment in the 18S range, as was observed earlier by pulse-labeling whole cells. A subclass of nucleoplasmic RNA molecules is defined in the approximate size range 110 to 250 x 10(3) daltons (330-740 nucleotides). By using heparin (2 mg/ml) to block the synthesis of smaller RNA, a peak in the chain-length range 330-740 nucleotides can be clearly resolved on 2.2% polyacrylamide/1% agarose gels in nuclei from control and DRB-treated cells. The synthesis of these molecules is largely ( approximately 90%) resistant to DRB but sensitive to alpha-amanitin at 1 mug/ml. The in vitro synthesis of molecules in the 140-330 residue range is also sensitive to alpha-amanitin at 1 mug/ml, and it is not at all affected by pretreatment of cells with DRB. Although the synthesis of the RNA in both the 330-740 and the 140-330 residue size ranges appears to be catalyzed by RNA polymerase II, the results with heparin suggest that there may be reinitiation of molecules in the 140-330 size range but not in the 330-740 range in vitro. The synthesis of 4.5S RNA ( approximately 100 nucleotides) and 5S RNA (120 nucleotides) is unaffected by pretreatment of cells with DRB and, as previously reported, is catalyzed by RNA polymerase III, with reinitiation occurring in vitro. Addition of DRB directly to isolated HeLa cell nuclei in vitro has no detectable effect on the overall rate of RNA synthesis.
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PMID:Definition of subclasses of nucleoplasmic RNA. 27 Jul 36

A cell-free system developed from human KB cells was used to transcribe 5.5S RNA from deproteinized adenovirus DNA in vitro. The cell-free RNA synthesis is dependent upon exogenous templates, ribonucleoside triphosphates, and cell-free postmitochondrial supernatant of human KB cells. The synthesis of 5.5S RNA is inhibited only by high levels of alpha-amanitin; therefore it is carried out by RNA polymerase III. The rate of synthesis was linear for at least 2 hr, indicating reinitiation. The 5.5S RNA synthesized in vitro is similar to the corresponding in vivo RNA in size, sequence, and coding region on adenovirus type 2 DNA. In this report is demonstrated in vitro synthesis of a facsimile of an in vivo transcript directed by deproteinized DNA in a mammalian cell-free postmitochondrial supernatant system.
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PMID:Adenovirus DNA-directed transcription of 5.5S RNA in vitro. 27 58

It has been shown that RNA synthesis in isolated hepatopancreas nuclei from Mytilus galloprovincialis is catalyzed by three DNA-dependent RNA polymerases (I, II and III) which resemble those identified in nuclei from mammalian cells. RNA polymerase I is active at 50 mM (NH4)2SO4, catalyzes the synthesis of GMP-rich ribosomal-like RNA and is completely resistant to the toadstool toxin alpha-amanitin. RNA polymerase II and III are active at higher (NH4)2SO4 concentrations, catalyze the synthesis of DNA-like RNA and are inhibited by very low (0.5-1 microgram/ml) and high (200 microgram/ml) concentrations of alpha-amanitin, respectively. Hepatopancreas nuclei retain considerable RNAase activity. Nuclear RNA polymerase activity may be underestimated since a part of the synthetized RNA is degraded.
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PMID:DNA-dependent RNA polymerase activities in hepatopancreas nuclei from Mytilus galloprovincialis Lamarck. 27 40


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