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 interaction of granaticin B, a quinone antibiotic produced by Streptomyces granaticolor, with some biologically important bivalent metal ions, DNA and ATP was demonstrated spectrophotometrically. The activity of isolated RNA polymerase was higher when the DNA of phage SP 50 served as template than with DNA isolated from Bacillus subtilis. Granaticin B inhibited in vitro RNA synthesis, similarly to certain other antibiotics (the inhibition was three times lower than that caused by actinomycin D or streptolydigin and slightly higher than that by epsilon-pyrromycinone). The inhibitory effect was higher when the Mg2+ concentration in the reaction mixture was decreased. The inhibition was then proportional to the concentration of the DNA template. DNA-dependent RNA synthesis is thus inhibited in vitro by granaticin B but this does not appear to be the only site of action of this antibiotic in vivo.
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PMID:Interaction of granaticin B with the transcription system of Bacillus subtilis. 41 19

Some properties of unprimed poly(A)-poly(U) synthesis by DNA-dependent RNA polymerase from Caulobacter crescentus were examined. The reaction required ATP and UTP as substrates and manganese as a divalent cation. Rifampicin completely inhibited the reaction at a concentration of 1 micron/ml, and the enzyme catalyzed the polymer synthesis well regardless of the presence of GTP, CTP or both. The chain length of the poly(A)-poly(U) synthesized was about one hundred base pairs, as estimated from a sedimentation velocity and the molar ratio of [3H]AMP to [gamma-32P]ATP incorporated into the poly(A)-poly(U). The reaction was dependent on the square of the enzyme concentration and the enzyme dimers formed complexes with poly(A)-poly(U) during the reaction.
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PMID:Properties of unprimed poly(A)-poly(U) synthesis by Caulobacter crescentus RNA polymerase. 42 57

When protein biosynthesis is inhibited by either cycloheximide of puromycine, the nucleolar RNA synthesis of Ehrlich ascites tumor cells decreases by approximately 70% within 1 h, while the removal of these protein synthesis inhibitors causes a rapid recovery of nucleolar RNA synthesis, largely within 1 h. A similar pattern of decrease and recovery of endogenous RNA polymerase activity in isolated nucleoli or in nuclei (in the presence of alpha-amanitin) may be demonstrated after addition and removal of these drugs. Analysis of the molecular species of RNA polymerase I on a phosphocellulose column indicates that only the IB form of the enzyme decreases in the nucleoli of drug-treated cells and recovers quickly after resumption of protein synthesis. The finding that the activity of the IB form enzyme remains unchanged in the whole nuclei indicates that during cessation of protein synthesis RNA polymerase IB is either released from the nucleoli into the extranucleolar compartment or becomes so loosely bound to the nucleoli that it is leached out from the nucleoli during their isolation. By using a system of assaying free, nucleolar-template bound and total RNA polymerase I activities, data supporting the above interpretation have been obtained. Namely, in isolated nuclei free enzyme activity increases with a concomitant decrease in bound enzyme activity during protein synthesis inhibition, while the total enzyme activity remains unchanged. In isolated nucleoli, both total and bound enzyme activities decreases on protein synthesis inhibition but recover quickly on its resumption. The putative bound enzyme, fractionated with the aid of actinomycin D, is exclusively IB form, whereas the unbound enzyme consists of both IA and IB forms as previously demonstrated (1). No conversion of IB form polymerase to IA form was noted on prolonged sonication in our system. The levels of ATP and GTP in the cell did not change appreciably either during cessation or resumption of protein synthesis in these cells. The data support the previous conclusion that some short-lived protein(s) is required to maintain the normal level of ribosomal RNA transcription (2) and further suggest that the protein is required to facilitate reinitiation of the transcription by RNA polymerase IB in the nucleolus.
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PMID:The mechanism of decrease in nucleolar RNA synthesis by protein synthesis inhibition. 42 65

Some properties of in vitro transcription by isolated Xenopus oocyte nucleoli were described. When incubated with labeled RNA precursors, Xenopus oocyte nucleoli exhibited prolonged incorporation of radioactivity into RNA. The synthetic activity was exclusively due to type I RNA polymerase as revealed by its insensitivity to low and high doses of alpha-amanitin. The size of the in vitro transcript was mostly larger than 28S at 10 minute incubation and became smaller as incubation proceeded. When [gamma-32P]ATP was included in the reaction mixture, 32P radioactivity was incorporated into RNA suggesting the possible initiation of transcription in this system. However, analysis of the terminal nucleotide of the transcript revealed that the incorporation of radioactivity from [gamma-32P]ATP was not due to the initiation of transcription but due to polynucleotide kinase activity in the nucleolar preparation. These results demonstrate that the incorporation of radioactivity from [gamma-32P] labeled nucleoside triphosphates cannot necessarily be regarded as an index of the initiation of transcription.
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PMID:Properties of in vitro transcription by isolated Xenopus oocyte nucleoli. 45 Jul 18

Although cordycepin 5'-triphosphate (3'-dATP), at low concentrations, preferentially inhibits chromatin-associated poly(A) synthesis in isolated nuclei, higher levels of the inhibitor prevent both rRNA (RNA polymerase I activity) and hnRNA (RNA polymerase II activity) synthesis in vitro (Rose, K.M., Bell, L.E. and Jacob, S.T. (1977) Nature 267, 178-180). The present studies demonstrate that this nucleotide can also inhibit tRNA and 5 S RNA synthesis (RNA polymerase III activity). At 50-200 microgram/ml, 3'-dATP inhibits incorporation of [3H]UTP into tRNA and 5 S RNA by approximately 65%, whereas the syntheses of these RNAs were completely blocked when [3H]GTP was used as the substrate. These data suggest the formation of poly(U) in the tRNA and 5 S RNA regions, which is resistant to 3'-dATP. In contrast, another ATP analog, Ara-ATP, which selectively inhibits poly(A) synthesis, does not block tRNA and 5 S RNA synthesis in isolated nuclei. The production of these RNA species in isolated nuclei is also insensitive to Ara-CTP and 2'-dATP. These data suggest that 3'-dATP exerts general inhibitory effects on RNA synthesis and further substantiate the conclusion that Ara-ATP is a selective inhibitor of the polyadenylation reaction in vitro.
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PMID:Differential effects of cordycepin triphosphate and 9 beta-D-arabinofuranosyladenine triphosphate on tRNA and 5 S RNA synthesis in isolated nuclei. 49 5

Bacteriophage T3-induced RNA polymerase is rapidly inactivated at 42 degrees C. Addition of T3 DNA delays this process for 30 s and reduces the rate with which the enzyme activity is lost indicating that a labile binary complex between T3 DNA and polymerase must have been formed. The ternary complex between T3-specific RNA polymerase, T3 DNA, and nascent RNA chains obtained when the enzyme is incubated with T3 DNA, GTP, ATP, and UTP is stable to heat (42 degrees C) and only slowly inactivated by polyvinyl sulfate. The optimal temperature for the formation of polyanionresistant ternary complexes is 30 degrees C while the elongation of T3 RNA chains proceeds fastest at 38 degrees C.
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PMID:Effect of temperature on the transcription by bacteriophage T3-induced RNA polymerase. 54 12

Amino acid starvation of Ehrlich ascites cells leads to a significant decrease of the intracellular ATP concentration concomitant with a marked decrease in nucleolar RNA polymerase activity. Addition of 8-bromoguanosine 3':5'-monophosphate (br8cGMP) to the amino-acid-deficient culture medium increased the cellular ATP levels and restored the rRNA synthesis capacity of nucleoli to control levels. Exogenous br8cAMP overcame the effects of br8cGMP. Administration of br8cAMP to exponentially growing ascites cells resulted in a shrinkage of ATP levels and in an inhibition of nucleolar RNA synthesis similar to that observed under shift-down conditions. These effects of br8cAMP could be antagonized by exogenous br8cGMP or hypoxanthine. Since the br8cGMP-induced increase in the total adenine nucleotides was abolished in the presence of azaserine (an inhibitor of the amidation of formylglycineamide ribonucleotide) it is concluded that cyclic nucleotides exert at least a part of their regulatory effects on cell proliferation by regulating nucleotide biosynthesis de novo.
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PMID:The effect of cyclic nucleotides on cellular ATP levels and ribosomal RNA synthesis in Ehrlich ascites cells. 56 46

The steady state kinetics of initiation of T7 DNA transcription by RNA polymerase holo enzyme from E. coli in the presence of rifampicin and the two substrates ATP and UTP were studied. Under these conditions, the enzyme catalyzes exclusively the promotor specific synthesis of pppApU. The kinetic data are in agreement with the mechanism of a truly ordered reaction. Binding of the initiating nucleotide ATP to the transcriptional complex occurs prior to the binding of the substrate UTP. Release of pppApU is most probably the rate limitinig step. Km constants were found to be 0.6 mM for ATP and 0.31 mM for UTP, respectively. The substrate inhibition pattern indicated that the substrate site exhibits a finite affinity for incorrect nucleoside triphosphate (Ki = 2.3 mM). A similar non specific binding to the 3-OH site could not be demonstrated.
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PMID:Steady state kinetic studies of initiation of RNA synthesis on T7 DNA in the presence of rifampicin. 59 91

HeLa nuclear homogenates incubated in vitro incorporate [beta-32P]ATP and S-[methyl-3H]-adenosylmeth-ionine ([3H]SAM) into blocked methylated 5' termini of newly synthesized RNA. Approximately 10% of the RNA chains initiated in vitro with [beta-32P]ATP are subsequently blocked by condensation of GMP to di- or triphosphate terminated RNA. The blocked termini can then be methylated by transfer of methyl groups from [3H]SAM to the 7 position of the guanosine and 2'-O position of the adenosine to form m7Gpp*pAm- capped terminus. In addition to conventional triphosphate caps, HeLa nuclear homogenates produce capping structures containing two phosphate residues in the pyrophosphate bridge. The two distinct cap forms were separated by DEAE-cellulose chromatography and analyzed. In contrast to triphosphate caps (m7GpppXm) in which X can be any one of the four nucleosides (G, A, C, or U), in diphosphate caps (m7GppXm), more than 95% of the penultimate nucleoside Xm is G. Incorporation of both [beta-32P]ATP and [3H]SAM into caps was markedly reduced by low concentrations of alpha-amanitin. However, an ammonium sulfate fraction of the nuclear homogenate can cap beta-32P-labeled RNA (pp*pA-RNA) to form m7Gpp*pA-RNA, in the presence of 0.5 microgram/mL of alpha-amanitin. Therefore, the nuclear capping enzyme is resistant to this drug. Our results indicate that RNA polymerase II primary transcripts are the substrate for the cellular capping enzyme and that the beta phosphate in the pyrophosphate bridge (m7GgammapbetapalphapXm) is derived from the 5' ends of the RNA chains.
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PMID:Methylation and capping of RNA polymerase II primary transcripts by HeLa nuclear homogenates. 62 55

Initiation of adenovirus transcription was analyzed by incubation of isolated nuclei from virus infected cells in the presence of beta-32P GTP or beta-32P ATP. Nucleotide analysis of RNA from nuclei incubated with beta-32P GTP shows that the label is incorporated exclusively into pppGp and ppGp. Under similar incubation conditions, the label from beta-32P ATP was incorporated primarily into the 5' phosphate of 5', 3' mononucleoside diphosphates, but label was also detected in pppAp, pppGp and in the 3' nucleoside monophosphates. Analysis of RNA, synthesized in the presence of different concentrations of alpha-amanitin, shows that only RNA polymerase III initiates virus specific transcription in isolated nuclei. The virus specific transcripts containing pppAp and pppGp in their 5' termini were identified as the 5.5S and 5.2S viral RNA species by hybridization and finger printing.
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PMID:Initiation of transcription in nuclei isolated from adenovirus infected cells. 64 9


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