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)

Chromatin-bound and poly[d(A-T)]dependent RNA polymerase I plus III and II activities of mouse liver were analysed 24 and 48 hr after partial hepatectomy. Chromatin-bound RNA polymerase I plus III activity showed an increase of 57% at 24 hr and 51% at 48 hr after partial hepatectomy. There was a decrease in chromatin-bound RNA polymerase II activity of 15% at 24 hr and 34% at 48 hr after partial hepatectomy. There was no significant changes in poly[d(A-T)]dependent RNA polymerase activities. Heparin caused an approximately 10-fold increase in chromatin-bound RNA polymerase II activity. The stimulation by heparin was significantly increased 48 h after partial hepatectomy. Anaesthesia and/or surgery had great influence on RNA polymerase activities. At 24 hr after operation, chromatin-bound RNA polymerase I plus III and II activities were depressed, and the liver cell chromatin was more susceptible to stimulation by heparin.
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PMID:Effects of partial hepatectomy on RNA polymerase activities in mouse liver. 669 89

The function of lysyl residues of the sigma subunit of the RNA polymerase from Escherichia coli was investigated by chemical modification with trinitrobenzenesulfonic acid (TNBS). Following reaction with TNBS, analysis of the modified sigma indicated that trinitrophenylation was limited to the epsilon-amino groups of lysyl residues. Progressive loss in the activity of sigma followed increasing trinitrophenylation as assayed by the ability to stimulate RNA polymerase core enzyme in a reaction directed by T7 DNA. Modification of five lysyl groups resulted in the complete loss of sigma activity. Kinetic analysis indicated that one lysyl group is critical for the function of sigma. TNP-sigma was able to form a holoenzyme complex with a binding affinity comparable to that of sigma. Promoter recognition studies were done by using HindIII fragments from T5 DNA. The TNP-sigma core complex was unable to form a tight binary complex with the T5 promoters. Studies on RNA chain initiation were carried out by using d(A-T)n and T7 DNA templates. TNP-sigma was unable to stimulate RNA chain initiation by core polymerase. Limited proteolytic digests of TNP-sigma or sigma using Staphylococcus aureus V8 protease were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The results suggested a change in the conformation of sigma following trinitrophenylation.
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PMID:Effect of lysine modification on the activity of the sigma subunit of Escherichia coli RNA polymerase. 681 85

Our previous work [Levinger, L. & Varshavsky, A. (1982) Cell 28, 375-385] has shown that D1, a 50-kilodalton chromosomal protein of Drosophila melanogaster, is specifically associated with isolated nucleosomes that contain a complex A + T-rich satellite DNA with buoyant density of 1.688 g/ml. We show here that D1 is also a component of nucleosomes containing a simple-sequence, pure A + T satellite DNA, buoyant density 1.672 g/ml. Furthermore, using a modification of a protein blotting technique in which proteins are not exposed to dodecyl sulfate denaturation, we have found that D1 preferentially binds to A + T-rich double-stranded DNA in vitro, and it is apparently the only abundant nuclear protein in cultured D. melanogaster cells that possesses this property. Synthetic poly[d(A-T)].poly[d(A-T)] and poly(dA).poly(dT) duplexes effectively compete in vitro with A + T-rich D. melanogaster satellite DNAs for binding to D1, whereas total Escherichia coli DNA is an extremely poor competitor. These findings strongly suggest that D1 is a specific component of A + T-rich, tandemly repeated, heterochromatic regions, which constitute up to 15-20% of the total D. melanogaster genome. Possible functions of D1 protein include compaction of A + T-rich heterochromatin and participation in microtubule-centromere interactions in mitosis. In addition, D1 may prevent nonspecific binding to A + T-rich satellite DNA of other nuclear proteins that have a preference for AT-DNA, such as RNA polymerase or regulatory proteins, and may also participate in the higher-order chromatin organization outside tandemly repetitive regions by binding to nonrandomly positioned stretches of A + T-rich DNA.
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PMID:Protein D1 preferentially binds A + T-rich DNA in vitro and is a component of Drosophila melanogaster nucleosomes containing A + T-rich satellite DNA. 681 40

Three transcription units are present in the adenovirus type 2 region EII. Transcription units EIIaE and EIIaL encode the mRNA for the 72,000-dalton DNA binding protein, early and late in the lytic cycle, respectively, and transcription unit EIIb encodes the mRNA for the protein that binds to the 5' termini of adenovirus DNA. By using a cell-free transcription system in the presence of purified RNA polymerase B (or II), we have obtained specific initiation of transcription from both the EIIaE promoter, which does not contain a T-A-T-A box, and the EIIaL promoter, which does. In addition, we have identified a new EII T-A-T-A box promoter that is located close to the early non-T-A-T-A box promoter and is used both in vivo and in vitro.
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PMID:Specific in vitro initiation of transcription on the adenovirus type 2 early and late EII transcription units. 695 Mar 83

Sea urchin (psammechinus miliaris) H2A histone genes shown to be promoter mutants from oocyte injection experiments were tested for their ability to initiate transcription in vitro. Circular templates were transcribed with HeLa cell extracts, and the transcripts were assayed by mung bean or S1 nuclease mapping of the 5' ends. The transcripts of the H2A mutants produced in vitro were qualitatively similar and, in most cases, identical to those seen in oocyte injection experiments, but quite large quantitative differences were observed for some H2A mutant genes. Both the T-A-T-A box and far upstream sequences residing in the modulator segment E [Grosschedl, R. & Birnstiel, M. L. (1980) Proc. Natl. Acad. Sci. USA 77, 7102--7106] were found to be essential for maximal transcription in vitro. Deletion of either of these sequence elements reduced transcription to 20%. A similar reduction in the amount of H2a transcripts was found when a T-A-T-A-to-T-A-G-A point mutant was tested in vitro. Essential far upstream sequences were mapped between nucleotides -139 and -111, 5' to the initiation site of transcription. In the standard run-off transcription test using restriction fragments, the effects of these sequences could be mimicked by free DNA ends, suggesting that the function of this in vitro upstream sequence might be to provide an entry side for RNA polymerase II.
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PMID:Delimitation of far upstream sequences required for maximal in vitro transcription of an H2A histone gene. 695 85

The mechanism of streptolydigin inhibition of RNA synthesis has been investigated with a combination of steady state kinetics and product analysis by employing the abortive initiation reaction of Escherichia coli RNA polymerase. The pattern of inhibition by streptolydigin on the poly[d(A-T)] . poly[d(A-T)]template (non-competitive versus AMP; competitive versus UTP) was consistent with one inhibitor binding site and with an ordered addition of AMP followed by UTP. The more complicated patterns observed on the poly[d(I-C)] . poly[d(I-C)] template and the bacteriophage T7 A2 promotor (noncompetitive versus CTP) were explained by assuming that streptolydigin could stabilize the translocated ternary complex containing the product dinucleotide. Product analysis of two other abortive initiation reactions showed that the product did not dissociate from the inhibitor-bound translocated ternary complex. Finally, rifampicin and streptolydigin were shown to be functionally independent during initiation on several templates.
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PMID:On the mechanism of streptolydigin inhibition of Escherichia coli RNA polymerase. 698 76

Open complexes of Escherichia coli RNA polymerase core enzyme with a poly[d(A-T)]-poly[d(A-T)]template have been characterized and compared with the previously characterized holoenzyme open complexes on the same template (Hansen, U. M., and McClure, W. R. (1979) J. Biol. Chem. 254, 5713-5717). The open complexes were monitored by the abortive initiation synthesis of the dinucleotide pApU, which is catalyzed by both enzymes. The major differences between the two complexes were: 1) the Michaelis constant for UTP was 60 times higher for core enzyme than for holoenzyme, 2) the intrinsic binding constant of core enzyme to the DNA was 3 orders of magnitude less than that of holoenzyme, and 3) cooperative binding of 2 core units was required for activity. Thus, the presence of the sigma subunit significantly altered the nature and extent of open complex formation. The rate of formation of the open complexes, however, was rapid for both enzymes. Rifampicin is shown to have a slight stimulatory effect on the extent of open complex formation of the core enzyme.
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PMID:Role of the sigma subunit of Escherichia coli RNA polymerase in initiation. I. Characterization of core enzyme open complexes. 700 Jul 58

The effect of the regulatory nucleotide ppGpp on transcription by Escherichia coli RNA polymerase in vitro has been studied using bacteriophage T7 and T3 DNAs as templates. We have previously described the development of transcriptional test systems using these templates that can sensitively detect changes in promotor or terminator recognition of an RNA polymerase (Wiggs, J.L., Bush, J.W., and Chamberlin, M.J. (1979) Cell 16, 97-109) or changes in the rate of any of the major steps of the transcription cycle (Chamberlin, M.J., Nierman, W.C., Wiggs, J.L., and Neff, N. (1979) J. Biol. Chem. 254, 10061-10069). Using these procedures we fail to detect any substantial alteration by ppGpp of the normal interaction of E. coli RNA polymerase with the several T7 major and minor promoter sites or of the rate of productive RNA chain initiation at either T7 promoter A1 (ATP start) or A2 (GTP start). However, at physiologically relevant concentrations (KI approximately 50 microM), ppGpp significantly lowers the overall elongation rate of T7 RNA chains, leading to a substantial reduction in the overall rate of RNA synthesis. Inhibition of transcriptional elongation does not appear to be competitive with the ribonucleoside triphosphate substrates. Furthermore, ppGpp does not inhibit chain elongation during transcription of the synthetic polynucleotide templates poly[d(A-T)] or poly[d(A-G):d(C-T)]. We conclude that ppGpp interacts directly with some site on RNA polymerase other than one of the sites used for substrate binding. Furthermore, the inhibition must depend on specific DNA sequences present in T7 DNA, but not in poly[d(A-T)] or poly[d(A-G):d(C-T)]. Analysis of T7 transcripts formed in the presence and absence of ppGpp by gel electrophoresis reveals that in the latter instance there is enhanced pausing of the transcriptional elongation complex at specific sites on the template. It is likely that the enzyme also pauses at such sequences in the absence of ppGpp, but for a far briefer time (Kassavetis, G.A., and Chamberlin, M.J. (1981) J. Biol. Chem. 256, 2777-2786). Thus, ppGpp appears to slow transcriptional elongation by binding to RNA polymerase and altering its structure in a manner that impedes passage of the enzyme through certain DNA sequences. The presence of similar transcriptional barriers in rRNA operons activated by the presence of ppGpp could lead selectively to large reductions in the rate of rRNA synthesis in vivo.
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PMID:A direct effect of guanosine tetraphosphate on pausing of Escherichia coli RNA polymerase during RNA chain elongation. 700 98

To characterize the interactions of RNA polymerase with DNA, we have investigated the thermal transition of poly[d(A-T] bound to RNA polymerase from Escherichia coli and the aggregation properties of the enzyme with DNA. The melting curve of the DNA-enzyme complex demonstrates a sharply lowered melting temperature for part of the DNA, whereas for another fraction the double helix is stabilized. This indicates that the DNA-binding site of RNA polymerase serves two functions: (1) to disrupt the double helix at one point, and (2) to maintain the duplex form at other points. The aggregation of DNA and RNA polymerase has been monitored by turbidity measurements, and conditions have been delineated under which aggregation is minimized. Holoenzyme added to double-stranded DNA or single-stranded DNA has little or no tendency to aggregate under most conditions. Core enzyme, on the other hand, aggregate extensively with double-stranded DNA, the only under conditions of low salt (10 mM KCl), without Mg2+, or at high salt (300 mM KCl), with or without Mg2+, can this aggregation be eliminated. Core enzyme also does not aggregate in the presence of single-stranded DNA. These aggregation properties are interpreted as evidence for more than one DNA-binding site on RNA polymerase.
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PMID:The interaction of RNA polymerase and DNA. Effects on the helix-coil transition and light scattering. 701 99

The azo dye Congo Red has a high affinity for nucleotide-binding enzymes. We have studied the binding of Congo Red to RNA polymerase by circular dichroism (CD) and difference absorption spectroscopy, steady-state kinetics, and nitrocellulose filter-binding. Induced CD shows that a large number of Congo Red molecules bind to the holoenzyme. CD also demonstrates that the core enzyme at low ionic strengths has a distinctive Congo Red binding site which is not present in the holoenzyme, nor in the core enzyme at higher ionic strengths or in the presence of poly(dT). CD studies indicate that Congo Red can readily displace double-stranded polynucleotides (T7 DNA or poly[d(A-T)] from RNA polymerase. Single-stranded DNA (poly(dT) and T7 DNA in open complexes) is not displaced from RNA polymerase except at high Congo Red concentrations. Both kinetics and nitrocellulose filter-binding measurements support this conclusion. Difference spectra indicate that the bound Congo Red molecules undergo stacking. We postulate that RNA polymerase binds Congo Red in a region with which a segment of DNA normally interacts, and that Congo Red is a potent inhibitor because the stacked dye has a polyanionic character.
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PMID:Spectroscopic studies of Congo Red binding to RNA polymerase. 702 Jul 64


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