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)

Although many researchers have reported that RNA synthesis in the ovary is enhanced by gonadotropin treatment, there are only a few papers concerning the character of newly synthesized RNA after gonadotropin treatment. In this paper, the RNA synthesized in the ovary of immature rats after HCG treatment was qualitatively studied. Immature female Sprague-Dawley rats were administered with 0.3 mc per rat of 3H-uridine at a certain time interval after injection of HCG (10 iu/rat) and the ovaries were subsequently isolated after 15, 30 or 60 minutes. RNA was extracted from the homogenate of the ovaries according to the hot phenol method after Scherrer and Darnell. The 3H-RNA thus extracted was treated with electrophoretically purified DNase to break down and remove DNA that mingled with it. The RNA solution ultimately obtained was analysed on a 3-20% sucrose gradient. The different fractions thus separated were then subjected to measurement of radioactivity and optical density at 260 mmug. The RNA extracted from the ovary of immature untreated rat labeled with 3H-uridine for 15 minutes showed a flat pattern of radioactivity from the top to the bottom fractions with low radioactivity. Otherwise, when labeled for one hour, the RNA showed a pattern of radioactivity like those of optical density at 260 mumu with peaks of r-RNAs and t-RNA. When the ovary was pulse-labeled with 3H-uridine for 15 minutes starting 2 hours after injection of HCG, the RNA with a large S value was synthesized and the pattern of variation in radioactivity was that of rising near the bottom fraction and declining with access to the top fraction. The results obtained by labeling for 15 minutes starting 40 hours after PMS administration were similar to those obtained in immature untreated rats. The patterns of radioactivity in RNA obtained by the labeling for 15 minutes starting 2 hours after HCG and 42 hours after PMS were similar to those starting 2 hours after only HCG injection. The patterns of radioactivity became similar to those of optical density at 260 mmu, when the ovaries were labeled for 30 or 60 minutes. From these results, it was suggested that the newly synthesized RNA 2 hours after HCG was constructed from m-RMA with rapid turn over and precursors of r-RNAs and t-RNA. This RNA synthesis was blocked by pretreatment with actinomycin but not by cycloheximide. From these results, it was suggested that enhancement in RNA polymerase activity or change in template capacity of DNA which would have an effect on RNA synthesis was not based on newly synthesized protein.
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PMID:[Studies on the RNA synthesized in the ovary of immature rats after HCG administration (author's transl)]. 5 Sep 55

Recent findings have confirmed the role of form A DNA-dependent polymerase activity as that which is responsible for the transcription of the ribosomal RNA-coding genes. Unfortunately, the form A enzymes have proved to be very labile and difficult to work with, especially under high ionic strength conditions. We have, therefore, investigated a method for the purification of the form AI and AII enzymes from rat liver using mild low-ionic-strength conditions. Since preparations from whole nuclei were found to be grossly contaminated with protein having similar properties, the enzymes are extracted from nucleoli. Forms AI and AII are separated on a phosphocellulose column, purified by further ion-exchange chromatography, and by sedimentation through a glycerol gradient. The purified enzymes each migrate as a single band on native polyacrylamide gels and have the expected characteristics of form A RNA polymerase. Sedimentation rates through glycerol gradients indicate that they both have a similar size to that of Escherichia coli RNA polymerase (Mr about 500,000). The purified enzymes are free of DNase and RNase. A method is also described for the purification of form B from the nucleoplasm remaining after isolation of nucleoli. The presence of form C activity was not detected.
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PMID:Purification of form AI and AII DNA-dependent RNA polymerases from rat-liver nucleoli using low-ionic-strength extraction conditions. 5 56

The binding characteristics of [125I]-labeled L-triiodothyronine (T3) to chromatin isolated from rat liver nuclei were investigated. Binding of T3 to chromatin showed temperature-, incubation time-, and DNA concentration-dependence. According to Scatchard analysis, the apparent equilibrium dissociation constant was 225 pM, with a maximum binding capacity of about 0.2 pmoles per mg DNA. Displacement studies with unlabeled thyroxine (T4) and T3 showed that the binding sites for T4 might be the same as T3 but the binding affinity of the former was less than that of the latter. The binding was completely inhibited by the eukaryotic RNA polymerase inhibitor, rifampicin AF/021, but not by the prokaryotic inhibitor, rifampicin and alpha-amanitin. These observations indicate that the receptors for T3 have certain properties in common with RNA polymerase or other enzyme proteins which are sensitive to the rifampicin derivative. The hormone--chromatin fragments complex was solubilized from residual chromatin by digestion with DNase, but not with RNase, suggesting that the T3 receptors localize in the DNase-sensitive regions of DNA in the chromatin. This provides a useful method to use to investigate the localization of the receptor proteins in the chromatin and the interaction of the hormone-receptor complex with DNA.
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PMID:Binding characteristics of L-triiodothyronine to isolated rat liver chromatin. 19 15

Cyclic AMP (cAMP) and its receptor protein (CRP) have a dual role in the regulation of the two promoters that control the galactose (gal) operon of Escherichia coli. One promoter, P1, requires cAMP-CRP for activity; the other, P2, is inhibited by these factors. We have examined the interactions site of cAMP-CRP on gal DNA by using two types of protection experiments, involving DNase digestion and methylation by dimethyl sulfate. Our results indicate that cAMP-CRP binds to gal DNA in a segment located between 50 and 24 base pairs preceding the P1 start point for transcription. Although the location of the cAMP-CRP interaction site is clearly different in gal and lac DNA, comparison of the DNA sequences suggests a similar recognition sequence. The location of the cAMP . CRP-binding site in gal further suggests that protein-protein interactions between RNA polymerase and cAMP . CRP play an important role in transcription initiation at the gal and possibly other cAMP-dependent promoters.
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PMID:Interaction site of Escherichia coli cyclic AMP receptor protein on DNA of galactose operon promoters. 22 78

Double-stranded RNA of some virus genomes can be used as template for the DNA-dependent RNA polymerase purified from Escherichia coli. The RNA synthesis requires all four nucleoside triphosphates and manganese ions and is dependent on the presence of sigma subunit. The reaction is inhibited by rifampicin, streptolydigin and ethidium bromide, but not by DNase and actinomycin D which does not bind to double-stranded RNA. The template activity of double-stranded RNA from various viruses is different in each case. The order of template efficiency is Penicillum chrysogenum virus greater than cytoplasmic polyhedrosis virus greater than rice dwarf virus greater than reovirus. The product obtained using cytoplasmic polyhedrosis virus double-stranded RNA as template is single-stranded and hybridizes specifically to the denatured template RNA. One of the major 5'-starting nucleotide sequences of the product RNA is pppA-A-Y--. These results indicate that transcription in vitro of double-stranded RNA by E. Coli RNA polymerase is initiated at specific sites on the template.
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PMID:Transcription of double-stranded RNA by Escherichia coli DNA-dependent RNA polymerase. 32 6

The dimethylsulphate method has been used to study the complexes of RNA polymerase (Escherichia coli) with DNA of T7 phage, poly[d(A--T)] and fragments of calf thymus DNA protected against DNase digestion by RNA polymerase. The binding of RNA polymerase to DNA significantly increases the formation of 1-methyl-adenine produced by methylation of the single-stranded DNA region, diminishes by about 10% the formation of 3-methyl-adenine by methylation within the minor groove and does not affect the formation of 7-methyl-guanine by methylation within the major DNA groove. The presence of nascent RNA decreases the formation of 1-methyl-adenine in DNA of the complex by about 30%. The initiation of RNA synthesis or RNA synthesis itself does not influence the methylation of the major groove but shielding of the minor groove increases by about twice as much. These results suggest that RNA polymerase, upon binding, breaks Watson-Crick base-pairing in a DNA region of about 15-base-pairs long, that nascent RNA forms a duplex with DNA of about 10-base-pairs long; and that the enzyme weakly interacts with DNA along its grooves and preferentially makes contacts with the minor groove.
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PMID:A study of unwinding of DNA and shielding of the DNA grooves by RNA polymerase by using methylation with dimethylsulphate. 34 67

T7 gene 6 exonuclease has been shown to have an RNase H activity as well as a double-strand specific DNase activity by the following experiments: The RNase H activity coelutes with the DNase activity from DEAE-cellulose, phosphocellulose, hydroxyapatite, and Sephadex G-200 columns. Gene 6 exonuclease specified by a T7 strain with a temperature sensitive mutation in gene 6 has an extremely heat-labile RNase H activity as well as a heat-labile DNase activity. T7 gene 6 exonuclease degrades the RNA region of a poly(A) . poly(dT) hybrid polymer exonucleolytically from the 5' terminus, releasing a ribonucleoside 5'-monophosphate product. When the RNA strand of a 0X174 RNA . DNA hybrid molecule synthesized with E. coli RNA polymerase is degraded, a ribonucleoside triphosphate is produced from the 5'-triphosphate terminus. Participation of T7 gene 6 exonuclease in the removal of primer RNA in discontinuous replication of T7 DNA is discussed.
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PMID:T7 gene 6 exonuclease has an RNase H activity. 36 24

DNA-dependent RNA polymerase class C (or III) has been solubilized from either uninfected or adenovirus-2-infected HeLa cells and purified by chromatography on phosphocellulose, DNA-cellulose, CM-Sephadex and DEAE-Sephadex. The last column separated the enzyme into three forms CI, CII and CIII, which were completely free of RNA polymerases class A and B and of DNase and RNase. The total and the relative amount of these different enzyme C forms did not vary whether purified from uninfected or infected cells. Irrespective of the stage of purification, the three enzyme forms transcribed deproteinized adenovirus-2DNA very efficiently. This transcription was highly sensitive to elevated ionic strength (especially in the presence of Mg2+) and was accompanied by continuous reinitiation as shown by adding poly(rI), a potent inhibitor of initiation. In addition heparin-resistant initiation complexes could be formed at elevated temperature. The RNA synthesized in vitro on deproteinized intact adenovirus-2 DNA by the different forms of RNA polymerase class C, has been characterized. Analysis of the transcripts by gel electrophoresis, RNA self-annealing, hybridization to separated adenovirus-2 DNA strands and to restriction endonuclease (BamHI, HindIII), adenovirus-2 DNA fragments have demonstrated that restriction endonuclease (BamHI, HindIII), adenovirus-2 DNA fragments have demonstrated that the various regions of the adenovirus-2 genome were randomly transcribed. In addition, hybridization of RNA transcripts labelled at their 5' end by either [gamma32P]ATP or [gamma-32P]GTP indicated that not only elongation but also initiation occurred randomly through the entire adenovirus-2 genome, irrespective of the form of the enzyme and of the origin of the cells (normal or infected). The results are discussed in terms of the components which are possibly involved in specific transcription.
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PMID:Transcription in vitro of adenovirus-2 DNA by RNA polymerases class C purified from uninfected and adenovirus-infected HeLa cells. 71 Apr 51

Methods developed for isolating bacterial nucleoids were applied to bacteria infected with phage T4. The replicating pool of T4 DNA was isolated as a particle composed of condensed T4 DNA and certain RNA and protein components of the cell. The particles have a narrow sedimentation profile (weight-average s=2,500S) and have, on average, a T4 DNA content similar to that of the infected cell. Their dimensions observed via electron and fluorescence microscopy are similar to the dimensions of the intracellular DNA pool. The DNA packaging density is less than that of the isolated bacterial nucleoid but appears to be roughly similar to its state in vivo. Host-cell proteins and T4-specific proteins bound to the DNA were characterized by electrophoresis on polyacrylamide gels. The major host proteins are the RNA polymerase subunits and two envelope proteins (molecular weights, 36,000 and 31,000). Other major proteins of the host cell were absent or barely detectable. Single-strand breaks can be introduced into the DNA with gamma radiation or DNase without affecting its sedimentation rate. This and other studies of the effects of intercalated ethidium molecules have suggested that the average superhelical density of the condensed DNA is small. However, these studies also indicated that there may be a few domains in the DNA that become positively supercoiled in the presence of high concentrations of ethidium bromide. In contrast to the Escherichia coli nucleoid, the T4 DNA structure remains condensed after the RNA and protein components have been removed (although there may be slight relaxation in the state of condensation under these conditions).
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PMID:Properties of condensed bacteriophage T4 DNA isolated from Escherichia coli infected with bacteriophage T4. 78 57

Although the bulk of RNA synthesized in vitro by vaccinia virus is 8 to 12 S, a small amount of high molecular weight RNA can be detected. This RNA is virion-associated and is not extruded from the virus as high molecular weight RNA. It is sensitive to pancreatic RNase digestion in high salt, has a density in neutral CS2SO4 of 1.68 g ml-1 and remains large after digestion with DNase or denaturation in dimethyl sulfoxide. In the presence of high concentrations of virus in the in vitro RNA polymerase reaction, pulse-labeling experiments indicate an RNA sedimenting heterogeneously between 20 and 30 S. Pulse-chase experiments indicate that a fraction of this high molecular weight RNA can be chased into RNA sedimenting at 8 to 12 S. Cleavage into smaller fragments is not dependent on continued RNA synthesis but does require ribonucleoside triphosphates. In the presence of ethidium bromide, the RNA is not cleaved.
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PMID:In vitro synthesis of a high molecular weight virion-associated RNA by vaccinia. 83 1


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