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

---

Query: EC:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

U6 small nuclear RNA (snRNA), an essential component of the eukaryotic spliceosomes, is unique in that it is synthesized by RNA polymerase III, while all other U-snRNAs are synthesized by RNA polymerase II. U6 genes are notable for functional upstream regulatory elements which resemble RNA polymerase II regulatory sequence motifs. In this study, the optimal conditions for transcription of the U6 snRNA gene in vitro were found to be similar to conditions optimal for transcription of 5S RNA genes. To purify the trans-acting factors necessary for the transcription of the U6 RNA gene, HeLa cell extracts were fractionated on a DEAE-Sephadex column, and three fractions, designated DE-50, DE-175, and DE-500, were obtained by stepwise elution with 50, 175, and 500 mM ammonium sulfate, respectively. DE-175 fraction transcribed tRNA and 5S RNA genes but not a mouse U6 RNA gene. Complementation of the DE-175 fraction with the DE-50 fraction resulted in the transcription of the U6 RNA gene. Experiments in which the transcription factor (TFIIIA) was selectively inactivated indicated that TFIIIA is not required for the transcription of the U6 RNA gene. These results show that the U6 snRNA gene, although transcribed by RNA polymerase III, differs from tRNA and 5S RNA genes in that factors other than TFIIIA, -IIIB, and -IIIC are required for U6 gene transcription in vitro.
...
PMID:Transcription of a U6 small nuclear RNA gene in vitro. Transcription of a mouse U6 small nuclear RNA gene in vitro by RNA polymerase III is dependent on transcription factor(s) different from transcription factors IIIA, IIIB, and IIIC. 318 77

Three subspecies of RNA polymerase II, designated IIO, IIA, and IIB, have been described in calf thymus and shown to differ in the apparent molecular weight of their largest subunits, designated IIo, IIa, and IIb, respectively. The objective of this study was to develop a procedure for the purification of RNA polymerase IIO. This form of the enzyme predominates in vivo and is responsible for the transcription of most cellular genes. RNA polymerase II is solubilized from isolated calf thymus nuclei in the presence of high concentrations of chelators, precipitated with polyethyleneimine, extracted with salt, and precipitated with (NH4)2SO4. The solubilized enzyme is resolved from factors that destabilize RNA polymerase IIO by chromatography on heparin-Sepharose CL-4B and DE52. RNA polymerase IIO is then partially resolved from RNA polymerases IIA and IIB by chromatography on DEAE-5PW and further purified by chromatography on Phenyl-Superose and Mono Q. RNA polymerase IIO was purified 1000-fold from the polyethyleneimine eluate resulting in about 130 micrograms of RNA polymerase IIO from 300 g of calf thymus. The specific activity of RNA polymerase IIO, in nonselective assays using calf thymus DNA as template, is 440 units/mg and not significantly different from that of RNA polymerases IIA and IIB. The similar transcriptional activities in nonselective assays suggest that the C-terminal domain of the largest RNA polymerase II subunit does not play a major role in the elongation phase of the reaction when deproteinized DNA serves as template. The small subunits of RNA polymerase IIO are indistinguishable from those of RNA polymerases IIA and IIB.
...
PMID:Purification of RNA polymerase IIO from calf thymus. 319 3

Experimental conditions are reported under which purified spinach chloroplast RNA polymerase catalyses the abortive elongation reaction on a synthetic poly[d(A-T)] template. The reaction only occurs under very stringent conditions and absolutely requires Mn2+ as the metal activator. No reaction can be detected in the presence of Mg2+. Furthermore, the rate of abortive elongation with the chloroplast enzyme is extremely sensitive to the presence of added salts, such as KCl or (NH4)2SO4, in the reaction assays. In the combined presence of Mn2+ and Mg2+, a marked inhibition of abortive elongation is associated with an activation of productive elongation and an increased length of RNA chains. Thus, whereas Mn2+ is more active than Mg2+ for phosphodiester bond formation, it appears that Mg2+ favors the stabilization of the ternary transcription complexes. These results are compared with those obtained under similar conditions for wheat germ RNA polymerase II and Escherichia coli RNA polymerase.
...
PMID:Abortive and productive elongation catalysed by purified spinach chloroplast RNA polymerase. 329 91

A DNA-dependent RNA polymerase has been isolated and characterized from the parasitic flagellated protozoan Leishmania mexicana. The initial stages of purification utilized high-ionic-strength extraction and protamine sulfate treatment. The enzyme was further purified by differential elution by heparin-Sepharose, DEAE-Sephadex, and carboxymethyl-Sephadex chromatography. Analysis of the chromatographically purified RNA polymerase on nondenaturing gels revealed two electrophoretic forms. The enzyme isolated had characteristics of true DNA-dependent RNA polymerase since it required DNA and all four nucleoside triphosphates for synthesis of RNase-sensitive products. Analysis of ammonium sulfate and metal ion optima, as well as relative activities of the enzyme with Mn2+ versus Mg2+, gave results similar to those reported for other RNA polymerase IIIs in eucaryotes. Formycin A triphosphate was found to be a noncompetitive inhibitor of RNA polymerase III, and cordycepin triphosphate was found to be inhibitory, although the exact mode of inhibition was not determined.
...
PMID:Isolation and characterization of DNA-dependent RNA polymerase III from Leishmania mexicana and inhibition by purine analogs. 343 22

The location of the repressor gene, blaI, for the beta-lactamase gene blaP of Bacillus licheniformis 749, on the 5' side of blaP, was confirmed by sequencing the bla region of the constitutive mutant 749/C. An amber stop codon, likely to result in a nonfunctional truncated repressor, was found at codon 32 of the 128 codon blaI open reading frame (ORF) located 5' to blaP. In order to study the DNA binding activity of the repressor, the structural gene for blaI, from strain 749, with its ribosome binding site was expressed using a two plasmid T7 RNA polymerase/promotor system (S. Tabor and C. C. Richardson. Proc. Natl. Acad. Sci. 82, 1074-1078 (1985). Heat induction of this system in Escherichia coli K38 resulted in the production of BlaI as 5-10% of the soluble cell protein. Repressor protein was then purified by ammonium sulfate fractionation and cation exchange chromatography. The sequence of the N-terminal 28 amino acid residues was determined and was as predicted from the DNA. Binding of BlaI to DNA was detected by the slower migration of protein DNA complexes during polyacrylamide gel electrophoresis. BlaI was shown to selectively bind DNA fragments carrying the promoter regions of blaI and blaP.
...
PMID:Purification and DNA binding properties of the blaI gene product, repressor for the beta-lactamase gene, blaP, of Bacillus licheniformis. 349 48

Nuclear extract from Morris hepatoma 3924A was fractionated by DEAE-Sephadex chromatography. The fraction eluting with 300 mM (NH4)2SO4 (DE-C) was used for transcribing cloned mouse metallothionein-I (MT-I) gene in a run-off assay. This fraction contained the majority of RNA polymerase II as well as the transcription factor(s). Accuracy of MT-I DNA transcription was confirmed by S1 nuclease mapping. Low concentrations (1 microgram/ml) of alpha-amanitin inhibited the reaction, indicating that RNA polymerase II directed the transcription. Unfractionated nuclear extracts from the hepatoma or a rat mammary adenocarcinoma as well as whole cell extract obtained from the mammary tumor also transcribed MT-I gene. The extent of transcriptional activity was in the following order: hepatoma nuclear fraction DE-C greater than whole cell extract derived from rat mammary adenocarcinoma cells greater than nuclear extract derived from rat hepatoma or rat mammary adenocarcinoma cells. These studies have demonstrated that a fractionated nuclear extract obtained from a tissue supports efficient and accurate RNA polymerase II-mediated transcription of MT-I DNA.
...
PMID:Accurate transcription of mouse metallothionein-I gene in a fractionated nuclear extract from a rat hepatoma. 355

The small nuclear RNAs U1, U2, U4, and U5 are cofactors in mRNA splicing and, like the pre-mRNAs with which they interact, are transcribed by RNA polymerase II. Also like mRNAs, mature U1 and U2 RNAs are generated by 3' processing of their primary transcripts. In this study we have investigated the in vitro processing of an SP6-transcribed human U2 RNA precursor, the 3' end of which matches that of authentic human U2 RNA precursor molecules. Although the SP6-U2 RNA precursor was efficiently processed in an ammonium sulfate-fractionated HeLa cytoplasmic S100 extract, the product RNA was unstable. Further purification of the processing activity on glycerol gradients resolved a 7S activity that nonspecifically cleaved all RNAs tested and a 15S activity that efficiently processed the 3' end of pre-U2 RNA. The 15S activity did not process the 3' end of a tRNA precursor molecule. As demonstrated by RNase protection, the processed 3' end of the SP6-U2 RNA maps to the same nucleotides as does mature HeLa U2 RNA.
...
PMID:Accurate and efficient 3' processing of U2 small nuclear RNA precursor in a fractionated cytoplasmic extract. 367 Mar 7

RNA polymerase type II from human term placenta has been isolated and characterized with respect to its template, ammonium sulfate, divalent cation, and buffer preferences. In addition, the apparent Michaelis constants for AMP and UMP incorporation have been determined. The enzyme was also analyzed by native and denaturing polyacrylamide gel electrophoresis, and evidence is presented that a single polypeptide is radiolabeled with azido purine nucleoside triphosphate photoprobes.
...
PMID:Characterization of RNA polymerase type II from human term placenta. 375 75

Nuclear DNA-dependent RNA polymerases I, II and III were purified from kidney, liver and spleen from Swiss mice (Mus musculis) and from seven transplantable murine tumors. In the presence of the optimal concentration of (NH4)2SO4 for each polymerase, 1-8 mM spermidine or spermine stimulated most polymerases several fold, and generally, enzyme I was stimulated more than either enzyme II or III. Spermine was more efficacious than spermidine as a stimulant of polymerase activity except for polymerase III from three tumors. Tumor polymerases I (or II) and the corresponding normal tissue enzymes responded similarly to the polyamines. Stimulation of a RNA polymerase by a polyamine could not be correlated with the growth rate of the tissues of polymerase origin or with the tissue's RNA polymerase or RNA synthetic activities.
...
PMID:Comparison of the effects of polyamines on the activities of RNA polymerases from murine normal tissues and transplantable tumors. 381 57

Nuclear extracts obtained from normal rat liver and from Morris hepatoma 3924A were fractionated by DEAE-Sephadex chromatography. The fraction eluted with 175 mM (NH4)2SO4 (DE-B), which contains greater than 90% of RNA polymerase I activity, supported accurate transcription of cloned rat rDNA. A similar fraction obtained from the cytosol had all of the factors required for rDNA transcription. However, its transcriptional activity was at most one-sixth that of the corresponding nuclear fraction, as determined by the amount of protein needed to produce a similar quantity of the transcript. Unfractionated nuclear or cytosol preparations did not yield an accurate transcript. Optimal KCl and magnesium concentrations for rDNA transcription were 60 mM and 5-7.5 mM, respectively. The extent of transcriptional activity was in the following order: hepatoma nuclear fraction DE-B greater than whole cell extract derived from rat mammary adenocarcinoma cells much greater than normal liver fraction DE-B. The hepatoma preparation produced at least 10 times the amount of transcript produced by the corresponding liver nuclear preparation. Transcriptional activity was proportional to the levels of RNA polymerase I and to the rate of rRNA synthesis in these tissues.
...
PMID:Accurate initiation of rat ribosomal RNA gene transcription using a fractionated nuclear extract from normal liver and a hepatoma. 385 47


<< Previous 1 2 3 4 5 6 7 8 9 10

---