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)

A cytoplasmic particulate fraction from human leukemic cells has been shown to contain reverse transcriptase and its associated high-molecular weight RHA template. We attempted to detect the reverse-transcriptase-template complex in morphologically normal peripheral blood leukocytes from patients with acute leukemia in complete remission. Our assay system consisted of a velocity glycerol gradient and cesium sulfate equilibrium gradient analysis of the endogenous reverse transcriptase reaction product. Three of nine patients in remission had positive reactions determined by glycerol gradient analysis, and eight of 10 patients in remission had positive reactions by cesium sulfate gradient analysis. We were unable to detect the template complex in leukocytes of normal persons. Thus, normal-appearing leukocytes in the peripheral blood of some leukemia patients in remission seem to retain a number of biochemical characteristics, possibly viral related, associated with leukemic cells.
...
PMID:Reverse transcriptase in leukocytes of leukemic patients in remission. 5 87

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.
...
PMID:Purification of form AI and AII DNA-dependent RNA polymerases from rat-liver nucleoli using low-ionic-strength extraction conditions. 5 56

DNA-dependent RNA polymerase B has been extensively purified from the larval fat body of the tobacco hornworm (Manduca sexta) by employing chromatography on ion-exchange columns of DEAE-Sephadex, DEAE-cellulose and phosphocellulose and centrifugation on glycerol gradients. The isolated enzyme after electrophoresis on acrylamide gels shows one main band and one minor band, both having enzyme activity sensitive to alpha-amanitin. The catalytic and physicochemical properties of the enzyme are similar to those of other eucaryotic B-type RNA polymerases. The enzyme has an apparent molecular weight of 530000, is inhibited 50% by alpha-amanitin at 0.04 microgram/ml and shows maximum activity on denatured DNA at 5 mM Mn2+ and 100 mM ammonium sulfate. An antibody was obtained that cross-reacts with the pure enzyme and forms a precipitin line. This antibody does not cross react with either Escherichia coli RNA polymerase or with wheat germ RNA polymerase but does react with one of the B polymerases isolated from wing tissue of the silkmoth, Antheraea pernyi.
...
PMID:Isolation and characterization of RNA polymerase B from the larval fat body of the tobacco hornworm, Manduca sexta. 10 73

We have devised a new procedure for the purification of highly active preparations of Bacillus subtilis RNA polymerase holoenzyme. A column of heparin-agarose A-15m is used to rapidly and quantitatively adsorb RNA polymerase from the initial crude extract fraction. This affinity procedure obviates the necessity of including nucleic acid precipitation or partitioning steps and allows for rapid separation of RNA polymerase from proteolytic activity. The enzyme is further purified by preparative glycerol gradient centrifugation resulting in an overall purification in 200-fold in 24 h with near quantitative recovery of polymerase protein and activity. RNA polymerase holoenzyme is obtained by chromatography on single-stranded DNA-agarose. The in vitro transcription products made by purified preparations of B. subtilis and Escherichia coli RNA polymerase holoenzymes in response to B. subtilis phage phi 29 DNA have been analyzed, and an in vitro transcription map is presented. The E. coli RNA polymerase holoenzyme initiates transcription from three promoter sites not efficiently utilized by the B. subtilis holoenzyme under optimal conditions for RNA synthesis.
...
PMID:Purification of Bacillus subtilis RNA polymerase with heparin-agarose. In vitro transcription of phi 29 DNA. 11 9

Adenosinetriphosphatase (ATPase) [EC 3.6.1,3] activity has been found to exist in most preparations of DNA-dependent RNA polymerase [EC 2.7.7.6] obtained from Escherichia coli by a number of purification procedures so far established. Electrophoretic analysis on polyacrylamide gels demonstrated that ATP hydrolysis and RNA synthesis were catalyzed by two distinct enzyme proteins. It appears that the two enzymes are associated or have similar molecular properties. Separation of the two enzymes, the object of the present work, was achieved by three independent methods: ion exchange chromatography on a phosphocellulose column, electrophoresis in glycerol gradients, or high-salt glycerol gradient centrifugation.
...
PMID:A novel adenosine triphosphatase isolated from RNA polymerase preparations of Escherichia coli. I. Copurification and separation. 13 29

A substance has been purified from isolated nuclei of Physarum polycephalum by equilibrium and velocity gradient centrifugations, ion exchange chromatography and gel filtration which has a high molecular weight, can be labeled in vivo with 32P, is heat stable and resistant to amylases, proteases, nucleases and phosphodiesterase but is sensitive to phosphatases or hydrolysis. This material consists of phosphate and glycerol. It selectively inhibits in vitro transcription of RNA polymerases, predominantly the homologous enzyme A by binding to the enzyme. In the presence of this inhibitor of transcription a stable RNA polymerase-template complex cannot be formed. Binding to and inactivation of RNA polymerase is reversible at high ionic strength.
...
PMID:Characterization of an endogenous transcription inhibitor from Physarum polycephalum. 15 53

Promoters of genes for bacteriophage lambda and for Escherichia coli ribosomal RNA (rrnB), elongation factor Tu (tufB), ribosomal proteins L11 (rplK), L1 (rplA), L10 (rplJ), and L7/L12 (rplL), and RNA polymerase subunits beta (rpoB) and beta' (rpoC) were studied by use of two types of filter binding assays which measured E. coli RNA polymerase binding and initiation of transcription on restriction fragments of lambda rifd 18 DNA. The DNA fragments selectively retained on filters were eluted, concentrated, and analyzed by gel electrophoresis. The binding characteristics of these promotor fragments were qualitatively determined by varying the RNA polymerase, salt, and glycerol concentrations in the polymerase binding assay with HaeIII fragments of lambda rifd 18 DNA. The approximate map locations of these small HaeIII fragments were determined by HaeIII digestion of the larger, previously mapped EcoRI, HindIII, and SmaI restriction fragments of the phage DNA. The base compositions proximal to the 5' ends of mRNA's from promoters on these DNA fragments were elucidated by the polymerase initiation assay, in which the addition of various combinations of nucleoside triphosphates to the reaction allowed RNA polymerase to form high-salt-resistant initiation complexes with some of the known SmaI + EcoRI, EcoRI + HindIII, or HaeIII restriction fragments of lambda rifd 18 DNA. The data obtained by this technique are consistent with the map positions and 5' mRNA base sequences of the known lambda promotors p'R, po, pR and pL. In the main focus of this work, we have determined the approximate map locations and 5' mRNA base compositions of several promoters for known E. coli genes including rrnB, tufB, rplK,A, and rplJ,L. No promoter was detected between rplL and the rpoB,C genes. Thus our data are consistent with the conclusion of Yamamoto and Nomura (1978) that the beta and beta' mRNA is probably cotranscribed from the promoter for rplJ,L. Finally, the approximate map positions and the NTP combinations which initiated transcription of several unknown lambda and E. coli in vitro promoters are reported. The methods reported should prove useful for studying the characteristics of promoters on other cloned DNA regions.
...
PMID:Escherichia coli RNA polymerase binding and initiation of transcription on fragments of lambda rifd 18 DNA containing promoters for lambda genes and for rrnB, tufB, rplC,A, rplJ,L, and rpoB,C genes. 15 6

H1 protein, a heat-stable low-molecular-weight DNA-binding factor previously described by Cukier-Kahn et al. [Proc. Nat. Acad. Sci USA (1972) 69, 3643-3647] markedly stimulates in vitro synthesis of lac-specific RNA directed by bacteriophage lambdah80 dlac or phi80 dlac DNA templates in the presence of purified E. coli RNA polymerase holoenzyme. The extent of stimulation obtained by addition of H1 alone is usually greater than that observed with the cAMP receptor protein-cAMP combination. H1 effect varies quite appreciably (from 4- to 16-fold) with the functional state of the promoter, being much larger with lambdah80 dlac p-s, a transducing DNA carrying a superpromoter mutation, than with lambdah80 dlac p+. H1 and cAMP receptor protein effects are nearly additive, although interpretation of the data obtained at high H1 concentration is complicated by the appearance of some inhibitory property. While the cAMP-receptor-protein-mediated synthesis is asymmetrical ("I" strand almost exclusively copied), the degree of asymmetry observed with H1 is less pronounced, suggesting asymmetrical copying from the lac promoter and symmetric transcription from other regions of the DNA. Synthesis of lac-specific RNA from lambdah80 dtrp/lac or phi80 dlac p-r uv5 templates, in which lac promoters are insensitive to cAMP receptor protein, either as a result of lac fusion to the trp operon or mutation in the lac promoter, is totally H1-insensitive. Glycerol (10-15% w/w) can fully substitute for H1 in stimulating lac RNA synthesis in a fashion analogous to that reported for the cAMP receptor protein-cAMP system. The possibility that H1 acts by causing conformational modifications at the promoter level in a way that increases its functional state, and that this effect is more pronounced with operons sensitive to cAMP receptor protein, is discussed.
...
PMID:Effect of a low-molecular-weight DNA binding protein, H1 factor, on the in vitro transcription of the lactose operon in Escherichia coli. 16 21

The RNA-dependent RNA polymerase associated with vesicular stomatitis virus was isolated to apparent homogeneity by a newly developed procedure, which includes stepwise removal of proteins from virions by successive treatment with high concentrations of cesium sulfate and cesium chloride, followed by glycerol gradient centrifugation or chromatography on phosphocellulose or DEAE-Sephadex column. The polymerase thus purified contained L (large protein) and NS proteins as the intrinsic subunits and multiple species of enzyme were found which differ in the molar ratio of L to NS. Since the enzyme with the highest activity was composed of equimolar amounts of the two subunits and exhibited the sedimentation coefficient of approximately 11 S in a buffer containing 0.2 M NaCl, the structure of active protomer was suggested to be (L)1(NS)1. In accordance with this conclusion, enzyme preparations deficient in the content of NS protein, were activated by the addition of preparations deficient in the content of NS protein. The purified RNA polymerase catalyzed the synthesis of poly(A), which was covalently attached to the 3' termini of RNA products, and RNA, only in the presence of all 4 substrates. The present finding might be the first which indicates that the transcriptase itself catalyzes post-transcriptional modification of mRNA by adding poly(A) sequences to the 3'-OH termini. The molecular mechanism of the switch from transcription to poly(A) synthesis, however, remains to be investigated.
...
PMID:Function and structure of RNA polymerase from vesicular stomatitis virus. 18 23

A template-dependent RNA polymerase has been isolated from poliovirus-infected cells by assaying for the ability of the enzyme to copy poly(A) complexed to an oligo(U) primer. The polymerase was solubilized with detergent, and RNA was removed by precipitation with 2 M LiCl. The solubilized polymerase required both poly(A) and oligo(U) for activity and was stimulated by Mg2+ but was inhibited by Mn2+. Poly(A)-oligo(U)-dependent poly(U) polymerase was not found in extracts of HeLa cells until about 2 hr after poliovirus infection, and then there was a linear increase in activity until about 5 hr. Analysis of the polymerase by glycerol gradient centrifugation showed that the majority of the activity sedimented at about 4 S, indicating that it was no longer complexed with high-molecular-weight RNA or cellular membranes. This poly(A)-oligo(U)-dependent polymerase activity could represent an important component of the poliovirus RNA-dependent RNA polymerase.
...
PMID:Poliovirus-specific primer-dependent RNA polymerase able to copy poly(A). 19 96


1 2 3 4 5 6 7 8 9 10 Next >>

---