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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A type C oncornavirus has been isolated from a continuous cell line of murine adrenal carcinoma in culture. The particles have a buoyant density of 1.165 g/cm(3), exhibit typical type C morphology by electron microscopy, possess an RNA-dependent DNA polymerase, and have a high molecular weight RNA (6.1 x 10(6)) which can be denatured to a homogeneous lower molecular weight species (3.2 x 10(6)) when extracted from rapidly harvested "immature" virions. The virus is related antigenically to other mammalian oncornaviruses and exhibits a similar, although much more complex, sodium dodecyl sulfate gel electrophoretic profile of virion proteins when compared to the profiles of other type C RNA tumor viruses.
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PMID:Physical and chemical properties of an oncornavirus associated with a murine adrenal carcinoma cell line. 413 66

Polycytidylic acid [poly(rC)] covalently linked to cyanogen bromide-activated agarose is an effective affinity matrix for the RNA-dependent DNA polymerase from avian myeloblastosis virus. Poly(rC)-agarose is capable of binding large quantities of avian myeloblastosis DNA polymerase, which is then eluted by using a linear KCl gradient of increasing concentration. The DNA polymerase isolated from crude, detergent-disrupted virions by a single pass through columns of poly(rC)-agarose appears nearly homogeneous (approximately 90% pure) as determined by sodium dodecyl sulfate-polyacrylamide disc gel electrophoresis. Complete recovery of input enzymatic activity was obtained. Results suggest that polyribonucleotide columns may provide a high-yield, rapid method for the purification of oncornaviral DNA polymerase.
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PMID:Purification of avian myeloblastosis virus DNA polymerase by affinity chromatography on polycytidylate-agarose. 413 57

Influenza B/LEE/40, B/Rome/1/67, B/Hong Kong/8/73, and B/Victoria/98926/70 viruses have a similar polypeptide composition as analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. These viruses are composed of six or seven polypeptides, depending on whether one or two high-molecular-weight polypeptides are resolved, ranging in molecular weights from 27,000 to 90,400. Three of these polypeptides, namely the heavy and light hemagglutinin chains and the neuraminidase, have attached carbohydrate. Highly purified influenza B/LEE/40 and B/Rome/1/67 virus preparations have RNA-dependent RNA polymerase activity equivalent to the incorporation of 100 and 30 pmol, respectively, of (3)H-UMP per mg of virus protein per h at 37 C, which is demonstrated only in detergent-treated virus suspensions. However, no RNA-dependent DNA polymerase enzyme activity was detected in the two viruses although virus suspensions were "activated" by heat, alpha-chymotrypsin, and detergents. Other enzymatic activities were associated with purified preparations of influenza B virus and were attributed to minor contamination of virus with host cell enzymes. Thus, nucleoside and deoxynucleoside phosphohydrolase enzymes were active in the absence of detergents and catalyzed the release of 1,200 and 1,800 nmol of P(i) per mg of virus protein in 30 min at 37 C from ATP and dATP substrates. Thin-layer chromatography indicated that the products of the phosphohydrolase enzymes of influenza B/LEE/40 were mainly nucleoside diphosphate and monophosphate. The latter enzymes were tightly bound to influenza B/LEE/40 virus and could not be removed completely by repeated centrifugation, including centrifugation of the virus to equilibrium in density gradients of 25 to 40% (wt/vol) cesium chloride. A low degree of RNase (approximately 0.01 mug% contamination) and phosphatase (10-30 nmol of P(i) released per mg of virus protein per 30 min) activity was detected in some, but not all, influenza B/LEE/40 virus preparations.
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PMID:Polypeptide composition of Influenza B viruses and enzymes associated with the purified virus particles. 435 55

An RNA-dependent DNA polymerase was isolated from purified virions of endogenous oncornaviruses released by the MOPC-315 murine myeloma cell line. Upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the purified enzyme was found to consist of two major polypeptides with molecular weights of about 28,000 and 26,500. The active enzyme had a molecular weight of approximately 56,000, as calculated from its sedimentation on glycerol density gradients, indicating that it is probably a dimer of the two subunit polypeptides. The isolated MOPC-315 virus polymerase exhibited all three activities known to be found in the DNA polymerase from oncornaviruses, namely, an RNA-dependent DNA polymerase, a DNA-dependent DNA polymerase, and an RNase H. The RNA-dependent polymerase activity showed a prounced preference for Mn2+ over Mg2+, whereas the DNA-dependent and RNase H reactions were catalyzed by these two cations to an almost equal extent. The purified polymerase was found to be immunologically related to the polymerase of Rauscher murine leukemia virus.
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PMID:RNA-dependent DNA polymerase of an endogenous type C virus of mice: purification and partial characterization. 615 78

Polyguanylic acid was found to be a potent inhibitor of RNase H associated with mammalian viral reverse transcriptase, indicating a strong interaction between polyguanylic acid and the reverse transcriptase protein. Based on this observation, we have developed three simple procedures for the purification of mammalian viral reverse transcriptases. In the first procedure, a nucleic acid-free extract of Rauscher murine leukemia virus was applied to a column of phosphocellulose and the reverse transcriptase was eluted by a low concentration (50 microM) of polyguanylic acid. Polyadenylic acid and polyuridylic acid could not replace polyguanylic acid for the elution. In the second procedure, a polyuridylic acid-Sepharose column was substituted for phosphocellulose, and the elution was again achieved by polyguanylic acid. In the third affinity procedure, the reverse transcriptase in a nucleic acid-free viral extract was incubated in the cold with 50 microM polyguanylic acid and the complex was adsorbed onto a DEAE-cellulose column. After washing to remove uncomplexed and weakly complexed proteins, the reverse transcriptase was eluted in a concentrated form at 0.3 M NaCl with a recovery of greater than 70%. by polyacrylamide gel analysis in the presence of sodium dodecyl sulfate, the enzyme appeared to be nearly pure.
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PMID:Simple affinity procedure for the purification of mammalian viral reverse transcriptases. 616 Feb 61

The effect of the ribonuclease inhibitor polyvinyl sulfate on the activity of retroviral DNA polymerase and terminal c transferase was examined. This substance was found to be a potent inhibitor of these enzymes by virtue of competition between the sulfated sidechains of the molecule and the template primer for a site on the enzyme. The significance of these findings in relation to searching for reverse transcriptase is discussed.
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PMID:Inhibition of reverse transcriptase by polyvinyl sulfate (PVS). 616 65

Poly(A)-containing RNA was isolated from total RNA of swine gastric mucosa by oligo(dT)-cellulose chromatography, and was translated in a wheat germ cell-free system. Analysis of the translation products by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that pepsinogen was a major translation product and was synthesized as two different molecular forms with apparent molecular weights of 45,000 and 43,000. The pepsinogen translated in the in vitro translation system had no autocatalytic activity. The pepsinogen mRNA was further purified by sucrose density gradient centrifugation, where the mRNA activity for pepsinogen was located around 15 S. At this stage, the translation product of the pooled fractions appeared to be almost exclusively pepsinogen. The peptide maps of the pepsinogen which was translated in vitro and digested by alpha-chymotrypsin and by Staphylococcus aureus V8 protease were nearly identical with the corresponding peptide maps of standard pepsinogen. The double-stranded complementary DNA which had been synthesized from the partially purified mRNA by avian myeloblastosis virus reverse transcriptase was cloned in Escherichia coli chi 1776, using plasmid pBR322 as a cloning vector. A colony carrying pepsinogen cDNA sequence was identified by in situ colony hybridization using the cDNA synthesized from the partially purified mRNA as a probe and further by a positive hybridization-translation assay. One of the recombinant clones (pSPcA1) had an insert of about 850 base pairs, and the nucleotide sequence analysis revealed that pSPcA1 codes for swine pepsinogen.
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PMID:Molecular cloning of complementary DNA to swine pepsinogen mRNA. 617 Jun 44

Visna virus is a retrovirus which replicates in fibroblast-like cells of the sheep choroid plexus through a lytic cycle. Visna virions contain three major low-molecular-weight proteins (p30, p16, and p14) which, together with the genomic RNA and several molecules of reverse transcriptase, constitute the core structure of the virions. The core is surrounded by an envelope containing a major glycoprotein (gp135). By analogy with the oncoviruses, these three groups of structural proteins (i.e., the internal proteins, the envelope glycoprotein, and the reverse transcriptase) are probably encoded by the gag, env, and pol genes, respectively. To elucidate the genetic organization of the visna virus genome and its expression, we studied the synthesis of viral proteins in infected sheep choroid plexus cells. Intracellular viral proteins were detected by immunoprecipitation of pulse-labeled cell extracts with monospecific sera raised against p30, p16, and gp135 and resolution of the proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Immunoprecipitation with anti-p30 and anti-p16 sera allowed the characterization of the 55,000-dalton polypeptide precursor to internal virion proteins p30, p16, and p14 (Pr55(gag)). Tryptic peptide mapping confirmed the precursor-product relationship between Pr55(gag) and the three internal proteins. In addition, a gag-related polypeptide of 150,000 daltons was also detected. This polypeptide, which was less abundant than Pr55(gag), is a likely precursor to the viral reverse transcriptase (Pr150(gag-pol)). Pr55(gag) and Pr150(gag-pol) are not glycosylated. The precursor related to major envelope protein gp135 is a glycosylated polypeptide with an average molecular weight of 150,000 (gPr150(env)). Pulse-chase experiments indicated that gPr150(env) matures into glycoprotein gp135 intracellularly; however, gp135 was never preponderant in cell extracts. The non-glycosylated from of gPr150(env), which accumulated in the presence of 2-deoxy-d-glucose, appeared as a polypeptide of about 100,000 daltons. These results indicated that visna virus codes for the largest non-glycosylated env-related precursor among all of the retroviruses and therefore probably contains the largest env gene.
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PMID:Precursor polypeptides to structural proteins of visna virus. 617 45

The p30 antigen from Rauscher leukemia virus (R-MuLV) was separated into two fractions by chromatography on either phosphocellulose or DEAE-cellulose. The p30-I and p30-II were indistinguishable immunologically or by isoelectrofocusing and gel electrophoresis. An ATPase activity was tightly associated with p30-II that could not be separated by ion-exchange chromatography, isoelectrofocusing, or glycerol velocity gradient sedimentation. The ATPase hydrolyzed the gamma phosphate from only ATP or dATP. Immunoglobulin directed against R-MuLV p30 completely inhibited the p30-II associated ATPase. Glycerol velocity gradient analysis showed that p30-I sedimented as a 30-kDa species while the p30-II and its associated ATPase sedimented as a 60-kDa species. The p30-II was converted entirely to a 30-kDa form by treatment with 0.2% (w/v) lithium dodecyl sulfate, suggesting that it represented a complexed species of p30. Finally, p30-II was found to stimulate the activity of R-MuLV reverse transcriptase, but p30-I had no effect on the activity of the enzyme. These results suggested the existence of at least two different forms of p30 in R-MuLV.
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PMID:Characterization of a p30 fraction from Rauscher leukemia virus which has an associated ATPase activity. 620 91

The diterpene ester promoter of mouse skin tumors, 12-O-tetradecanoylphorbol-13-acetate (TPA), efficiently induces Epstein-Barr virus (EBV)-associated DNA polymerase (DNA nucleotidyltransferase) activity in the EBV-producing lymphoblastoid cell line, P3HR-1. With the use of intervent dilution chromatography followed by sequential DEAE-cellulose and phosphocellulose column chromatography, the virus-associated enzyme has been isolated and purified 300-fold. The partially purified EBV DNA polymerase activity could be distinguished from cellular polymerases by its activation with salt and its degree of sensitivity to N-ethylmaleimide and phosphonoacetic acid. The enzyme showed maximum activity for copying activated calf thymus DNA in the presence of 100 mM ammonium sulfate. In the absence of salt, the enzyme utilized with high efficiency deoxyoligomer-homopolymer templates, but failed to copy poly(rA) . oligo(dT)10 and oligo(dT)10, showing that the enzyme had properties distinct from DNA polymerase gamma, reverse transcriptase, and terminal deoxynucleotidyltransferase. The partially purified enzyme is strongly inhibited by acyclovir triphosphate and thus has properties similar to herpes simplex virus DNA polymerase.
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PMID:Induction of Epstein-Barr virus-associated DNA polymerase by 12-O-tetradecanoylphorbol-13-acetate. Purification and characterization. 624 99


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