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

The synthesis of a complementary DNA copy (cDNA) of hen ovalbumin mRNA using AMV RNA-directed DNA polymerase was studied under different conditions of salt, deoxyribonucleotide concentrations, temperature, and time. It was observed that in the absence of monovalent cation at 46 degrees C a complete transcript of ovalbumin mRNA could be effected by the enzyme. The minimum deoxyribonucleotide requirement for complete synthesis was 35 muM for dATP, dGTP, and dCTP and 200 muM dTTP. By a number of different experimental criteria which included sedimentation on alkaline sucrose gradients and electrophoresis in polyacrylamide gels containing 98% formamide, direct electron microscope visualization, and protection of ovalbumin [25I]mRNA from nuclease digestion it could be demonstrated that a considerable fraction of a complete mRNA transcript was indeed synthesized. The cDNA/ovalbumin mRNA hybrid had a Tm on hydroxylapatite of 92 degrees C, indicating the synthesis of a RNA transcript with a high fidelity. When such a complete ovalbumin [3H]cDNA was synthesized with a specific activity of 10(8) cpm/mug and hyfridized to an excess of chick DNA, the kinetics of hybridization indicated that the cDNA was comprised of a nonrepetitive sequence.
...
PMID:The synthesis and properties of the complete complementary DNA transcript of ovalbumin mRNA. 5 72

A cat kidney cell line, CRFK-F2, was successfully inoculated in suspension and in monolayer culture with a purified mouse mammary tumor virus derived from RIII milk. The virus produced by the infected cells was identified by immunogluorescence, electron microscopy, and RNA-directed DNA polymerase assays; it was a B-type virion that did not cross-react with mouse or feline leukemia-sarcoma viruses, had spikes on its envelope, and had a RNA-directed DNA polymerase reaction that was typical of mouse mammary tumor virus. The producing cells were identified as cat cells by chromosome number, cytotoxic assays, and isoenzyme migratory patterns. A standardized method for the in vitro inoculation of cat cells is described that presently permits highly reproducible results. For the first time, the mouse mammary tumor virus is seen replicating in cells from another species, thus offering an opportunity to study the kinetics of infection of that virus.
...
PMID:Experimental infection of a cat kidney cell line with the mouse mammary tumor virus. 5 5

The synthesis of DNA products complementary to artificial templates by the enzyme RNA-directed DNA polymerase isolated from avian myeloblastosis virus has been studied. Of the single base polyribonucleotides, poly (rC), poly(rA), and poly(rI) were active while poly (rG) and poly (rU) were almost inactive. The minimum length showing activity for an oligo (rC) template was 9; the minimum primer length of oligo(dG) was 3 or 4. In order to examine the fidelity of transcription, single base oligoribonucleotides of defined length were studied. Using (rC)13 as template and (dG)8as primer, the oligo (dG) product coelectrophoresed with the template. However, using (rA)-20 as template and (dT)10 as primer, a large (10-16s) product was formed. Similarly, using oligo (rI) (2.5S) as template and (dC)10 as primer, a large (greater than 22s) product was formed. No significant activity was obtained with oligo (rU) templates. RNA-directed DNA polymerase transcribes the various oligonucleotides differently: slippage with oligo (rA) and oligo (rI), faithful transcription with oligo (rC), and poor transcription with oligo (rU).
...
PMID:Transcription of single base oligonucleotides by ribonucleic acid-directed deoxyribonucleic acid polymerase. 5 99

The major species of primer RNA required for the initiation of DNA synthesis by the Rous sarcoma virus RNA-directed DNA polymerase can be aminoacylated by tryptophan. Furthermore, an intact 3' terminus is required for the primer to function in the initiation of DNA synthesis.
...
PMID:Initiation of DNA synthesis by the avian oncornavirus RNA-directed DNA polymerase: tryptophan tRNA as the major species of primer RNA. 5 57

The alpha beta DNA polymerase of avian myeloblastosis virus was treated with dimethyl sulfoxide to dissociate the enzyme subunits. The dimethyl sulfoxide treated enzymes were passed over phosphocellulose to purify and characterize the dissociated subunits as well as to remove the dimethyl sulfoxide. RNA-directed DNA polymerase, RNase H, and nucleic acid-binding activity were monitored, as well as the subunit structure (on sodium dodecyl sulfate-polyacrylamide gels) of the various enzyme species obtained. With 30% dimethyl sulfoxide, the majority of DNA polymerase and RNase H activities as well as the alpha subunit were displaced from the alpha beta DNA polymerase position on phosphocellulose (0.23 M potassium phosphate) to the alpha DNA polymerase position (0.1 M). The association of DNA polymerase and RNase H activities with the alpha subunit suggests that alpha is the enzymatically active subunit in alpha beta. In addition to alpha DNA polymerase, a minor polymerase species eluted from phosphocellulose at 0.4 M potassium phosphate. The dissociated beta subunit eluted from phosphocellulose at a wide range of salt concentrations (0.28 to 0.5 M potassium phosphate). The dissociated beta subunit bound 3H-labeled murine leukemia virus RNA and [3H]poly(dT)-poly(dA) approximately 20-fold more avidly than alpha DNA polymerase alone. In contrast to the results with the alpha subunit, there was no correlation between DNA polymerase and RNase H activity profiles and the elution profile of the beta subunit from phosphocellulose. These observations suggest the beta subunit is either enzymatically inactive or possesses limited DNA polymerase and RNase H activity when compared with the alpha subunit.
...
PMID:Dissociation of alpha beta DNA polymerase of avian myeloblastosis virus by dimethyl sulfoxide. 5 61

Milk from a number of species (e.g., man, mouse, rat, dog, and cow) contains inhibitors of the RNA-directed DNA polymerase. When attempts are made to isolate virions from the milk, part of the inhibitors follow the virions in the purification. The amount of inhibitors varies in different milk samples. These inhibitors can probably account for the large discrepancies reported in studies of the presence of oncornaviruses in human milk. Phosphatases bound to subcellular particles or fragments seem to be the most important inhibitors in the milk interfering with the RNA-directed DNA polymerase assay. It is shown that the inhibitory enzymes can be completely removed by sedimentation of the milk through a Metrizamide gradient.
...
PMID:Removal of inhibitors against RNA-directed DNA polymerase activity in human milk. 5 90

The RNA-directed DNA polymerase (deoxynucleosidetriphosphate:DNA deoxynucleotidyltransferase EC 2.7.7.7) of avian oncornavirus requires a tryptophan tRNA (tRNATrp) primer molecule located close to the 5' end of the viral RNA genome for the initiation of DNA synthesis in vitro. In this communication we demonstrate that the DNA product, transcribed from avian myeloblastosis virus (AMV) 35S RNA containing only tRNATrp as primer, is located also at the 5' end of the RNA genome. More importantly, we demonstrate that these 5' terminal DNA transcripts contain nucleotide sequences complementary to the 3' end of the genome. We have interpreted these results to mean that the genome. We have interpreted these results to mean that the 3' and 5' termini of the AMV 35S RNA genome become juxtaposed with each other either before or immediately after DNA synthesis has begun. These results are discussed in regard to the mechanism for synthesis of the circular forms of oncornavirus proviral DNA.
...
PMID:Evidence for circularization of the avian oncornavirus RNA genome during proviral DNA synthesis from studies of reverse transcription in vitro. 5 20

Type C viruses were isolated from embryo cultures of two different rat strains, Sprague-Dawley and Fischer. Both viruses (termed rat leukemia virus, RaLV) were released spontaneously from rat embryo cells, have a density of 1.14 to 1.15 g/cm(3) based on equilibrium sedimentation in sucrose gradients, contain 60-70S RNA, RNA-directed DNA polymerase, and rat type C virus-specific 30,000 molecular-weight-protein determinants. Molecular hybridization studies using the Sprague-Dawley RaLV 60-70S RNA show that the virus-specific nucleotide sequences are present in the DNA of rat embryos. Both Sprague-Dawley and Fischer RaLV can rescue the murine sarcoma virus genome from Kirsten murine sarcoma virus-transformed nonproducer cells and are neutralized by antisera to the RPL strain of RaLV. In contrast to previous RaLV's, these viruses propagate in their own cells of origin as well as in cells of heterologous rat strains.
...
PMID:Spontaneous release of endogenous ecotropic type C virus from rat embryo cultures. 5 77

The inoculation of L2C guinea pig leukemia cells into strain 2 guinea pigs results in the death of the animals within 12 to 15 days. Death is preceded by the simultaneous appearance in the plasma of (i) elevated leukocyte levels, (ii) extracellular virus particles, and (iii) a particle-associated RNA-directed DNA polymerase. This enzyme activity has a cation preference identical to that of the type B bromodeoxyuridine-induced guinea pig virus, i.e., an Mg2+ optimum at 20 mM and no activity using Mn2+. Competitive molecular hybridization studies also revealed that the plasma of leukemic guinea pigs contained approximately 2 X 10(9) genome equivalents per ml of an RNA that is homologous to the RNA of the bromodeoxyuridine-induced guinea pig virus. Morphological observations indicate that most, but not all, of the extracellular particles observed in leukemia plasma are derived from the intracisternal particles seen in the L2C tumor cells. The possibilities that either two viral populations are present or that the in vivo morphogenesis of the type B bromodexoyuridine-inducible guinea pig virus is markedly different from its in vitro morphogenesis are discussed.
...
PMID:Characterization of the oncornavirus particles in the plasma of guinea pigs with L2C leukemia. 5 78

We have compared the relative merits of several procedures for the isolation of RNA-directed DNA polymerase (EC 2.7.7.7.) from cells using a reconsituted model system consisting of a mixture of woolly monkey (simian) sarcoma virus and a cultured human lymphoblastoid cell line, NC-37. When the cell-virus mixture was gently disrupted and fractionated by differential centrifugation, most of the added polymerase was recovered associated with a particulate fraction obtained from the post-mitochondrial supernatant. Purification of the polymerase was best achieved starting from this fraction. The particulate fraction itself can be purified by gel filtration through a Sepharose 2 B column. This procedure did not significantly alter the composition of viral and cellular DNA polymerases. Whereas as little as 7.5 - 10(5) viral particles were sufficient for the detection of RNA-directed DNA polymerase activity, a minimum of about 10(11) particles were necessary for the isolation and unequivocal characterization of the enzyme from the cell-virus mixture by subcellular fractionation and chromatographic separation from cellular DNA polymerases. Purified RNA-directed DNA polymerase had the same primer-template characteristics, sedimentation properties, and immunological cross reactivity as the enzyme purified from density gradient-banded virions of simian sarcoma virus. Methods involving total extraction of the cell-virus mixture either by repeated freezing and thawing followed by detergent treatment or by Dounce homogenization and treatment with high salt and detergent failed to provide RNA-directed DNA polymerase free of cellular DNA polymerases. Because of this, low levels of cellular RNA-directed DNA polymerase may be missed when these approaches are used.
...
PMID:A comparative evaluation of methods for isolation of RNA-directed DNA polymerase from cells in a reconstituted system. 5 69


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

---