Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.48 (transcriptase)
 9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The in vitro transcription reaction of bluetongue virus (BTV) is characterized by a core-mediated, temperature-dependent inhibition at high core concentrations and temperatures (Van Dijk & Huismans, 1980; Huismans, Van Dijk & Els, 1987a). It has been found that this inhibition is reversible and that an inactivated transcriptase reaction mixture can be reactivated by lowering the temperature of the reaction from 37 degrees C to 28 degrees C. In the same way it is possible to inactivate a reaction by increasing the incubation temperature from 28 degrees C to 37 degrees C. It was also found that the inhibition is counteracted by the addition of sucrose or glycerol. At relatively low core concentrations and in the presence of sucrose it is possible to obtain conditions under which transcription at 37 degrees C is more efficient than at 28 degrees C. The latter conditions probably reflect much better the in vivo temperature optimum for the BTV transcriptase than the in vitro conditions at very high core concentrations.
Onderstepoort J Vet Res 1987 Dec
PMID:The identification of factors capable of reversing the core-mediated inhibition of the bluetongue virus transcriptase. 283 1

The nucleotide sequence of segment 1 of the double stranded RNA genome of bluetongue virus serotype 10 (BTV-10), encoding the largest viral core protein, VP1, has been determined. Linear sequence analysis of the predicted amino acid sequence of the 149-K Da protein, a putative component of the viral RNA-directed RNA polymerase, revealed extensive homology with the vaccinia virus 147K Da DNA-directed RNA polymerase subunit. Similar homologies were detected between the VP1 polypeptide and the beta chain subunit of Escherichia coli and common tobacco chloroplast RNA polymerases, yeast RNA polymerase II and III and fruit fly polymerase II.
Nucleic Acids Res 1988 Dec 23
PMID:Evidence for genetic relationship between RNA and DNA viruses from the sequence homology of a putative polymerase gene of bluetongue virus with that of vaccinia virus: conservation of RNA polymerase genes from diverse species. 285 May 42

Acidic chloroform-methanol soluble proteins possessing hydrophobic properties and capable of inhibiting in vitro transcriptase activity of influenza virus RNP were detected in native and partially purified human leukocyte interferon (IFN) preparations. Purification of IFN resulted in the removal of at least a portion of such proteins; however, no proteins have been found in highly-purified IFN preparations.
Acta Virol 1985 Dec
PMID:Studies of proteins soluble in acidic chloroform-methanol isolated from crude human leukocyte interferon preparations. 286 58

Using morphological and cell biological techniques, we have shown that the RNA replicase of Semliki Forest and Sindbis virus (two closely related alphaviruses) is located in complex ribonucleoprotein structures associated with the cytoplasmic surface of modified secondary lysosomes and endosomes. These nucleoprotein complexes often form a bridge between the membrane of the endocytic vacuole and the rough endoplasmic reticulum where the synthesis of the structural proteins of these viruses occurs. The results suggest that these cytopathic vacuoles constitute sites not only for viral RNA synthesis, but also for translation of structural proteins, and for the assembly of nucleocapsids.
J Cell Biol 1988 Dec
PMID:Alphavirus RNA replicase is located on the cytoplasmic surface of endosomes and lysosomes. 290 46

The large gene, L, of vesicular stomatitis virus (VSV), which codes for the multifunctional RNA-dependent RNA polymerase, was assembled from five overlapping cDNA clones. The sequence of the 6.4-kilobase gene of the final construct was identical to the consensus sequence reported earlier. The gene was inserted into the simian virus 40 transient expression vector pJC119. Antibodies directed against synthetic peptides corresponding to the amino and carboxyl termini of the L protein were raised in rabbits. Both antibodies specifically immunostained the cytoplasm of COS cells that had been transfected with the vector DNA. The expressed L protein was immunoprecipitated from cell extracts and it was identical in size to the L protein of the virion (241 kilodaltons). Most importantly, COS cells that expressed the recombinant L protein transcribed, replicated, and consequently complemented and rescued temperature-sensitive RNA polymerase mutants of VSV at the nonpermissive temperature. The kinetics of virus release were similar to those of a wild-type VSV infection. We conclude that the recombinant RNA polymerase protein L is indistinguishable in its size and its functions from the VSV polymerase.
Proc Natl Acad Sci U S A 1985 Dec
PMID:Expression of a cDNA encoding a functional 241-kilodalton vesicular stomatitis virus RNA polymerase. 299 88

Replication of the infectious RNA genome of poliovirus is accomplished in cells by the viral RNA polymerase through negative-strand RNA intermediates. Full-length negative-strand poliovirus RNA was synthesized in vitro by transcription of infectious poliovirus cDNA with bacteriophage SP6 DNA-dependent RNA polymerase. When provided with this negative-strand RNA as template, the poliovirus RNA-dependent RNA polymerase synthesized full-length positive-strand molecules. The positive-strand RNAs synthesized in vitro were infectious when transfected into HeLa cells. In contrast, positive-strand copies of poliovirus RNA synthesized in vitro by SP6 polymerase, using a poliovirus cDNA template, were not infectious. Production of infectious positive-strand RNA by the poliovirus polymerase was not observed when magnesium or negative-strand RNA template was omitted from the reaction mixture. Infectivity of the product RNA was not destroyed by DNase treatment. The specific infectivity in HeLa cells of in vitro-synthesized positive-strand RNA was 4 X 10(4) plaque-forming units/micrograms of RNA.
Proc Natl Acad Sci U S A 1985 Dec
PMID:In vitro synthesis of infectious poliovirus RNA. 300 3

To facilitate further studies of flavivirus transcription, cell extraction methods and in vitro reaction conditions which increased West Nile virus (WNV) RNA-dependent RNA polymerase activity were determined. Subcellular fractions from WNV-infected BHK-21/W12 cells were characterized with regard to their protein and RNA content and in vitro polymerase activity. In both a cytoplasmic fraction, designated S1, and a fraction enriched for outer nuclear membranes, designated S2, seven virus-specific proteins, NS5 (96 kilodaltons [kDa]), NS3 (67 kDa), E (48 kDa), NS1 (47 kDa), ns4a (26 kDa), ns2a (17 kDa), and ns2b (14.5 kDa), were detected. The fractions also contained virus-specific RNA and cellular rRNA and mRNA. Polymerase activity in S1 and S2 fractions from WNV-infected cells was concentrated by pelleting and consisted of two types of enzyme activities: the WNV RNA-dependent RNA polymerase and terminal transferases of cellular origin. Enhanced levels of WNV polymerase activity were obtained from these cell fractions by altering several of the in vitro reaction conditions. Although Mg2+ was the divalent cation preferred by WNV polymerase, virus-specific in vitro transcription was detected at reduced levels when Mn2+ (0.05 or 0.5 mM) was present as the sole divalent cation. Product analysis revealed that the viral polymerase incorporated radiolabeled ribonucleotides into three distinct RNA species. Free single-stranded genome-sized RNA which was LiCl insoluble and RNase sensitive was found by fingerprint analysis to have an oligonucleotide pattern similar to that of WNV genomic RNA. RNA molecules which comigrated as a broad band near the top of the gel were separable into LiCl-insoluble, partially RNase-sensitive replicative-intermediate RNA and LiCl-soluble, RNase-resistant replicative-form RNA. The cellular transferases added UMP or AMP residues to the 3'-termini of cellular mRNA, tRNA, and 18S and 28S rRNA. Although a cellular terminal transferase has been reported to function in initiation of poliovirus transcription, no labeling of the WNV RNA by either of these cellular enzymes was detected. Therefore, they appear to play no specific role in flavivirus RNA synthesis.
J Virol 1986 Dec
PMID:Characterization of West Nile virus RNA-dependent RNA polymerase and cellular terminal adenylyl and uridylyl transferases in cell-free extracts. 302 63

Subviral particles were isolated from lysates of simian rotavirus SA11-infected cells by sedimentation through sucrose gradients and separated by equilibrium centrifugation in CsCl gradients. A cell-free system that supports rotavirus RNA replication and transcription was used to identify particles in the CsCl gradients with associated polymerase activity. These data indicated that particles with densities of 1.34 and 1.38 g/cm3 were responsible for most of the transcriptase activity present in infected cells. Electrophoretic analysis showed that particles at 1.34 g/cm3 were analogous to double-shelled virus, consisting of the inner shell proteins VP1, VP2, and VP6, the outer shell proteins VP3 and VP7, and DS RNA. Particles of 1.38 g/cm3 were similar to single-shelled virus containing the inner shell proteins and DS RNA. The pellets of the CsCl gradients were enriched for subviral particles with replicase activity. Analysis of the pellets suggested that replicase particles contain a core of VP1 and VP2 that is similar to that found in single- and double-shelled virus but contain significantly less VP6 protein per particle than those with transcriptase activity. Two particles were detected in infected cells that contain no detectable polymerase activity; one consisted primarily of the structural proteins VP2, VP3, and VP6 and the other of the nonstructural protein NS35.
Virology 1986 Dec
PMID:Characterization of subviral particles in cells infected with simian rotavirus SA11. 302 5

Flaviviruses encode seven non-structural proteins for which functions have not yet been described. The identification of the viral and possible host proteins which may be involved in flavivirus replication has been impeded by the fact that the viral replication complexes are tightly associated with endoplasmic reticular membranes within infected cells and that in vitro polymerase activity is associated with large membrane fragments. To facilitate further study of flavivirus replication complexes, selected ultrapure detergents were analysed for their effect on West Nile virus (WNV) in vitro RNA-dependent RNA polymerase activity and for their ability to release functional replication complexes from partially purified intracellular BHK-21 membrane fragments. A few previous reports indicated that flavivirus in vitro polymerase activity was sensitive to detergent treatment. The present study indicates that WNV polymerase activity is variably inhibited depending on the concentration and identity of the detergent used. Of the five detergents (Tween 20, maltoside, octylglucoside, lubrol PX and sodium deoxycholate) tested, sodium deoxycholate was the most efficient at releasing functional viral replication complexes from intracellular membranes.
J Gen Virol 1988 Dec
PMID:Separation of functional West Nile virus replication complexes from intracellular membrane fragments. 319 3

The arenavirus genomic L RNA segment represents approximately 70% of the viral genetic material but current understanding of the organization, regulation, and functioning of the viral L products remains limited. Biological studies with reassortant viruses have implicated the L RNA segment in the lethal infection of adult guinea pigs produced by LCMV-WE but no further explanation of the pathogenic process is presently available. We have initiated a detailed molecular analysis of LCMV L products based on construction and characterization of L-specific cDNA clones and synthesis of L-specific hybridization probes. Nucleotide sequencing studies have allowed the derivation of a partial amino acid sequence for a predicted L protein and antisera raised against synthetic peptides have demonstrated an L protein in Western blotting experiments. Using this approach we have identified a single high molecular weight protein (approximately 200,000 Da) in purified virions and in viral ribonucleoprotein complexes extracted from acutely infected tissue culture cells. This L protein is translated from a nonpolyadenylated, genomic complementary L mRNA and potentially represents part or all of the viral RNA-dependent RNA polymerase.
Virology 1987 Dec
PMID:Analysis of the genomic L RNA segment from lymphocytic choriomeningitis virus. 331 94


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