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 presence of an RNA-dependent RNA polymerase is demonstrated in purified rotavirus particles. Optimum polymerase activity was found between 45 to 50 degrees C, at pH 8, and in the presence of 10 mM-magnesium ions. The polymerase product was highly sensitive to pancreatic RNase (97%) in low or high salt concentration. The enzyme was activated by EDTA treatment of intact particles or heat shock. The similarities between reovirus, blue-tongue virus and rotavirus polymerases are discussed.
J Gen Virol 1977 Sep
PMID:Ribonucleic acid polymerase activity associated with purified calf rotavirus. 2 Dec 25

Upon infection of Chinese hamster ovary cells (CHO), vesicular stomatitis (VSV) virus synthesizes two membrane proteins (the VSV glycoprotein and the VSV matrix or membrane (M) protein) and three nonmembrane proteins (the VSV nucleocapsid, the viral transcriptase, and an NS protein). We have used the VSV-infected cell as a model system for the study of the site of synthesis of these membrane and nonmembrane proteins. We have isolated VSV mRNA from free polyribosomes, membrane-bound polyribosomes, and the postribosomal supernatant, and identified the individual species of VSV mRNA present in each fraction. The mRNA which encodes the VSV glycoprotein is found exclusively on membrane-bound polyribosomes, while the mRNAs which encode the VSV, M, N, and NS proteins are found in free polyribosomes, in the membrane fraction of the cell, and in the postribosomal supernatant. Our results suggest that the VSV glycoprotein is synthesized exclusively on membrane polyribosomes, while at least some of the M, N, and NS proteins are made on free polyribosomes.
J Biol Chem 1975 Sep 10
PMID:Site of synthesis of membrane and nonmembrane proteins of vesicular stomatitis virus. 16 63

The RNA polymerase in cells infected with three group I mutants of vesicular stomatitis virus has been examined. Mouse L cells were incubated at the permissive temperature (30 degrees C) for a few hours after infection to allow the development of secondary transcription. The temperature dependence of the secondary transcription system was determined from the incorporation of labelled uridine, in the presence of cycloheximide, at 30 and at 38 degrees C, the later temperature being non-permissive for viral replication. In cells infected with mutants W14, W28, and G11 at a low multiplicity (20 PFU/cells) secondary transcriptase activity was markedly temperature-sensitive after 3 and 5 h of infection at 30 degrees C. At a high multiplicity of infection (1000 PFU/cell) cells infected with W28 showed considerable RNA synthesis at 38 degrees C after 3 h at 30 degrees C. RNA synthesis was also observed in W28-infected cells in which protein synthesis was allowed to continue after the shift from 30 to 38 degrees C. In the latter two cases the RNA synthesized contained 12-18S species but little or no 30S mRNA.
Can J Microbiol 1976 Sep
PMID:Temperature-sensitive mutants of vesicular stomatitis virus: viral RNA synthesis in cells infected with mutants belonging to complementation group I. 18 5

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.
Proc Natl Acad Sci U S A 1977 Sep
PMID:Poliovirus-specific primer-dependent RNA polymerase able to copy poly(A). 19 96

The L and NS proteins of vesicular stomatitis virions (New Jersey serotype) were solubilized with Triton X-100 and high-salt buffer and recombined with purified nucleocapsids under conditions similar to those used to reconstitute transcriptase activity in vitro. The nucleocapsid-bound L and NS proteins were separated from unbound proteins on a glycerol gradient. The rebinding of L and NS proteins mimics the in vivo binding in that at saturation the ratio of L and NS molecules to N molecules is approximately the same as observed in the intact virion. L and NS proteins were separated and added back independently and in combination to the template. The purified NS protein bound to the template in the absence of L protein. However, the L protein binding appeared to depend on the presence of NS protein. The presence of Mg2+ and nucleotides, which is required for transcription, was not necessary for the rebinding of L and NS proteins.
J Virol 1978 Sep
PMID:Rebinding of transcriptase components (L and NS proteins) to the nucleocapsid template of vesicular stomatitis virus. 21 81

Phage SP RNA-dependent RNA polymerase (SP replicase) was purified from Escherichia coli infected with RNA phage SP. The enzyme was found to be composed of four non-identical polypeptides, i.e. subunits I, II, III, and IV and molecular weights of 74,000, 69,000, 47,000, and 36,000 daltons, respectively. As in the case of phage Qbeta replicase, the largest polypeptide is identical with the ribosomal protein S1, and subunits III and IV with polypeptide chain elongation factors EF-Tu and EF-ts, respectively.. This is based on the behaviour of the subunits on SDS-polyacrylamide gel electrophoresis, isoelectric focusing and immunological cross-reaction. Subunits I, III, and IV of SP replicase are derived from the host cell, while subunit II is coded by phage RNA genome. The striking coincidence of the composition and entity of the structural components of SP replicase with those of Qbeta replicase may indicate the structural and functional requirements of host-derived polypeptides in RNA replicase. The binding activity of S1 (in 70S ribosome comples) to poly (U) is retained in SP replicase complex. In contrast, the GDP binding activity of EF-Tu is masked in SP replicase. It is concluded that S1 is required functionally whereas EF-Tu.EF-Ts are required structurally in RNA replicase.
J Biochem 1978 Sep
PMID:Identification of host-derived subunits of phage SP RNA-dependent RNA polymerase (SP replicase). 36 4

A purification method for Semliki Forest virus-specified RNA-dependent RNA polymerase from BHK cells is described. The procedure entails (i) the preparation of a crude cell lysate by Dounce homogenization of cells 3-5 h post-infection, (ii) differential centrifugation to give a 15 000 g 'mitochondrial' pellet, (iii) equilibrium centrifugation on discontinuous sucrose gradients (Friedman et al. 1972) to give a membranous band of density 1-16 g/ml, (iv) solubilization with Triton N-101 and velocity centrifugation to give a 25S solubilized polymerase complex and (v) affinity chromatography through an oligo (dT)-cellulose matrix bearing immobilized 42S virus particle RNA. The overall purification was approx. 360-fold with a 5% recovery of activity. Of the various intermediate fractions in the purfication procedure, only the relatively crude post-nuclear supernatant fraction was competent to synthesize the major single-stranded RNAs found in infected cells. Other fractions incorporated precursor only into replicative intermediate (RI) or replicative from (RF). Analysis of the product RF showed that it was of the same size and could bind to the same extent to oligo (dT)-cellulose as the RF isolated directly from lysates of infected cells. Displacement hybridization and ribonuclease digestion suggested that the purified polymerase could only complete previously initiated progeny positive strands using negative strands as template and, even in its most highly purified form, was still tightly bound to its template. Analysis on polyacrylamide slab gels revealed the presence of three 35S-labelled polypeptides in the purified polymerase preparation, but a polypeptide which had identical electrophoretic mobility to the lowest mol. wt. polypeptide of the purified polymerase was also present in material from mock-fected cells which had been taken through the purification procedure. From these results we conclude that only two virus-specified polypeptides are present in the polymerase. A scheme for the synthesis of these polypeptides is presented in the accompanying paper.
J Gen Virol 1976 Sep
PMID:Purification and polypeptide composition of Semliki Forest virus RNA polymerase. 96 47

The avian viral agent S1133 has previously been classified serologically as a member of the avian reovirus group. This viral agent grows in chicken embryo fibroblast cells, bands at a density of 1.37 g/ml in CsCl equilibrium density gradients, has a particle diameter of 75 nm, and has a morphology similar to that of human reovirus type 3. Its nucleic acid is comprised of double-stranded RNA and adenosine-rich oligonucleotides. The dsRNA is distributed among 10 segments with molecular weights of 2.7 x 10(6), 2.6 x 10(6), 1.7 x 10(6), 1.5 x 10(6), 1.3 x 10(6), 1.2 x 10(6), 0.80 x 10(6), 0.74 x 10(6), and 0.68 x 10(6) for the largest (L1) to the smallest (S4) segment, respectively, as determined by polyacrylamide gel electrophoresis. These 10 segments migrate differently on polyacrylamide gels compared to those of human reovirus type 3. The capsid proteins of avian reovirus consist of eight species of polypeptides as determined by polyacrylamide gel electrophoresis. These are lambda1, lambda2, lambda3, mu1, mu2, sigma1, sigma2, and sigma3 with molecular weights of 140, 125, 115, 85, 72, 40, 36, and 32 x 10(3), respectively. Only polypeptide sigma2, which resides in the inner capsid or core, comigrated with the sigma2 polypeptide of type 3 reovirus. Antiserum against type 3 reovirus did not neutralize avian reovirus. Avian reovirus core particles were found to possess a transcriptase and a methylase activity.
J Virol 1976 Sep
PMID:Physical and chemical characterization of an avian reovirus. 98 52

A comparative study of the in vitro reaction kinetics of the virion RNA polymerase of influenza A strains WS and WSN was conducted to establish phenotypic differences for enzyme activity that might be exchanged as genetic markers among recombinants of these viruses. Characteristically, the RNA polymerase activity of WS virus showed an initial rate of synthesis about two- to threefold higher than that of WSN when assayed at 32 C. The two strains were also distinguishable by comparing the transcription rates of each strain at 32 and 37 C. The initial rate of WS was invariably higher at 37 than at 32 C, whereas the opposite was found with WSN. When a series of recombinants obtained from mixed infections with the WS and WSN viruses were examined for virion transcriptase activity, it was found that the two polymerase related markers behaved as properties which segregated independently of each other and of additional nonselective markers that were scored. Seven temperature-sensitive mutants of WSN virus representing distinct recombination-complementation groups were found to show a diminished transcriptase activity as compared to wild-type virus, and one of these clones (ts 24) was largely deficient for this function. None of these mutants appeared to possess a heat-liable virion polymerase.
J Virol 1975 Sep
PMID:Virion-associated transcriptase activity of influenza recombinant and mutant strains. 115 93

Temperature sensitive mutants of bacteriophage Qbeta have been isolated which fail in the synthesis of their virus RNA at the non-permissive temperature (42 degrees C). Nine mutants have been studied in some detail. Cells infected with these mutants at 37 degrees C and incubated long enough to produce substantial amounts of Qbeta RNA cease Qbeta RNA replication when shifted to 42 degrees C. The mutants can be classified into 3 groups according to the amount of Qbeta RNA replicase activity exhibited in extracts from infected cells isolated at various times after shift to 42 degrees C: in group 1 mutants, enzyme activity is the same, regardless of the time of isolation after shift; in group 2 mutants enzyme activity increases with time of isolation after shift; in group 3 mutants, enzyme activity decreases with time of isolation after shift. Synthesis of all virus proteins is suppressed at 42 degrees C in cells infected with group 2 of group 3 mutants. In cells infected with group 2 mutants, synthesis of Qbeta RNA replicase subunit beta is increased, but synthesis of other virus proteins is depressed at 42 degrees C. The inhibition of virus RNA and protein synthesis is reversible. A detailed analysis of these experiments suggests that a defective Qbeta RNA replicase is involved in the inhibition of both virus RNA and protein synthesis.
J Gen Virol 1975 Sep
PMID:Studies of temperature sensitive mutants of bacteriophage Qbeta, defective in both replication and translation. 117 68


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