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:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Sodium p-chloromercuribenzoate (PCMB) caused a noticeable reduction of infectivity of prototype strains of type A and Lee strain of type B influenza viruses at concentrations of 100 and 200 mug/ml, respectively, after an incubation at 37 C for 60 min. The virucidal effect on A/AA/2/60 (H2N2) strain was dependent on the concentration of the drug and temperature as well as on the time of incubation. The reagent exerted this effect at a concentration which induced little change in the hemagglutinating and neuraminidase activities of the virus. PCMB inhibited by 50% the virus particle-associated RNA polymerase activity of all prototype strains of type A influenza virus at about 2 mug/ml and that of Lee strain of type B influenza virus at 8.5 mug/ml. Other sulfhydryl reagent such as phenylmercuric nitrate also exhibited virucidal effect on A/AA/2/60 virus which paralleled their inhibition of the virus particle-associated RNA polymerase activity. From these results it was considered likely that the virucidal action of PCMB on influenza viruses was attributable to inhibition of the virus particle-associated RNA polymerase activity.
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PMID:Virucidal effect of sodium p-chloromercuribenzoate on influenza viruses attributable to inhibition of virus particle-associated RNA-dependent RNA polymerase. 116 Feb 1

Cloned cDNA encoding the Sendai virus (SV) hemagglutinin-neuraminidase (HN) envelope glycoprotein was expressed in cultured cells in two ways: (I) infection with HN-expressing recombinant vaccinia virus, or (II) transfection with a plasmid with T7 promoter and termination sequences flanking the HN gene, with intracellular T7 RNA polymerase supplied by coinfection with recombinant vaccinia virus that expresses the enzyme. The HN expressed was indistinguishable from the authentic SV protein in antigenicity, cell surface location, and formation of oligomeric structures. In addition, HN expressed from cDNA functioned normally in both hemadsorption and neuraminidase activities. The usefulness of cDNA expression for analyzing HN structure and function was evaluated by mutating the HN cDNA and observing the consequences for HN protein activity. Since previous work indicated that the lysine residue at position 461 is important for the neuraminidase activity of HN, we used site-directed mutation to produce HN protein with this lysine residue changed to glutamic acid. The mutated HN had neuraminidase activity with significantly increased thermal stability, indicating that residue 461 may be essential to the protein's conformation.
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PMID:Expression of cDNA encoding the Sendai virus hemagglutinin-neuraminidase gene: characterization of wild-type and mutant gene products. 131 81

The complete nucleotide sequence of the simian virus 41 (SV41) large (L) protein gene was determined. The L gene spanned 6883 nucleotides including a putative trailer RNA, and the L mRNA contained a single large open reading frame encoding a polypeptide of 2269 amino acids. Dot-matrix comparisons under stringent conditions identified domains highly conserved among paramyxoviruses. Domain 3 is the most highly conserved, and has been hypothesized to be the RNA polymerase active site. A phylogenetic tree was constructed from the sequences of the L proteins of seven paramyxoviruses. SV41 was most closely related to human parainfluenza virus type 2 (HPIV-2), and SV41, HPIV-2 and SV5 form a subgroup. The intergenic sequences at the nucleocapsid protein-phosphoprotein and haemagglutinin-neuraminidase-L protein gene junctions, and the 5' trailer sequence of SV41 were also determined, and it was shown that the first 13 nucleotides of the 5' trailer sequence are complementary to those of the 3' leader sequence. The intergenic, and gene-start and -end sequences of SV41, HPIV-2 and SV5 are shown.
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PMID:Nucleotide sequence analysis of the simian virus 41 gene encoding the large (L) protein and construction of a phylogenetic tree for the L proteins of paramyxoviruses. 132 85

We succeeded in rescuing infectious influenza virus by transfecting cells with RNAs derived from specific recombinant DNAs. RNA corresponding to the neuraminidase (NA) gene of influenza A/WSN/33 (WSN) virus was transcribed in vitro from plasmid DNA and, following the addition of purified influenza virus RNA polymerase complex, was transfected into MDBK cells. Superinfection with helper virus lacking the WSN NA gene resulted in the release of virus containing the WSN NA gene. We then introduced five point mutations into the WSN NA gene by cassette mutagenesis of the plasmid DNA. Sequence analysis of the rescued virus revealed that the genome contained all five mutations present in the mutated plasmid. The ability to create viruses with site-specific mutations will allow the engineering of influenza viruses with defined biological properties.
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PMID:Introduction of site-specific mutations into the genome of influenza virus. 233 22

Sixteen temperature-sensitive mutants of Sendai virus were isolated from mutagenized stocks (10 mutants, designated numerically) and persistently infected cultures (6 mutants, designated alphabetically). Based on complementation tests, virion-associated activities, thermal inactivation, and viral RNA and hemadsorbing antigen synthesis as well as virion production in chick lung embryo cells at nonpermissive temperature, these mutants were divided into seven groups as follows. i) HANA group mutants (ts-5, -9, -10, -201), defective in hemagglutinin-neuraminidase protein, complementation group I. ii) F group mutants (ts-18, -108), defective in hemolytic and cell-fusing activity, complementation group II. iii) Ts-43, defective in RNA polymerase activity, complementation group III. iv) Ts-23, defective in RNA polymerase activity, interfered with the other mutants in complementation tests. v) Ts-25, defective in the incorporation of hemagglutinin-neuraminidase protein into the virion at the stage of virus assembly. vi) Ts-110, belongs to F group mutants on one hand, but is considered to carry another undetermined defect. vii) C group (carrier culture-borne group) mutants (ts-a, -b, -c, -d, -e, -f), defective lesion not yet determined and belong to neither complementation group I nor II. Assignment of mutants in groups iv), v), vi), and vii) to complementation groups could not be achieved.
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PMID:Isolation and characterization of temperature-sensitive mutants of Sendai virus. 626 Oct 90

A systematic and comparative study was performed on the polypeptide composition and the RNA polymerase activity associated with virions of various strains of influenza A virus, including four human and two avian viruses. Significant differences were found in the molecular weights of not only hemagglutinin (HA) but also both nucleoprotein (NP) and membrane protein (M), as determined by polyacrylamide gel electrophoresis under denaturing conditions. The results indicate that, among viruses sharing the same serotype determined by the surface proteins HA and NA (neuraminidase), considerable variations exist in the structure of viral proteins, including inner proteins. The relative contents of viral proteins also varied among these strains grown under similar conditions. The total content of three P proteins, the putative RNA polymerase subunits, was within the range between 1.1 and 2.2% of total viral proteins and roughly paralleled the virion-associated RNA polymerase activity. The virion-associated RNA polymerase of all the strains tested were stimulated by the same dinucleotide primers, ApG or GpG, indicating that the specificity of transcription initiation is conserved among wide varieties of influenza virus.
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PMID:RNA polymerase of influenza virus. I. Comparison of the virion-associated RNA polymerase activity of various strains of influenza virus. 627 72

Two paramyxovirus isolates recovered from the peripheral blood leukocytes of a patient with subacute sclerosing panencephalitis were characterized by biological, immunological, and molecular techniques. Both virus isolates possessed neuraminidase activity but demonstrated reduced hemagglutination potential. Neutralization titrations showed that the viruses were clearly related to, but were distinct from, simian virus 5. Characterization of purified virus preparations by SDS-PAGE showed that the structural polypeptides of the viruses were similar to those of simian virus 5, but distinct differences were noted as well. In addition, the virus isolates were found to differ from one another, particularly with regard to the major putative transcriptase protein (L).
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PMID:Characterization of nonmeasles paramyxovirus isolates from a patient with subacute sclerosing panencephalitis. 733 93

Influenza viruses are spherical, about 1000 A in diameter, and consist of an as yet undefined central structure containing the eight negative-sense RNA molecules of the genome (1) in association with the transcriptase required for mRNA synthesis, an abundant nucleoprotein, and an equally abundant matrix protein. This core is surrounded by a membrane derived from the cell surface in a budding process by which newly formed viruses are released from the infected cell. During infection cell membranes are modified by the incorporation of newly synthesized virus membrane proteins, and the finally released viruses contain exclusively two different types of virus-specified glycoprotein, hemagglutinin and neuraminidase, and a proton channel protein, M2. All three of these molecules have been studied extensively, particularly the glycoproteins, and in this paper information on their structures and functions will be summarized and related to modifications in cellular membranes that occur during virus infection.
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PMID:Influenza viruses and cell membranes. 755 5

Cleavage activation of the Sendai virus (Fushimi strain) fusion (F) protein was analyzed by site-directed mutagenesis of the amino acids proximal to the highly conserved fusion peptide. In addition, the functional properties of the wild-type and mutant proteins were examined to determine their ability to elicit the formation of syncytia when co-expressed with the hemagglutinin-neuraminidase (HN) glycoprotein. Viral genes were expressed from recombinant T7 transcription vectors (pT7/T3 plasmids) containing F or HN genes, after transfection into cells previously infected with a recombinant vaccinia virus expressing T7 RNA polymerase (vTF7-3). The wild-type F protein sequence (112VPQSRF) which contains a monobasic cleavage activation site was altered to include a tribasic, 112VPRKRF (mB3), or a pentabasic sequence, 112RRRKRF (mB5) adjacent to the fusion peptide. Although addition of basic residues to the normal protein sequence resulted in enhanced cleavage activation of the mB3 and mB5 proteins, only the mB5 protein was able to induce syncytia formation in CV-1 or HeLa T4 cells. Further analysis by the introduction of acidic residues upstream of the cleavage activation site was performed to determine whether increased hydrophilicity of the surrounding residues might contribute to cleavage activation. The mutants examined, mAcB1 (104NDDEENAGVPQSRF), mAcB3 (104NDDEENAGVPRKRF), and mAcB5 (104NDDEENAGRRRKRF) all contained DEE in replacement for the wild-type TTQ sequence (104NDTTQNAGVPQSRF). Analysis showed that only mAcB3 was efficiently cleaved by the endogenous cellular proteases, while mAcB1 was minimally cleaved, and mAcB5 not at all. Consequently, only the mAcB3 mutant was able to support fusion of CV-1 or HeLa T4 cells when co-expressed with HN.
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PMID:Role of basic residues in the proteolytic activation of Sendai virus fusion glycoprotein. 762 24

To study the contributions of the hemagglutinin-neuraminidase (HN) and the fusion (F) glycoproteins in virus-induced membrane fusion, the HN and F proteins of human parainfluenza virus type-1 (hPIV-1) and Sendai virus (SV) were expressed in HeLa T4+ cells using the vaccinia virus-T7 RNA polymerase transient expression system. Expression of F protein alone did not induce cell fusion. However, coexpression of homologous F and HN proteins resulted in extensive syncytium formation by hPIV-1 or SV glycoproteins, which supports the proposal that both the F and HN glycoproteins are necessary for membrane fusion. To investigate the function of HN in membrane fusion, we coexpressed heterologous combinations of the HN and F proteins of hPIV-1 and SV. No fusion was observed when SV HN and hPIV-1 F proteins were coexpressed. In contrast, the coexpression of hPIV-1 HN and SV F induced extensive cell fusion. These results suggest that specific interaction between HN and F is required to induce membrane fusion. To locate regions that are essential to the fusion promoting activity, chimeric HN proteins of SV and hPIV-1 were constructed. The chimeric proteins coexpressed with the SV or hPIV-1 F proteins indicated that some regions in the middle 62% of HN contribute to the fusion-promoting activity. To determine the role of the transmembrane region of HN on fusion-promoting activity, mutant HN proteins were expressed and their biological activities examined. Mutation of hPIV-1 HN at residue 55 from cysteine to tryptophan did not affect cell binding, neuraminidase activities, or homooligomer formation, but did result in the loss of cell fusion activity. The mutation of the same cysteine residue to glycine retained the fusion-promoting activity, suggesting that a sulfhydryl moiety is not specifically required at position 55, but the structure of the residue that occupies the position is important in fusion-promoting activity.
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PMID:Regions on the hemagglutinin-neuraminidase proteins of human parainfluenza virus type-1 and Sendai virus important for membrane fusion. 794 17


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