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:2.7.1.21 (thymidine kinase)
7,561 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The previous demonstration of the efficacy and tolerability of the Oka strain of varicella-zoster virus (VZV) in clinical trials involving vaccination of both normal and immunocompromised individuals has laid the foundation for its use in preventing chickenpox. In this context, VZV could be useful as a vector for vaccinating against other infectious agents as well. As an initial application, a live recombinant VZV expressing Epstein-Barr virus (EBV) membrane glycoproteins (gp350/220) was generated by inserting a gene fusion of the VZV gpI promoter and hydrophobic leader-encoding sequence with the gp350/220 coding sequence into the thymidine kinase (TK) gene of VZV (Oka). Insertion of the foreign DNA into the thymidine kinase gene was demonstrated by Southern blot analysis and the ability of the recombinant virus to replicate in the presence of bromodeoxyuridine. RNA splicing, glycosylation, and plasma membrane presentation of gp350/220 in cells infected with the recombinant virus were similar to those seen in EBV-infected cells. In addition, the expression of VZV-specific glycoproteins was unaltered by the concomitant expression of this large foreign glycoprotein. Thus, VZV can be used as a live viral vector for active immunization against EBV and other pathogens.
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PMID:Varicella-zoster virus as a live vector for the expression of foreign genes. 303 57

A modified-live pseudorabies virus (PRV) vaccine, designated PRV(dlg92/d1tk), with deletions in the thymidine kinase (tk) and glycoprotein-gIII (g92) genes, was derived from the PRV (Bucharest [BUK]-d13) vaccine strain. The vaccine virus also contained a deletion in glycoprotein gI. Despite 3 deletions, PRV(dlg92/d1tk) replicated to high titers in cell culture from 30 C to 39.1 C. Enzyme assays and autoradiography revealed that PRV(dlg92/d1tk) did not induce a functional tk activity in infected tk- RAB(BU) cells (rabbit skin). Rabbit skin cells were infected with PRV(dlg92/d1tk), with vaccine strains derived from BUK or Bartha K strains of PRV or with the virulent Illinois (ILL), Indiana-Funkhauser (IND-F), and Aujeszky (Auj) strains of PRV and were labeled with [3H]mannose from 4 or 5 to 24 hours after infection to investigate whether these viruses induced the synthesis of glycoprotein gIII. Nonionic detergent extracts were prepared and immunoprecipitated with antisera from pigs vaccinated with tk(-)-PRV(BUK-d13) or tk+-Bartha K, pigs vaccinated with tk+-PRV(BUK) strains and then challenge exposed to tk+-PRV(IND-F), naturally infected domestic or feral pigs, and pigs vaccinated with tk-)-PRV(dlg92/d1tk). Mouse monoclonal antibodies against PRV glycoproteins gIII, gp50, and gII were also studied. After immunoprecipitation, labeled PRV-specific proteins were analyzed by sodium dodecylsulfate-polyacrylamide gel electrophoresis and autoradiography. The PRV glycoprotein-gII complex, but not glycoprotein gIII, was synthesized in PRV(dlg92/d1tk)-infected cells. Glycoprotein gII and gIII were made in cells infected with PRV vaccine strains BUK, Bartha K, and BUK-d13 and with virulent PRV strains ILL, IND-F, and Auj. Cells infected with PRV(dlg92/d1tk) and with PRV strains ILL, IND-F, Auj, Bartha K, BUK, and BUK-d13, excreted into the cell culture medium a highly sulfated glycoprotein gX of about 90 kilodaltons. Antibodies to glycoprotein gIII were not detected in the sera of pigs inoculated with PRV(dlg92/d1tk), but were found in all other swine sera.
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PMID:Second-generation pseudorabies virus vaccine with deletions in thymidine kinase and glycoprotein genes. 303 72

Human alpha interferons (IFN-a) cause a reorganization of internal cell membranes into tubuloreticular inclusions (TRI). Morphogenesis and cytochemistry indicate a pre-Golgi intracisternal origin from the endoplasmic reticulum. Clinically, TRI formation in human blood mononuclear cells correlates with systemic IFN-a treatment or with endogenous overproduction of IFN-a in viral or autoimmune diseases (e.g., rubella syndrome, AIDS, systemic lupus erythematosus). In vitro, TRI formation can be produced by treatment of Daudi lymphoblasts or vascular endothelial cells with IFN-a, and is blocked by actinomycin-D. In Daudi lymphoblasts or vascular endothelial cell cultures, TRI formation parallels induction of 2'-5' A synthetase, inhibition of thymidine kinase and growth inhibition; however, heavy water treatment of Daudi cells prevented TRI formation while induction of 2'-5' A synthetase and growth inhibition persisted. TRI formation was dissociated from IFN-a antiproliferative activity in a mutant clone of Daudi lymphoblasts. Decreased glycoprotein biosynthesis and increased phospholipid biosynthesis may accompany progressive TRI accumulation.
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PMID:Tubuloreticular reorganization of cytomembranes in cells treated with human alpha interferons--a review. 307 Jul 35

The inherited human disorders sialidosis and galactosialidosis are the result of deficiencies of glycoprotein-specific alpha-neuraminidase (acylneuraminyl hydrolase, EC 3.2.1.18; sialidase) activity. Two genes were determined to be necessary for expression of neuraminidase by using human-mouse somatic cell hybrids segregating human chromosomes. A panel of mouse RAG-human hybrid cells demonstrated a single-gene requirement for human neuraminidase and allowed assignment of this gene to the (pter----q23) region of chromosome 10. A second panel of mouse thymidine kinase (TK)-deficient LM/TK- -human hybrid cells demonstrated that human neuraminidase activity required both chromosomes 10 and 20 to be present. Analysis of human neuraminidase expression in interspecific hybrid cells or polykaryocytes formed from fusion of mouse RAG (hypoxanthine/guanine phosphoribosyltransferase deficient) or LM/TK- cell lines with human sialidosis or galactosialidosis fibroblasts indicated that the RAG cell line complemented the galactosialidosis defect, but the LM/TK- cell line did not. This eliminates the requirement for this gene in RAG-human hybrid cells and explains the different chromosome requirements of these two hybrid panels. Fusion of LM/TK- cell hybrids lacking chromosome 10 or 20 (phenotype 10+,20- and 10-,20+) and neuraminidase-deficient fibroblasts confirmed by complementation analysis that the sialidosis disorder results from a mutation on chromosome 10, presumably encoding the neuraminidase structural gene. Galactosialidosis is caused by a mutation in a second gene required for neuraminidase expression located on chromosome 20.
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PMID:Sialidosis and galactosialidosis: chromosomal assignment of two genes associated with neuraminidase-deficiency disorders. 308 2

A cloned cDNA (1.65 kb) containing the complete glycoprotein gene of the Josiah strain of Lassa virus was inserted into the thymidine kinase (TK) gene of the New York Board of Health (WYETH) strain of vaccinia virus. The Lassa virus glycoprotein precursor, GPC, and the posttranslational cleavage products, G1 and G2, were shown by Western blot analysis to be properly expressed in cells infected with the recombinant virus. Northern blot hybridization of total cytoplasmic RNA extracted from recombinant virus infected cells demonstrated the presence of RNA transcripts of appropriate size considering the site of transcription initiation from the vaccinia P7.5 promoter, the size of the Lassa glycoprotein gene, and the presumed location of the transcription terminator in the vaccinia thymidine kinase gene. All guinea pigs vaccinated with the recombinant virus survived a lethal challenge infection with Lassa virus, whereas 80% of control animals died. The vaccinated guinea pigs did, however, develop transient, low-grade, fevers and detectable viremias following infection with Lassa virus, indicating that protection was not complete.
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PMID:Construction of a recombinant vaccinia virus expressing the Lassa virus glycoprotein gene and protection of guinea pigs from a lethal Lassa virus infection. 335 60

The major glycoprotein, G, of human respiratory syncytial (RS) virus is a Mr 84,000-90,000 species that has about 60% of its mass contributed by carbohydrate, most of which is in the form of O-linked oligosaccharides. The G protein contains neither a hydrophobic N-terminal signal sequence nor a hydrophobic C-terminal anchor region. Instead, its amino acid sequence reveals only one region with significant hydrophobic character, which is between residues 38 and 66. In order to study the synthesis, processing, and functions of this unusual viral glycoprotein, full-length cDNA copies of the G protein mRNA were inserted into the DNA genome of vaccinia virus (VV) in a position that was adjacent to a strong VV promoter and within the VV gene for thymidine kinase (TK). The resulting TK- recombinant viruses were selected, plaque-purified, and characterized by Southern blot analysis of restriction enzyme digests of the viral DNA. Recombinant RNA transcripts that contained both G-specific and VV-specific sequences accumulated in cells infected with recombinant viruses having the G protein gene in the positive orientation. The translation product of these transcripts in infected cells was a Mr 84,000-90,000 glycoprotein that was indistinguishable from authentic RS virus G protein. It could be detected in cell lysates after metabolic labeling with [3H]glucosamine and was immunoprecipitated by anti-RS-virus antiserum. Immunofluorescence studies showed that the G protein accumulated intracellularly with the perinuclear distribution that is characteristic of newly synthesized glycoproteins. Furthermore, the protein was also clearly detectable on the surface of recombinant-infected cells, showing that it was transported to and inserted into the plasma membrane.
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PMID:Expression of the major glycoprotein G of human respiratory syncytial virus from recombinant vaccinia virus vectors. 345 62

Six recombinants of New York Board of Health (NYBH) vaccinia virus containing cDNA for Challenge Virus Standard (CVS) rabiesvirus glycoprotein (G) were produced by directing gene insertion into the vaccinia thymidine kinase (TK) locus. To regulate expression of G the promoter P7.5 (functions at early and late times postinfection) from the gene for the vaccinia 7.5 kilodalton (kD) protein was used in two of the recombinants; late promoter P11 of the vaccinia 11 kD protein was used in four recombinants. The six differed in nucleotide sequences flanking the translation start codon; in two constructs the encoded signal peptide of G was fused to several additional amino acids. Cells infected with each recombinant made G that reacted with G-specific antibodies, comigrated with authentic G, and was transported to the plasma membrane. The highest amounts of G were made with fusion or standard versions of G with P11 provided that the mRNA leader sequences were identical to the natural gene. Each recombinant in mice and one in dogs induced rabiesvirus neutralizing antibodies and protection against lethal rabiesvirus challenge.
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PMID:Vaccinia virus recombinants expressing rabiesvirus glycoprotein protect against rabies. 350 40

A cDNA copy of the G glycoprotein gene of human respiratory syncytial virus (RSV) was placed under control of a vaccinia virus promoter and inserted into the thymidine kinase locus of the vaccinia virus genome. The recombinant vaccinia virus retained infectivity and expressed a 93-kDa protein that migrated with the authentic RSV G glycoprotein upon polyacrylamide gel electrophoresis. Glycosylation of the expressed protein and transport to the cell surface were demonstrated in the absence of other RSV proteins. Cotton rats that were inoculated intradermally with the infectious recombinant virus produced serum antibody to the G glycoprotein that neutralized RSV in vitro. Furthermore, the vaccinated animals were resistant to lower respiratory tract infection upon intranasal inoculation with RSV and had reduced titers of RSV in the nose.
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PMID:Resistance to human respiratory syncytial virus (RSV) infection induced by immunization of cotton rats with a recombinant vaccinia virus expressing the RSV G glycoprotein. 351 91

A cDNA clone representing the mRNA coding sequence of the fusion glycoprotein (F) gene of human respiratory syncytial virus (RSV) was constructed and inserted into the thymidine kinase gene of vaccinia virus (WR strain) under the control of a vaccinia virus promoter. The resulting recombinant vaccinia virus, vaccinia F, expressed the F1 and F2 cleavage products (48 and 20 kDa, respectively) of the F glycoprotein in cell culture. F1 and F2 were indistinguishable from their authentic RSV counterparts with respect to glycosylation, disulfide linkage, electrophoretic mobility, cell-surface expression, and antigenic specificity. Cotton rats infected intradermally with vaccinia F developed a high titer of serum F-specific antibodies, which neutralized infectivity of RSV. This neutralizing antibody response exceeded that induced by infection of the respiratory tract with RSV and was 6-fold higher than that induced by vaccinia G, a recombinant vaccinia virus that expressed the RSV G glycoprotein gene. Immunization with vaccinia F stimulated almost complete resistance to replication of RSV in the lower respiratory tract as well as significant resistance in the upper respiratory tract. The degree of resistance conferred by vaccinia F exceeded that induced by vaccinia G.
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PMID:Expression of the F glycoprotein of respiratory syncytial virus by a recombinant vaccinia virus: comparison of the individual contributions of the F and G glycoproteins to host immunity. 353 15

Vaccinia virus (VV) recombinants were constructed that contained full-length cDNA copies of the fusion (F) protein gene of human respiratory syncytial (RS) virus. The F protein gene was placed next to the strong early-late VV 7.5-kilodalton promoter and was located within the VV thymidine kinase (tk) gene. Full-length recombinant transcripts that initiated at both the tk and the 7.5-kilodalton promoters accumulated in cells early in infection, and one or more of these transcripts was translated to yield a glycoprotein which comigrated with Fo, the fusion protein precursor. This precursor was processed by proteolytic cleavage to produce the two disulfide-linked subunits F1 and F2, which were both glycosylated and of the same electrophoretic mobility as authentic F1 and F2. Immunofluorescence studies demonstrated that the mature F protein was transported to and expressed on the surface of recombinant VV-infected cells. Inoculation of rabbits with a recombinant vector expressing F resulted in the production of antiserum specific for the RS virus F protein. This antiserum neutralized virus infectivity and was capable of preventing fusion in RS virus-infected cells. Mice were vaccinated with recombinants expressing the F protein. At 3 weeks postinoculation, these animals had serum antibody against RS virus F protein. At 5 days after intranasal challenge with RS virus, the lungs of the mice previously vaccinated with recombinants expressing F protein were free of detectable RS virus, whereas the lungs of unvaccinated mice contained 10(4.2) PFU of virus per g.
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PMID:Expression of the fusion protein of human respiratory syncytial virus from recombinant vaccinia virus vectors and protection of vaccinated mice. 380 89


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