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 conditions required for the production of varicella-zoster virus (VSV)-induced deoxythymidine kinase (dTk) have been studied. Extracts from Vero cells harvested 62 h after VZV infection were found to contain VZV-induced dTk activity, with a minimal contribution from the cellular dTk activity. VZV dTK was shown to have a broad substrate specificity phosphorylating both deoxythymidine, deoxycytidine, and iododeoxyuridine. Deoxythymidine triphosphate inhibition studies revealed an intermediate deoxythymidine triphosphate sensitivity when compared with that of the cellular cytosolar enzyme and the deoxythymidine triphosphate-insensitive herpes simplex virus dTk. An assay for VZV dTk-blocking antibodies was developed, with [125I]iododeoxyuridine as a substrate in the presence of a deoxythymidine triphosphate concentration which selectively blocked the dTK of host cell origin. A total of 79 serum samples were studied; these included serum pairs from patients with varicella or herpes zoster and single sera from immune and nonimmune adults. VZV dTk blocking antibodies were detected exclusively in sera from patients with herpes zoster. All serum pairs showing VZV dTK seroconversion also showed a parallel conversion of complement fixation titers. The VZV dTk antibodies were found to be of the immunoglobulin G class. The immunological specificity of VZV dTK was investigated, and no cross-reactivity with herpes simplex virus type 1 or 2 dTk was found.
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PMID:Human serum antibodies to varicella-zoster virus thymidine kinase. 617 44

Mice infected with three different isolates of herpes simplex virus (HSV) and treated with acyclovir (acycloguanosine; ACV) showed low levels of virus replication during the acute phase of infection. However, virus isolated from such treated mice did not show increased resistance to ACV. In contrast, resistant virus was readily isolated in vitro by passaging HSV in the presence of the drug. The degree of resistance was determined, in part, by the nature of the cells used to test the virus. The majority of ACV-resistant strains induced low or undetectable levels of HSV-specified thymidine kinase (TK), the enzyme responsible for phosphorylating ACV in infected cells. The TK-resistant strains were attenuated when injected into mice as indicated by reductions in virus replication, inflammation, and establishment of latent infections in sensory ganglia. The reduced virulence of the TK- strains was most marked after intracerebral inoculation, where the lethal dose was increased more than 100-fold compared with the parental isolates. However, one mutant is described which induced high levels of TK but was highly resistant to ACV and retained virulence for mice.
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PMID:Pathogenicity in mice of strains of herpes simplex virus which are resistant to acyclovir in vitro and in vivo. 624 69

When Xenopus laevis oocyte nuclei are injected with a recombinant plasmid containing the Herpes Simplex Virus (HSV) thymidine kinase (tk) gene, a 100-fold increase in tk enzymatic activity is observed. Three lines of evidence show that this increase in tk activity is a result of the expression of the HSV tk gene. First, the enzymatic activity is selectively inactivated by the IgG fraction of antiserum raised against HSV tk protein. Second, a polypeptide that comigrates with authentic HSV tk on polyacrylamide gels is synthesized uniquely by oocytes injected with the HSV tk gene. Third, the induced tk activity found in injected oocytes is capable of phosphorylating deoxycytidine, a substrate that is utilized by HSV tk but not by cellular tk. We have used these observations to establish an assay for examining the activity of mutated variants of the HSV tk gene. Two sets of deletion mutants of the tk gene were constructed in vitro. In one set varying amounts of 5' flanking and intragenic sequences are deleted. The other set is deleted at the 3' end of the gene. By testing the activity of each mutant in the oocyte injection assay we have delimited functional boundaries corresponding to the 5' and 3' termini of the HSV tk gene.
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PMID:Expression of the herpes thymidine kinase gene in Xenopus laevis oocytes: an assay for the study of deletion mutants constructed in vitro. 625 55

Acyclovir reduced mortality and organ virus titers in mice inoculated intraperitoneally with 10 50% lethal doses of mouse cytomegalovirus. This susceptibility to acyclovir of a herpesvirus which lacks thymidine kinase is surprising. Alternative phosphorylating enzymes may account for this susceptibility.
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PMID:Efficacy of acyclovir against mouse cytomegalovirus in vivo. 626 90

The replication of unselected strains of herpesvirus saimiri (HVS) was sensitive to bromodeoxyuridine and bromovinyldeoxyuridine (BVdU) but insensitive to acycloguanosine (ACG), in contrast to the growth of herpes simplex virus (HSV) which was sensitive to all three analogues. Mutants of HVS resistant to bromodeoxyuridine and BVdU could be selected by growth in the presence of these inhibitors. Productive infections of owl monkey kidney or Vero cell cultures by unselected strains of HVS resulted in increases in a thymidine kinase (TK) activity which was deficient in cells infected with bromodeoxyuridine-resistant mutants of the virus. Induction of the virus enzyme promoted a net increase in the uptake and incorporation of exogenous labelled thymidine in the face of the progressive inhibition of the overall incorporation of [35S]methionine and [3H]uridine into productively infected cells. The TK induced in cells infected with HVS differed from the major activity of uninfected cells and resembled that encoded by HSV in its capacity to phosphorylate iododeoxyuridine and in the sensitivity of all the thymidine phosphorylating activity to competition by BVdU. However, in contrast to the HSV TK, which phosphorylated deoxycytidine and iododeoxycytidine relatively efficiently and was sensitive to ACG, the HVS enzyme did not phosphorylate deoxycytidine or iododeoxycytidine and was insensitive to ACG. Whilst HVS, therefore, shares the characteristic of other members of the herpesvirus group of inducing a novel TK, the properties of the HVS-induced enzyme differ significantly from the enzyme of the prototype herpesvirus, HSV. The properties of the HVS TK are nonetheless sufficiently distinct from those of the uninfected cell to provide a possible basis for selective antiviral chemotherapy based on preferential phosphorylation of nucleoside analogues such as BVdU by infected cells.
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PMID:Comparison of thymidine kinase activities indiced in cells productively infected with herpesvirus saimiri and herpes simplex virus. 627 57

Acyclovir, an acrylic purine nucleoside analog, is a highly potent inhibitor of herpes simplex virus (HSV), types 1 and 2, and varicella zoster virus, and has extremely low toxicity for the normal host cells. This selectivity is due to the ability of these viruses to code for a viral thymidine kinase capable of phosphorylating acyclovir to a monophosphate; this capability is essentially absent in uninfected cells. The acyclovir monophosphate (acyclo-GMP) is subsequently converted to acyclovir triphosphate (acyclo-GTP) by cellular enzymes. Acyclo-GTP persists in HSV-infected cells for many hours after acyclovir is removed from the medium. The amounts of acyclo-GTP formed in HSV-infected cells are 40 to 100 times greater than in uninfected Vero cells. Acyclo-GTP acts as a more potent inhibitor of the viral DNA polymerases than of the cellular polymerases. The DNA polymerases of HSV-1 and HSV-2 also use acyclo-GTP as a substrate and incorporate acyclo-GMP into the DNA primer-template to a much greater extent than do the cellular enzymes. The viral DNA polymerase binds strongly to the acyclo-GMP-terminated template, and in thereby inactivated.
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PMID:Mechanism of action and selectivity of acyclovir. 628 36

Three mutant strains, one conditional, of Tetrahymena thermophila were defective in thymidine phosphorylating activity in vivo and in thymidine kinase activity in vitro. Nucleoside phosphotransferase activity in mutant cell extracts approached wild-type levels, suggesting that thymidine kinase is responsible for most, if not all, thymidine phosphorylation in vivo. Thymidine kinase activity in extracts of the conditional mutant strain was deficient when the cells were grown or assayed or both at the permissive temperature, implying a structural enzyme defect. Analysis of the reaction products from in vitro assays with partially purified enzymes showed that phosphorylation by thymidine kinase and nucleoside phosphotransferase occurred at the 5' position. Genetic analyses showed that the mutant phenotype was recessive and that mutations in each of the three mutant strains did not complement, suggesting allelism.
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PMID:Mutant strains of Tetrahymena thermophila defective in thymidine kinase activity: biochemical and genetic characterization. 629 Aug 73

The specific activity of thymidine kinase (TK) was higher in spleen than in thymus or unseparated tonsillar lymphocytes, while deoxycytidine kinase (dCK) specific activity was lowest in spleen and was much higher in thymus and in unseparated tonsillar lymphocytes. The ratio of dCK to TK was always high in thymus, in unseparated and in B-cell-enriched tonsillar lymphocytes (between 2 and 5), but it was always low in spleen (0.3-0.4). The difference in the pyrimidine nucleoside phosphorylating enzyme activities of the thymus and spleen does not seem to be a mere consequence of different DNA synthesis rates, because the activities of DNA polymerase-alpha were practically the same in these organs. Unseparated and B-cell-enriched tonsillar lymphocytes resemble the thymus with respect to the ratio of dCK to TK activities, while the T-cell-enriched fraction contained 3-5 times lower activities of both enzymes. These results suggest that the metabolic pathways of CdR and TdR utilization for DNA synthesis differ in the lymphocyte populations independently from their rate of DNA polymerization and they may be in connection with their maturation processes.
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PMID:Differences between lymphoid organs with respect to the phosphorylation of deoxycytidine and thymidine. 630 28

Acyclovir [9-(2-hydroxyethoxymethyl)guanine] (ACV), a potent antiviral compound, was phosphorylated to the same extent by extracts from untreated and iododeoxyuridine-treated Epstein-Barr virus-containing latent D98/HR-1 somatic hybrid cells. ATP was the preferred phosphate donor over other nucleoside triphosphates. The cytosol extract from D98/HR-1 cells effected optimum phosphorylation of thymidine at pH 8.0, whereas ACV was phosphorylated equally well over a wide pH range. Electrophoretic analysis of thymidine kinase-, deoxycytidine kinase-, and ACV-phosphorylating activities from both untreated and iododeoxyuridine-treated cell extracts displayed identical properties. A small part (5 to 10%) of the loaded ACV-phosphorylating activity seemed to migrate with the deoxycytidine kinase activity from cytosol. dTTP and dCTP, at relatively high concentrations, partially inhibited ACV-phosphorylating activity. The results suggest that Epstein-Barr virus does not code for its own thymidine kinase and that phosphorylation of ACV in Epstein-Barr virus-producing cells is carried out by multiple or as yet unidentified ATP-dependent nonspecific cellular phosphotransferases.
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PMID:Phosphorylation of acyclovir in vitro in activated Burkitt somatic cell hybrids. 631 70

The metabolism of 9-(1,3-dihydroxy-2-propoxymethyl)guanine (DHPG), one of the most promising new anti-herpes virus compounds, in HeLa cells infected with herpes simplex virus type 1 was compared with that in the uninfected HeLa cells. In the virus-infected cells, the uptake of DHPG was enhanced and the major metabolites were found to be the mono-, di-, and triphosphate derivatives. The formation of these metabolites was dependent on the extracellular concentration of DHPG (0.5 to 5.0 microM). Virus-induced thymidine kinase was capable of phosphorylating DHPG to its monophosphate which could be further phosphorylated to the di- and triphosphate derivatives by the host cellular enzymes. Incorporation of the DHPG into DNA was observed in virus-infected cells. In contrast with 9-(2-hydroxyethoxymethyl)guanine, DHPG seemed not to serve as a chain terminator, but to be incorporated internally into DNA strands.
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PMID:Metabolism of 9-(1,3-dihydroxy-2-propoxymethyl)guanine, a new anti-herpes virus compound, in herpes simplex virus-infected cells. 631 60


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