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)

4'-Azidothymidine (ADRT) is a novel nucleoside analog, that selectively inhibits human immunodeficiency virus replication in human lymphocytes. Unlike the dideoxyribonucleoside analogs and 3'-azido-2',3'-dideoxythymidine (AZT), ADRT retains the 3'-hydroxy group. The pathways of ADRT metabolism were elucidated by determining: (i) the kinetics of the interactions of ADRT and its metabolites with enzymes of thymidine metabolic pathways, (ii) the pool sizes of phosphorylated metabolites, and (iii) the nature of ADRT incorporation into human DNA. ADRT is not a substrate for thymidine phosphorylase, but is metabolized by kinases. Thymidine kinase phosphorylates ADRT to ADRT monophosphate (ADRT-MP). For this enzyme, ADRT has a Ki value of 5.2 microM, in comparison to a Km value of 0.7 microM for thymidine. The Km value of ADRT toward thymidine kinase is 8.3 microM and the rate of ADRT phosphorylation is 1.4% that of thymidine phosphorylation. ADRT-MP has a low affinity toward thymidylate kinase (a Ki value of 28.9 microM versus a Km value of 0.56 microM for thymidylate), and toward thymidylate synthase (a Ki value of 180 microM versus a Km value of 8 microM for deoxyuridylate). The results suggest that ADRT can be activated effectively by cellular kinases without significant interference of normal thymidine metabolism. In cultured human lymphocytes (A3.01, H9, and U937 cells), ADRT was phosphorylated efficiently to ADRT 5'-triphosphate (ADRT-TP), which is the major metabolite of ADRT. The intracellular concentrations of ADRT-TP ranged from 1 to 3.3 microM after 24 h of incubation with 2 microM of ADRT and the half-life of ADRT-TP varied from 3 to 6 h. Although ADRT-TP is a poor competitive inhibitor against dTTP toward DNA polymerases alpha and beta with Ki values of 62.5 and 150 microM, respectively. ADRT-MP was found to be internally incorporated into cellular DNA. The extent of ADRT-MP substitution for dTMP in DNA was 1 in 6979 for A3.01 cells incubated with 2.9 microM ADRT for 24 h. Internal incorporation of ADRT-MP contrasts with the mechanism of other 2',3'-dideoxynucleoside analogs (i.e. AZT, ddC, ddI, d4T...), which are DNA chain terminators. This finding indicates that a 3'-deoxy structure in a nucleoside analog is not a prerequisite for anti-human immunodeficiency virus activity.
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PMID:Metabolism of 4'-azidothymidine. A compound with potent and selective activity against the human immunodeficiency virus. 173 May 94

Thymidine kinase (EC 2.7.1.21) from regenerating rat liver has been purified 70,000-fold to apparent homogeneity by affinity chromatography. Molecular weight of the native enzyme was found to be about 54,000, as determined by gel filtration. Electrophoresis in polyacrylamide gels containing sodium dodecyl sulfate yielded a single band with a molecular weight of 26,000, suggesting that thymidine kinase is a dimer of very similar or identical subunits. The Michaelis constant for thymidine is 2.2 microM. ATP acts as a sigmoidal substrate with a 'Km' of 0.2 mM. Reaction kinetics and product inhibition studies reveal the enzymatic mechanism to be sequential.
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PMID:Purification and characterization of thymidine kinase from regenerating rat liver. 191 61

Thymidine kinase was isolated and purified 1600-fold from sea urchin (Strongylocentrotus intermedius) oocytes. The molecular mass of the enzyme is 66 kDa, pI is 5.2. The enzyme activity needs Mg2+, ATP and the corresponding phosphate acceptor. The pH optimum of the enzyme activity is at 9.0-9.5. The isolated enzyme does not exhibit any strict substrate specificity and can phosphorylate thymidine, deoxycytidine and some other pyrimidine nucleosides and their derivatives, the phosphorylation rate being maximal for thymidine, ATP or dATP. The TMP formed via the enzymatic reaction does not influence the thymidine kinase activity.
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PMID:[Thymidine kinase from sea urchin oocytes]. 196 86

Thymidine kinase is an enzyme involved in DNA precursor metabolism and DNA replication. The synthesis of this enzyme is highly regulated during the cell cycle and the activity of the enzyme is also regulated by feedback inhibition. Genes encoding thymidine kinase have been extremely useful as selectable markers for introducing DNA into a number of cells. In order to study cell cycle regulation of thymidine kinase, the gene which encodes this enzyme, as well as aspects of DNA replication in the ciliated protozoan Tetrahymena thermophila, we have purified thymidine kinase from Tetrahymena. Two forms of thymidine kinase with native molecular masses of 59 kDa and 80 kDa have been identified and purified 6800- and 4600-fold, respectively. The 59-kDa enzyme, a homodimer of 30-kDa subunits, has been purified to near homogeneity and polyclonal antibodies have been raised against the 30-kDa subunit. Serological studies indicate that the two enzymes are antigenically distinct. The antibody against the Tetrahymena protein cross-reacts with a polypeptide in Chinese hamster ovary (CHO) cell extracts of 26 kDa which corresponds to the reported size of Chinese hamster thymidine kinase protein.
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PMID:Thymidine kinase from Tetrahymena thermophila. Purification and immunological analysis. 199 76

1. An affinity column for the purification of thymidine kinase (TK) from the cestode Hymenolepis diminuta is described. Using an epoxy-activated Sepharose 6B affinity column containing thymidine as a ligand, a 698-fold purification of thymidine kinase was obtained. 2. Thymidine kinase eluted from this affinity column was partially characterized as having an apparent Km value of 3.94 microM thymidine. This value is very similar to those observed in mammalian systems. 3. Thymidine kinase appears to be an extremely active and ubiquitous enzyme, whose primary function is to rapidly phosphorylate incoming thymidine and thus "trap" it for the cell's use, reducing efflux to a minimum. 4. The apparent Km for TK is two orders of magnitude lower than the Kt for thymidine transport. Thus, theories postulating that long-term (2 min) uptake kinetics for thymidine actually represent subsequent metabolism must look further along the thymidine phosphorylating pathway, beyond TK and its very active role.
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PMID:Independent characterization of thymidine transport and subsequent metabolism in Hymenolepis diminuta--II. Purification and preliminary analysis of thymidine kinase. 206 Feb 79

Concanavalin A-activated lymphocytes were made polyamine deficient by treatment with alpha-difluoromethylornithine and ethylglyoxal bis(guanylhydrazone). Thymidine kinase activity in polyamine-deficient cells was 17% of the level in normal cells. Thymidine kinase mRNA increased with time after concanavalin A activation and reached a maximum at 36 h after concanavalin A addition. The amount of thymidine kinase mRNA in polyamine-deficient cells was approximately 75% of that in normal cells. The transcription of thymidine kinase gene in isolated nuclei of polyamine-deficient cells was also 75% of that from normal cells. The turnover rate of thymidine kinase mRNA in both normal and polyamine-deficient cells was nearly equal. In normal cells, 95% of thymidine kinase mRNA was polysome associated, while in polyamine-deficient cells, 60% of the mRNA was polysome associated. In addition, the size of polysomes associated with thymidine kinase mRNA in polyamine-deficient cells was smaller than that in normal cells. Synthesis of thymidine kinase was stimulated approximately seven-fold by 0.3 mM spermidine in a rabbit reticulocyte polyamine-free protein synthetic system. The half-life of thymidine kinase activity in both normal and polyamine-deficient cells was nearly equal. Thymidine kinase activity was not influenced significantly by 0.3 mM spermidine. These combined results suggested that the synthesis of thymidine kinase was mainly regulated by polyamines at the level of translation.
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PMID:Polyamine regulation of the synthesis of thymidine kinase in bovine lymphocytes. 210 6

Thymidine kinase enzymes isolated from a variety of sources are generally considered to have a native molecular weight of 80-90 kDa composed of two 40-45 kDa subunits. Although these parameters may accurately describe the atypical deoxypyrimidine kinases expressed by members of the Herpesviridae, the nucleotide sequences of thymidine kinase genes isolated from human, mouse, chicken and variety of poxviruses (vaccinia virus, monkeypox virus, variola virus, fowlpox virus and capripoxvirus) predict molecular weights on the order of 20-25 kDa for the derived primary translation products. To resolve this apparent dilemma, velocity sedimentation centrifugation, gel filtration chromatography and protein cross-linking procedures were employed to provide experimental evidence that enzymatically-active vaccinia virus thymidine kinase is a homotetrameric complex of 20 kDa monomers with a native Mr of 80 kDa.
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PMID:Quaternary structure of vaccinia virus thymidine kinase. 211 4

The enzyme thymidine kinase is associated with DNA synthesis. Thymidine kinase serum levels were studied in normal controls (n = 20), patients with primary breast cancer (n = 60), patients with systemic breast cancer (n = 20) and as a non-cancer disease control group in patients with inflammatory gastrointestinal disorders (n = 20). Comparison of pretreatment values in the cancer patients with the normal controls showed a significant difference between the three groups in relation to stage of disease: mean values 4.22 (+/- 1.08), 6.22 (+/- 2.24) and 9.79 (+/- 7.56) pmol ml-1 h-1 for normal controls, operable breast cancer and systemic breast cancer respectively (P less than 0.005; analysis of variance). Patients with systemic breast cancer had a significantly elevated serum thymidine kinase level compared to controls (P less than 0.01) and patients with primary operable cancer (P less than 0.05). Patients with primary operable cancer had significantly higher serum thymidine kinase levels over normal controls (P less than 0.01). Mean serum TK in patients with inflammatory gastrointestinal diseases was similar to normal controls but significantly less than both patients with primary operable breast cancer and patients with systemic breast cancer. Twenty patients with operable breast cancer were followed up after primary surgery by serial 3-monthly thymidine kinase levels in the disease free interval. Four patients have developed systemic recurrence with a rise in the mean thymidine kinase value to 14.3 pmol ml-1 h-1. Ten patients with advanced breast cancer had serial thymidine kinase levels measured 2-monthly during the first 6 months of primary hormone therapy. The serum values fell in all five responders (mean 9.12-4.78 pmol ml-1 h-1) and rose in all five progressors (mean 8.62-38.5 pmol ml-1 h-1). Serum thymidine kinase reflects stage of disease in breast cancer. Serial thymidine kinase levels in patients with systemic breast cancer reflected response to systemic therapy.
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PMID:Thymidine kinase in breast cancer. 222 87

We have measured by a radioenzymatic assay the thymidine kinase in the cytosol of 182 primary infiltrating breast cancers. Maximal follow-up is 95 months. Thymidine kinase was found to be related to SBR grade, tumour size and absence of oestradiol receptors (RE). Univariate analysis has pointed out a significant linkage between overall or metastase free survival and thymidine kinase, using a cut-off level of 80 mU/mg protein which is the most discriminating value. Thymidine kinase appeared to be particularly useful in lymph-node-positive, RE-negative and grade 3 patients. Multivariate analysis of the overall survival and of the metastase free survival (Cox model) revealed that they were strongly related to thymidine kinase status.
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PMID:[Prognostic value of thymidine kinase in cancer of the breast]. 224 17

The effect of chlordecone (CD) on hepatic repair, measured either as recovery of microsomal enzymatic functions or as the induction of cytosolic thymidine kinase (TK) activity, was evaluated in rats given carbon tetrachloride (CCl4). Carbon tetrachloride was administered to CD-potentiated and control animals using doses of this hepatotoxin which produce similar degrees of damage at 24 hours in both groups of animals (6 and 100 microliters CCl4 per 100 g body weight, respectively). Chlordecone had no significant effect on the time course of recovery of microsomal cytochrome P-450 content or glucose-6-phosphatase activity following CCl4 administration. Hepatic TK activity was measured 48 hours after CCl4 administration as a biochemical index of the hepatic regenerative response. Thymidine kinase activity was increased eightfold in CD-treated rats receiving 6 microliters CCl4 per 100 g body weight, whereas in controls a similar induction of TK activity was produced by 100 microliters CCl4 per 100 g body weight. Therefore, the TK response in CD-treated rats receiving CCl4 is appropriate for the amount of damage produced, suggesting that CD does not inhibit the hepatic regenerative response to CCl4-induced injury. The effect of CD on hepatic repair was also examined in rats receiving a two-thirds partial hepatectomy. Pretreatment of animals with CD had no significant effect on the increase in TK activity produced 24 hours after partial hepatectomy. These results offer no support for the idea that CD impairs hepatic repair after either partial hepatectomy or CCl4 administration.
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PMID:Chlordecone does not interfere with hepatic repair after carbon tetrachloride or partial hepatectomy. 246 95


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