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)

Measurements of the deoxyribonucleoside triphosphate (dNTP) contents, the [14C] thymidine and deoxyuridine incorporation and the "key enzymes" of the thymidine triphosphate (dTTP) synthesis, thymidine kinase and ribonucleotide reductase, in diploid Ehrlich-ascites carcinoma, in Yoshida sarcoma-ascites cells and to a smaller extent in surgically removed malignant human tumours show 1. A distinctly increased dTTP content compared with the remaining dNTP is not a characteristic of tumour cells generally but a peculiarity of sarcoma and a sign of differentiation of a malignant tumour. 2. With simultaneous linear deoxyribonucleoside incorporation the dTTP content and the mix-proportion of [14C] dTTP to total dTTP in ascites tumour cells in short-term in-vitro incubation (120 min) remain constant. 3. Thymidine addition to the medium leads to a distinct rise of dTTP concentration even at a dosage of 3 X 10(-5) M. 4. The dNTP contents of ascites tumour cells are within the range of the endproduct-inhibiting concentrations of thymidine kinase and ribonucleotide reductase.
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PMID:[Studies on the thymidine-triphosphate synthesis in malignant tumors. I. Effects of thymidine on deoxyribonucleoside triphosphate pools and deoxyribonucleic acid synthesis (author's transl)]. 13 36

Deoxythymidine kinase activities were induced in HeLa TK- (deoxythymidine kinase-deficient) cells infected with either herpes simplex virus type I or herpes simplex virus type II. The herpes simplex virus type I-induced enzyme was found in the cytoplasmic and nuclear fractions of the infected cells, whereas the herpes simplex type II-induced deoxythymidine kinase could only be found in the cytoplasm. Herpes simplex virus type I and II specific deoxythymidine kinases were purified by affinity column chromatography. Both purified deoxythymidine kinases retained the deoxycytidine kinase activity present in the crude preparation. The purified herpes simplex virus type I deoxythymidine kinase had a different mobility on electrophoresis, but the same sedimentation rate on a glycerol gradient as the corresponding unpurified enzyme, whereas the purified herpes simplex virus type II deoxythymidine kinase had the same mobility and sedimentation rate as the corresponding unpurified enzyme. In the presence of Mg2+ATP and dithiothreitol, herpes simplex virus type II deoxythymidine kinase was more stable than herpes simplex virus type I deoxythymidine kinase at both 45 degrees and 4 degrees. The deoxycytidine kinase activity present in the purified preparations was inactivated at the same rate as the deoxythymidine kinase activity. In the presence of the other substrate, deoxythymidine, herpes simplex virus type I deoxythymidine kinase was more stable than herpes simplex virus type II kinase. The purified herpes simplex virus type I and II deoxythymidine kinase had different activation energies when Mg2+ATP and deoxythymidine were used as substrates, but showed the same sensitivity toward ammonium sulfate inhibition.
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PMID:Deoxythymidine kinase induced in HeLa TK- cells by herpes simplex virus type I and type II. II. Purification and characterization. 17 18

1. Extracts of several plant species contained nucleoside-AMP phosphotransferase activity. The ratio of activity with thymidine to that with uridine as nucleoside substrate was essentially constant, both between species and throughout plant development. Evidence is presented that the total thymidine-AMP phosphotransferase activity of the leaves of Asplenium nidus (bird's-nest fern) and of Helianthus tuberosus (Jerusalem artichoke) increases during maturation. 2. Thymidine-AMP phosphotransferase was purified 22-fold from a very rich source of this activity, extracts of A. nidus. 3. A broad specificity towards both nucleoside and nucleoside 5'-monophosphate substrates is displayed by this preparation, and the evidence suggests that all could be due to a single enzyme. 4. Nucleosides that act as substrates will also inhibit phosphotransfer to other nucleosides, with Ki values close to the corresponding Km values found when utilized as substrates. 5. Ca2+-activated ATP phosphohydrolase was separated from the phosphotransferase by differential complexing to Blue Dextran in the presence of urea, whereas an AMP phosphohydrolase activity was closely associated with thymidine-AMP phosphotransferase through all separation techniques used. 6. Metal ions did not activate either of the latter two activities, and 1,10-phenanthroline was found to inhibit the phosphotransferase. 7. Km values for AMP for the respective activities were 0.11 mM (thymidine phosphotransferase) and 0.20 mM (AMP phosphohydrolase) and for thymidine (phosphotransferase only) 0.88 mM. 8. 3':5'-Cyclic AMP was found to inhibit both phosphotransferase and AMP phosphohydrolase activities, with Ki values of 0.056 mM and 0.15 mM respectively. It is suggested that this inhibitor would be of value in revealing the existence of thymidine kinase in plant extracts with high thymidine phosphotransferase activity.
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PMID:Thymidine phosphotransferase and nucleotide phosphohydrolase of the fern Asplenium nidus. General properties and inhibition by adenosine 3':5'-cyclic monophosphate. 18 31

Deoxythymidine kinases (EC 2.7.1.--) induced in HeLa TK- cells by Herpes simplex Type I and Type II viruses both had a requirement for divalent cations. The enzymes had the highest activities in the presence of Mg2+, followed by Mn2+, Ca2+, Fe2+, and in that order, whereas they were inactive in the presence of Zn2+ and Cu2+. The amount of Mg2+ required for optimal activity was dependent on the amount of ATP present, so that optimal activities were found when the concentration of Mg2+ was equal to that of ATP; an excess of Mg2+ inhibited the reaction. The activities of various nucleoside triphosphates as phosphate donors for Herpes simplex virus Type I deoxythymidine kinase were in the order: ATP = dATP = ara ATP greater than CTP greater than dCTP greater than UTP greater than dUTP greater than GTP greater than dGTP. Those for Herpes simplex virus Type II deoxythymidine kinase were in the order: CTP greater than dCTP = ara CTP greater than dATP greater than ATP greater than UTP greater than GTP greater than dUTP = dGTP. For both deoxythymidine kinases induced by Herpes simplex virus, the nucleoside triphosphates tested exerted cooperative effects. The Km values of ATP and CTP for the Herpes simplex virus Type I enzyme were 30 and 70 muM respectively; whereas those for the Herpes simplex virus Typr II enzyme were 140 and 450 muM. Studies on binding of various thymidine analogs with free 5'-OH to these deoxythymidine kinases indicated that 5-substituted ethyl-, vinyl-, allyl-, propyl-, iodo- and bromo-dUrd as well as iodo5 dCyd and bromo5 dCyd had good affinity to both enzymes. In contrast, vinyl5 Urd, iodo5 Urd and arabinosylthymidine had good affinity only to the Herpes simplex virus Type I enzyme but not to the Herpes simplex virus Type II deoxythymidine kinase. All of these thymidine analogs were competitive inhibitors, with KI values in the range of 0.25 to 1.5 muM. Herpes simplex virus Type I deoxythymidine kinase was less sensitive to either dTTP or iodo dUTP inhibition than Herpes simplex virus Type II. Both dThd and dCyd could serve as substrates and competed with each other for Herpes simplex viruses Type I and Type II induced kinases, but they differed in their Km values for these enzymes. The Km values of dThd and dCyd were 0.59 muM and 25 muM for Herpes simplex virus Type I deoxythymidine kinase; while they were 0.36 muM and 88 muM respectively for the Herpes simplex virus Type II enzyme.
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PMID:Deoxythymidine kinase induced in the HELA TK- cells by herpes simplex virus type I and type II. Substrate specificity and kinetic behavior. 18 65

Malignant cells have enhanced sensitivity to inhibition of growth by thymidine. Cell growth of the permanent lymphoid cell line CCRF-CEM, originating from a patient with acute lymphoblastic leukemia, is inhibited by 3 x 10(-5) M thymidine, compared to 1 to 5 x 10(-3) M thymidine required to inhibit growth of normal lymphoid lines. Thymidine-resistant cells were isolated at a frequency of approximately 1/100,000 cells after cloning CCRF-CEM cells in medium containing 5 x 10(-4) M thymidine. The resistant cells lacked the enzyme thymidine kinase, had a 20-fold decrease of thymidine uptake, and were resistant to 1 x 10(-4) M 5-bromo-2-deoxyuridine. The cells were sensitive to 1 x 10(-5) M methotrexate, even in the presence of exogenously added thymidine and hypoxanthine. The data indicate that a small fraction of malignant cells may escape the toxic effect of high thymidine therapy and therefore, require additional chemotherapy for their control.
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PMID:Isolation of thymidine-resistant cells from a thymidine-sensitive acute lymphoblastic leukemia cell line. 28 39

The rate of DNA synthesis is exponentially growing cells was determined by isotopedilution analysis of the incorporation of [me-3H]thymidine. Thymidine concentrations greater than 7 micrometer were used so that the rate-limiting step governing incorporation would be at the level of DNA polymerase rather than at the level of thymidine kinase [Sjostrom & Forsdyke (1974) Biochem. J. 138, 253-262]. In early exponential phase the rate determined by isotope-dilution analysis closely correlated with the rates calculated either from growth curves or from known cell-cycle parameters. However, in late-exponential phase the rate calculated from the growth curve was less than that determined by isotope-dilution analysis. We conclude that, under certain conditions, the pool-corrected rate of incorporation of [me-3H]thymidine, as determined by isotope-dilution analysis, can accurately reflect the rate of DNA synthesis. Discrepancies between the observed rate of DNA synthesis and increase in cell number could reflect an exponential degeneration of post-S-phase cells.
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PMID:The rate of deoxyribonucleic acid synthesis by cultured Chinese-hamster ovary cells. An application of isotope-dilution analysis. 34 2

We have examined the interaction of dexamethasone with the ZR75-1 human breast cancer cell line to determine if glucocorticoids might directly inhibit growth of breast cancer cells. Growth of these cells in serum-free medium was stimulated significantly by physiological concentrations of insulin (0.1 to 1.0 nM). Pharmacological concentrations of dexamethasone (10 nM) reduced cell number below that found in controls and nearly abolished the effect of insulin after several days in culture. Thymidine and uridine, but not leucine, incorporation into macromolecules or acetate incorporation into fatty acids were similarly inhibited by dexamethasone in the presence of absence of insulin. Dexamethasone did not inhibit insulin effects by altering insulin receptor affinity or concentration, as determined by Scatchard analyses of insulin binding. Net thymidine uptake into the trichloroacetic acid-soluble fraction of the cell was stimulated by insulin and inhibited by dexamethasone also inhibited thymidine kinase activity multiple potential sites of glucocorticoid action that directly oppose the effects of insulin. They also suggest that glucocorticoids have a direct inhibitory effect on proliferation of human breast cancer cells, which may help explain breast tumor regression following pharmacological glucocorticoid therapy.
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PMID:Direct inhibition of growth and antagonism of insulin action by glucocorticoids in human breast cancer cells in culture. 44 41

A line of mouse 3T3 cells lacking deoxythymidine kinase (dTK-) was stably transformed to see dTK+ phenotype after exposure to ultraviolet-irradiated equine herpesvirus type 1 (EHV-1). Deoxythymidine kinase (dTK) was purified from the biochemically transformed mouse cells by affinity chromatography on deoxythymidine-Sepharose. The purified dTK from EHV-1-transformed 3T3 cells was identical to the dTK purified from dTK- 3T3 cells lytically infected with EHV-1 with respect to its electrophoretic mobility, molecular weight, substrate specificity, phosphate donor specificity, and immunological specificity. The sedimentation velocity of the purified dTK from the transformed 3T3 cells was similar to that previously reported for the enzyme in lytically infected dTK- 3T3 cells, and its molecular weight was estimated to be 87,000. Antiserum prepared against the EHV-1 dTK induced in horse cells inactivated the dTK purified from the transformed mouse cells. The Km for deoxythymidine (5 micrometers) of purified dTK from the EHV-1-transformed cells was the same as that reported for the EHV-1-induced dTK. These results further support the notion that the dTK acquired by dTK- mouse 3T3 cells after transformation by EHV-1 is of viral and not of cellular origin.
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PMID:Purification and biochemical characterization of deoxythymidine kinase of deoxythymidine kinase-deficient mouse 3T3 cells biochemically transformed by equine herpesvirus type 1. 48 23

Thymidine transport was studied in isolated rat hepatocytes. In these cells no phosphorylation of the substrate by thymidine kinase occurred subsequent to transport. Results from studies of the concentration-dependent uptake of thymidine indicated two transport systems with about 80-fold differences in their kinetic constants. These systems were denoted as high affinity [Km = 5.3 micron, V = 0.47 pmol/(10(6) cells X s)] and low affinity systems [Km = 480 micron, V = 37.6 pmol/(10(6) cells X s)]. From intracellular to extracellular distribution ratios of [3H]thymidine it could be concluded that the uptake by the high affinity system was a concentrative process while the transport by the low affinity system was non-concentrative. The uptake of [3H]-thymidine by the high affinity system could only be inhibited by unlabeled thymidine. In contrast, all other nucleosides tested (uridine, 2'-deoxycytidine, and 2'-deoxyguanosine) were equally effective in inhibiting the low affinity system competitively. The results would suggest that in hepatocytes lacking phosphorylation by thymidine kinase, thymidine is taken up by a high and a low affinity system working in tandem. The high affinity system seems to be an active transport process with narrow substrate specificity. Thymidine uptake by the low affinity system is a facilitated diffusion process. This system is considered to be a common transport route for nucleosides of different structures.
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PMID:Uptake of thymidine into isolated rat hepatocytes. Evidence for two transport systems. 68 Jun 44

Thymidine kinase has been measured in phytohaemagglutinin (PHA)-stimulated lymphocytes from 13 normal subjects and eight patients with megaloblastic anaemia. The levels in normal subjects ranged from 0.20 to 2.10 units/mg protein (mean 0.903 units/mg protein) and in megaloblastic anaemia from 2.99 to 9.97 units/mg protein). All the patients showed raised levels of the enzyme which were partly but not completely reduced to normal by addition of folic acid in vitro. Vitamin B12 in vitro had a lowering effect in the five vitamin-B12-deficient patients and two patients with combined deficiencies but not in one 'pure' folate-deficient patient. Thymidine kinase activity was highest in the cells of the least anaemic patients, suggesting that the degree of anaemia in megaloblastic anaemia may be determined in part by the ability of the cells to utilize thymidine by the 'salvage' pathway when the de novo pathway of thymidylate synthesis is failing. The rise in thymidine kinase activity in megaloblastic anaemia is presumably due to induction of the enzyme. Addition of methotrexate or 5-fluorouracil, drugs known to inhibit de novo thymidylate synthesis, caused an increase in thymidine kinase activity in normal PHA-stimulated lymphocytes after 24 h (but not after 1 h) which could be completely blocked by addition of puromycin. Thymidine mono- and di-phosphate kinases were also measured in normal PHA-stimulated lymphocytes. The activities were substantially higher than that of thymidine kinase and their activities were unaffected by methotrexate addition.
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PMID:Thymidine kinase in megaloblastic anaemia. 100 25


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