Csapó Z, Sasvári-Székely M, Spasokoukotskaja T, Talianidis I, Eriksson S, Staub M
Department of Medical Chemistry, Molecular Biology and Pathobiochemistry, Semmelweis University, H-1444 8, Budapest, Hungary.
Biochem Pharmacol. 2001 Jan 15;61(2):191-7. doi: 10.1016/s0006-2952(00)00534-7.
Deoxycytidine kinase (dCK, EC.2.7.1.74) is a key enzyme in the intracellular metabolism of 2-chlorodeoxyadenosine, 1-beta-D-arabinofuranosylcytosine, difluorodeoxycytidine, and other drugs used in chemotherapy of different leukaemias and solid tumours. Recently, stimulation of dCK activity was shown by these analogues and by other genotoxic agents such as etoposide and NaF, all of which cause severe inhibition of DNA synthesis in cell cultures. Here we describe that direct inhibition of DNA polymerases by aphidicolin stimulated dCK activity in normal lymphocytes and acute myeloid leukaemic cells, as well as in HL 60 promyelocytic cell cultures. Increased dCK activity was not due to new protein synthesis under our conditions, as measured by immunoblotting. Partial purification by diethylaminoethyl-Sephadex chromatography revealed that the activated form of dCK survived purification procedure. Moreover, it was possible to inactivate purified dCK preparations by recombinant protein phosphatase with Ser/Thr/Tyr dephosphorylating activity. These data suggest that the activation of dCK may be due to phosphorylation, and that deoxynucleoside salvage is promoted during inhibition of DNA synthesis in human lymphocytes.
脱氧胞苷激酶（dCK，EC.2.7.1.74）是2-氯脱氧腺苷、1-β-D-阿拉伯呋喃糖基胞嘧啶、二氟脱氧胞苷以及其他用于治疗不同白血病和实体瘤的化疗药物细胞内代谢的关键酶。最近，这些类似物以及其他遗传毒性剂如依托泊苷和氟化钠均显示出对dCK活性的刺激作用，所有这些物质在细胞培养中均会导致DNA合成的严重抑制。在此我们描述，在正常淋巴细胞、急性髓性白血病细胞以及HL 60早幼粒细胞培养物中，阿非科林对DNA聚合酶的直接抑制作用刺激了dCK活性。如通过免疫印迹法所测，在我们的实验条件下，dCK活性的增加并非由于新蛋白质的合成。通过二乙氨基乙基-葡聚糖凝胶色谱法进行部分纯化显示，dCK的活化形式在纯化过程中得以保留。此外，具有丝氨酸/苏氨酸/酪氨酸去磷酸化活性的重组蛋白磷酸酶能够使纯化的dCK制剂失活。这些数据表明，dCK的激活可能是由于磷酸化作用，并且在人淋巴细胞DNA合成受抑制期间，脱氧核苷补救途径得到了促进。
