Johansson M, Karlsson A
Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, Sweden.
Biochem Pharmacol. 1995 Jul 17;50(2):163-8. doi: 10.1016/0006-2952(95)00129-n.
Human and mouse deoxycytidine kinase (dCK) (EC 2.7.1.74) were cloned and expressed in Escherichia coli. Michaelis-Menten kinetics were determined for the purified enzymes with 2'-deoxycytidine (dCyd), 2'-deoxyadenosine (dAdo), 2-chloro-2'-deoxyadenosine (CdA), 2',3'-dideoxycytidine (ddCyd) and 9-beta-D-arabinofuranosylguanine (araG) as substrates and ATP and UTP as phosphate donors. Both human and mouse dCK showed highest affinity to dCyd with Km values of 0.05-0.2 microM. The anti-leukaemic compound CdA was the superior substrate of the nucleoside analogues tested. Both enzymes were able to efficiently utilize ATP and UTP as phosphate donors. However, the use of UTP instead of ATP as phosphate donor decreased Km values for all substrates investigated. The kinetic properties of mouse and human dCK differed in that the human enzyme showed higher affinity for the substrates dAdo, CdA, ddCyd and araG. The human enzyme also showed higher affinity for ATP and UTP. The ability to phosphorylate dCyd was, however, similar for both human and mouse dCK. At physiological concentration of the feedback inhibitor dCTP, mouse dCK showed lower activity than human dCK for all substrates investigated.
人源和鼠源脱氧胞苷激酶（dCK）（EC 2.7.1.74）被克隆并在大肠杆菌中表达。以2'-脱氧胞苷（dCyd）、2'-脱氧腺苷（dAdo）、2-氯-2'-脱氧腺苷（CdA）、2'，3'-双脱氧胞苷（ddCyd）和9-β-D-阿拉伯呋喃糖基鸟嘌呤（araG）为底物，ATP和UTP为磷酸供体，测定了纯化酶的米氏动力学。人源和鼠源dCK对dCyd的亲和力最高，Km值为0.05 - 0.2微摩尔。抗白血病化合物CdA是所测试的核苷类似物中的优质底物。两种酶都能够有效地利用ATP和UTP作为磷酸供体。然而，用UTP代替ATP作为磷酸供体降低了所有研究底物的Km值。鼠源和人源dCK的动力学性质有所不同，人源酶对底物dAdo、CdA、ddCyd和araG表现出更高的亲和力。人源酶对ATP和UTP也表现出更高的亲和力。然而，人源和鼠源dCK磷酸化dCyd的能力相似。在反馈抑制剂dCTP的生理浓度下，对于所有研究的底物，鼠源dCK的活性均低于人源dCK。
