5-氯苯并三唑对重组人蛋白激酶CK2全酶活性的影响

Effect of 5-chlorobenzotriazole on recombinant human protein kinase CK2 holoenzyme

目的:观察5-氯苯并三唑对重组人蛋白激酶CK2全酶的直接作用及其酶动力学机制。方法:在体外等物质的量混合CK2α’和β亚基构成重组CK2全酶。以重组人CK2全酶为分子靶点,通过测定转移到CK2底物上的γ-32PATP的32P放射活度来检测不同浓度5-氯苯并三唑对CK2活性的影响,并进行动力学分析。结果:0、1、3、9、27、81及243μmol/L的5-氯苯并三唑对重组人CK2全酶均具有抑制作用,且该作用随浓度的增加而增强(F=165.552,P<0.001),IC50为20.10μmol/L;它与ATP呈竞争性抑制CK2的活性,抑制常数Ki为18.63μmol/L;与酪蛋白呈非竞争性抑制CK2的活性,抑制常数Ki为22.58μmol/L。结论:5-氯苯并三唑是重组人蛋白激酶CK2的抑制剂。

Aim：To observe the direct effect of 5-chlorobenzotriazole on recombinant protein CK2 holoenzyme and its kinetics.Methods：The CK2 α＇and β subunits were mixed at the same molar ratio and reconstituted CK2 holoenzyme.The recombinant human protein kinase CK2 holoenzyme was used as a molecular target.The CK2 activity and its kinetic analysis were assayed by detecting incorporation of 32P of [γ-32P]ATP into the substrate in the various conditions.Results：5chlorobenzotriazole at 0,1,3,9,27,81 and 243 μmol/L inhibited the holoenzyme activity of recombinant human protein kinase CK2,and the effect strengthened with the increase of the concentration（F = 165.552,P 0.001）.IC50 = 20.10 μmol/L.Kinetic studies of 5-chlorobenzotriazole on CK2 showed that the inhibition was competitive with ATP with the Ki of 18.63 μmol/L and noncompetitive with casein with the Ki of 22.58 μmol/L.Conclusion：5-chlorobenzotriazole is an effective inhibitor of CK2 holoenzyme.