Sarno Stefania, de Moliner Erika, Ruzzene Maria, Pagano Mario A, Battistutta Roberto, Bain Jenny, Fabbro Doriano, Schoepfer Joseph, Elliott Matthew, Furet Pascal, Meggio Flavio, Zanotti Giuseppe, Pinna Lorenzo A
Venetian Institute for Molecular Medicine (VIMM), University of Padova, 35121 Padova, Italy.
Biochem J. 2003 Sep 15;374(Pt 3):639-46. doi: 10.1042/BJ20030674.
IQA [[5-oxo-5,6-dihydro-indolo(1,2-a)quinazolin-7-yl]acetic acid] is a novel ATP/GTP site-directed inhibitor of CK2 ('casein kinase 2'), a pleiotropic and constitutively active protein kinase whose activity is abnormally high in transformed cells. The K (i) value of IQA (0.17 microM) is lower than those of other CK2 inhibitors reported so far. Tested at 10 microM concentration in the presence of 100 microM ATP, IQA almost suppresses CK2 activity in vitro, whereas it is ineffective or weakly effective on a panel of 44 protein kinases and on phosphoinositide 3-kinase. In comparison, other CK2 inhibitors, notably apigenin and quercetin, are more promiscuous. The in vivo efficacy of IQA has been assessed by using the fact that treatment of Jurkat cells with IQA inhibits endogenous CK2 in a dose-dependent manner. IQA has been co-crystallized with maize CK2alpha, which is >70% identical with its human homologue, and the structure of the complex has been determined at 1.68 A (1 A=0.1 nm) resolution. The inhibitor lies in the same plane occupied by the purine moiety of ATP with its more hydrophobic side facing the hinge region. Major contributions to the interaction are provided by hydrophobic forces and non-polar interactions involving the aromatic portion of the inhibitor and the hydrophobic residues surrounding the ATP-binding pocket, with special reference to the side chains of V53 (Val53), I66, M163 and I174. Consequently, mutants of human CK2alpha in which either V66 (the homologue of maize CK2alpha I66) or I174 is replaced by alanine are considerably less sensitive to IQA inhibition when compared with wild-type. These results provide new tools for deciphering the enigmatic role of CK2 in living cells and may pave the way for the development of drugs depending on CK2 activity.
IQA [[5-氧代-5,6-二氢吲哚并(1,2-a)喹唑啉-7-基]乙酸]是一种新型的针对酪蛋白激酶2（CK2）的ATP/GTP位点定向抑制剂，CK2是一种多效性且组成型激活的蛋白激酶，其活性在转化细胞中异常高。IQA的K（i）值（0.17微摩尔）低于迄今报道的其他CK2抑制剂。在100微摩尔ATP存在下，以10微摩尔浓度进行测试时，IQA几乎能在体外抑制CK2活性，而它对一组44种蛋白激酶和磷酸肌醇3-激酶无效或效果微弱。相比之下，其他CK2抑制剂，尤其是芹菜素和槲皮素，具有更强的混杂性。利用IQA处理Jurkat细胞能以剂量依赖方式抑制内源性CK2这一事实，评估了IQA的体内功效。IQA已与玉米CK2α共结晶，玉米CK2α与其人类同源物的同源性超过70%，并且复合物的结构已在1.68埃（1埃 = 0.1纳米）分辨率下确定。抑制剂位于ATP嘌呤部分所占据的同一平面内，其更疏水的一侧面向铰链区。相互作用的主要贡献来自疏水力和涉及抑制剂芳香部分以及ATP结合口袋周围疏水残基的非极性相互作用，特别是V53（缬氨酸53）、I66、M163和I174的侧链。因此，与野生型相比，人类CK2α中V66（玉米CK2α I66的同源物）或I174被丙氨酸取代的突变体对IQA抑制的敏感性显著降低。这些结果为解读CK2在活细胞中的神秘作用提供了新工具，并可能为基于CK2活性的药物开发铺平道路。
