A novel allosteric site in casein kinase 2α discovered using combining bioinformatics and biochemistry methods
A novel allosteric site in casein kinase 2α discovered using combining bioinformatics and biochemistry methods
复制标题
结合生物信息学和生物化学方法发现酪蛋白激酶 2α 中的一个新的变构位点。
DOI：
10.1038/aps.2017.55
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发表时间：
2017-12-01
影响因子：
8.2
通讯作者：
Zhang, Jian
中科院分区：
文献类型：
--
作者：
Jiang, Hai-ming;Dong, Jiang-kai;Zhang, Jian
酪蛋白激酶2（Casein kinase 2，CK 2）是一种高度多效性的丝氨酸-苏氨酸激酶，可催化300多种蛋白质的磷酸化，参与信号转导、转录调控、细胞凋亡和细胞周期等多种细胞功能的调控。另一方面，CK 2在多种肿瘤中异常升高，被认为是有希望的治疗靶点。然而，目前可用的ATP竞争性CK 2抑制剂缺乏选择性，这阻碍了它们在癌症治疗中的发展。由于变构抑制剂可以避免传统激酶抑制剂的缺点，本研究旨在发现CK 2 α中新的变构位点，并研究该位点突变对CK 2 α活性的影响。使用基于蛋白质动力学和结构排列的Allosite，我们预测了一个新的变构位点，该位点部分位于CK 2 α的α C螺旋中。对暴露在该位点表面上的五个残基进行突变以验证预测。通过改变肽底物的浓度，使用发光ADP检测测定进行动力学分析，结果显示突变1780和178 W降低CK 2 α活性，而V31 R、K75 E、1820和P1090增加CK 2 α活性。使用蒙特卡罗路径生成方法鉴定了潜在的变构途径，并且这些预测的变构途径的结果与突变分析一致。使用ClustaIX方法进行CK 2 α与该家族中其他激酶的多重序列比对，其揭示了该位点中残基的多样性。总之，我们确定了一个新的变构位点CK 2 α，可以改变调节激酶的活性。由于该位点中残基的高度多样性，可以使用针对生物相关性的特定CK 2 α抑制剂的合理药物设计来靶向该位点。
Casein kinase 2 (CK2) is a highly pleiotropic serine-threonine kinase, which catalyzed phosphorylation of more than 300 proteins that are implicated in regulation of many cellular functions, such as signal transduction, transcriptional control, apoptosis and the cell cycle. On the other hand, CK2 is abnormally elevated in a variety of tumors, and is considered as a promising therapeutic target. The currently available ATP-competitive CK2 inhibitors, however, lack selectivity, which has impeded their development in cancer therapy. Because allosteric inhibitors can avoid the shortcomings of conventional kinase inhibitors, this study was aimed to discover a new allosteric site in CK2 alpha and to investigate the effects of mutations in this site on the activity of CK2 alpha. Using Allosite based on protein dynamics and structural alignment, we predicted a new allosteric site that was partly located in the alpha C helix of CK2 alpha. Five residues exposed on the surface of this site were mutated to validate the prediction. Kinetic analyses were performed using a luminescent ADP detection assay by varying the concentrations of a peptide substrate, and the results showed that the mutations 1780 and 178W decreased CK2 alpha activity, whereas V31R, K75E, 1820 and P1090 increased CK2 alpha activity. Potential allosteric pathways were identified using the Monte Carlo path generation approach, and the results of these predicted allosteric pathways were consistent with the mutation analysis. Multiple sequence alignments of CK2 alpha with the other kinases in the family were conducted using the ClustaIX method, which revealed the diversity of the residues in the site. In conclusion, we identified a new allosteric site in CK2 alpha that can be altered to modulate the activity of the kinase. Because of the high diversity of the residues in the site, the site can be targeted using rational drug design of specific CK2 alpha inhibitors for biological relevance.