A chip-based screening system for IκB kinase β (IKKβ) has been developed by physically immobilizing the substrate IκBα on a glass matrix using a calixarene linker. Phosphorylation of IκBα by IKKβ and ATP was quantitated using a fluorescently labeled antibody. Using this efficient assay system a chemical library of 2,000 bioactive compounds was screened against IKKβ and four were identified as good inhibitors, namely, aurintricarboxylic acid, diosmin, ellagic acid, and hematein. None of them have been reported to be an inhibitor of IKKβ although they were implicated in various NFκB-mediated biological processes. Our enzyme-based assay showed that IC50 of the four inhibitors is comparable with that of IKK-16, a previously known strong inhibitor. Molecular docking simulation shows that the hydrophobic moiety of an inhibitor interacts with the four hydrophobic residues (Leu21, Val29, Val152, and Ile165) of the active site. The MM-PBSA calculation suggests that these hydrophobic interactions appear to be the predominant contributor to the binding free energy. As IKKβ is ubiquitously expressed in various cell types and executes many biological functions, the enzyme and cell specificity of the four inhibitors need to be rigorously tested before accepted as a drug candidate.

通过使用杯芳烃接头将底物IκBα物理固定在玻璃基质上，已开发出基于芯片的IκB激酶β（IKKβ）筛选系统。使用荧光标记的抗体定量IKKβ和ATP对IκBα的磷酸化。使用这一高效的测定系统，针对IKKβ筛选了2,000种生物活性化合物的化学文库，并确定了四种良好的抑制剂，即金精三羧酸，薯os皂素，鞣花酸和血凝素。尽管它们与各种NFκB介导的生物学过程有关，但没有报道它们是IKKβ抑制剂。我们的基于酶的测定表明，四种抑制剂的IC50与以前已知的强抑制剂IKK-16相当。分子对接模拟显示抑制剂的疏水部分与活性位点的四个疏水残基（Leu21，Val29，Val152和Ile165）相互作用。MM-PBSA计算表明，这些疏水相互作用似乎是结合自由能的主要贡献者。由于IKKβ在各种细胞类型中普遍表达并且具有许多生物学功能，因此在被接受为候选药物之前，必须对四种抑制剂的酶和细胞特异性进行严格测试。