Duong-Ly Krisna C, Peterson Jeffrey R
Cancer Biology Program, Fox Chase Cancer Center, 333 Cottman Avenue, Philadelphia, PA, 19111, USA.
Methods Mol Biol. 2016;1360:87-95. doi: 10.1007/978-1-4939-3073-9_7.
Aberrant kinase signaling has been implicated in a number of diseases. While kinases have become attractive drug targets, only a small fraction of human protein kinases have validated inhibitors. Screening of libraries of compounds against a kinase or kinases of interest is routinely performed during kinase inhibitor development to identify promising scaffolds for a particular target and to identify kinase targets for compounds of interest. Screening of more focused compound libraries may also be conducted in the later stages of inhibitor development to improve potency and optimize selectivity. The dot blot kinase assay is a robust, high-throughput kinase assay that can be used to screen a number of small-molecule compounds against one kinase of interest or several kinases. Here, a protocol for a dot blot kinase assay used for measuring insulin receptor kinase activity is presented. This protocol can be readily adapted for use with other protein kinases.
异常的激酶信号传导与多种疾病有关。虽然激酶已成为有吸引力的药物靶点，但只有一小部分人类蛋白激酶有经过验证的抑制剂。在激酶抑制剂开发过程中，常规会针对一种或多种感兴趣的激酶对化合物库进行筛选，以识别针对特定靶点的有前景的骨架结构，并识别感兴趣化合物的激酶靶点。在抑制剂开发的后期阶段，也可能会进行更有针对性的化合物库筛选，以提高效力并优化选择性。斑点印迹激酶测定法是一种强大的高通量激酶测定法，可用于针对一种感兴趣的激酶或几种激酶筛选多种小分子化合物。在此，介绍一种用于测量胰岛素受体激酶活性的斑点印迹激酶测定法的方案。该方案可轻松适用于其他蛋白激酶。
