Franks Caroline E, Hsu Ku-Lung
Department of Chemistry, University of Virginia, Charlottesville, Virginia.
Curr Protoc Chem Biol. 2019 Sep;11(3):e72. doi: 10.1002/cpch.72.
Human kinases are a large family of proteins (500+) that catalyze ATP-dependent phosphorylation of protein and metabolite substrates to regulate diverse facets of cell biology. Dysregulation and mutations of protein kinases are linked to human disease, providing opportunities for developing pharmacological agents as potential therapy. Assessing the selectivity of pharmacological compounds targeting this enzyme class is critical given that off-target activity of kinase inhibitor drugs may result in toxicity. This set of protocols outlines use of ATP acyl phosphate activity-based probes to evaluate the potency and selectivity of kinase inhibitors via fluorescent gel- and mass spectrometry-based detection methods. These competitive chemical proteomic assays can evaluate engagement of >200 native kinase targets directly in complex proteomes. © 2019 by John Wiley & Sons, Inc.
人类激酶是一个庞大的蛋白质家族（超过500种），可催化蛋白质和代谢物底物的ATP依赖性磷酸化，以调节细胞生物学的多个方面。蛋白激酶的失调和突变与人类疾病相关，这为开发潜在治疗药物提供了机会。鉴于激酶抑制剂药物的脱靶活性可能导致毒性，评估针对这类酶的药理化合物的选择性至关重要。本套方案概述了基于ATP酰基磷酸活性的探针的使用，通过基于荧光凝胶和质谱的检测方法来评估激酶抑制剂的效力和选择性。这些竞争性化学蛋白质组学分析可以直接在复杂蛋白质组中评估超过200种天然激酶靶点的结合情况。© 2019约翰威立父子公司版权所有
