色谱2011，Vol.29Issue(8)：706-711,6.DOI:10.3724/SP.J.1123.2011.00706
蛋白质组衍生肽段文库用于鉴定酪蛋白激酶2的底物模体
Proteomic peptide library for determination of substrate motif of casein kinase 2
摘要
Abstract
Substrate motif is of great importance in kinasc -substrate recognition and contributes a lot to the understanding of signal transduction. In this study, a proteomic peptide library was generated to determine substrate motif of casein kinase 2. Firstly, whole cell lysate was digested by trypsin to generate a large number of candidate peptides, which were then incubated with alkaline phosphotase to dephosphorylate intrinsic phosphopeptides. Then, the unphospho-rylated peptide mixture was incubated with casein kinase 2 (CK2) and adenosine-triphosphate (ATP) for 30 min, and the resulting phosphopeptides were enriched by immobilized metal affinity chromatography (IMAC) followed by reversed-phase liquid chromatographic separation coupled with tandem mass spectrometry detection. Finally, the 472 unique phosphopeptides and 451 unique phosphorylation sites were identified, resulting in the determination of a motif of S/T-D/E-x-D/E for CK2. This method enables accurate determination of substrate motif in a short time and can be readily applied to other kinases.关键词
固定化金属离子亲和色谱/反相液相色谱-串联质谱/模体/激酶/肽段文库/蛋白质组学Key words
immobilized metal affinity chromatography (IMAC)/ reversed-phase liquid chro-matography-tandem mass spectrometry (RPLC-MS/MS)/ motif/ kinase/ peptide library/ pro-teomics分类
化学化工引用本文复制引用
张曼玉,王春丽,边阳阳,程凯,卫晓鸾,叶明亮,邹汉法..蛋白质组衍生肽段文库用于鉴定酪蛋白激酶2的底物模体[J].色谱,2011,29(8):706-711,6.基金项目
国家自然科学基金创新研究群体科学基金项目(No.21021004) （No.21021004）
