Identification of novel p53-binding proteins by biomolecular interaction analysis combined with tandem mass Spectrometry
Identification of novel p53-binding proteins by biomolecular interaction analysis combined with tandem mass Spectrometry
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DOI：
10.1385/mb:23:3:203
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发表时间：
2003-03-01
影响因子：
2.6
通讯作者：
Hayashi, N
中科院分区：
文献类型：
--
作者：
Kikuchi, J;Furukawa, Y;Hayashi, N
将电喷雾串联质谱（ESI-MS/MS）与生物分子相互作用分析（BIA）相结合，建立了一种实时分析蛋白质-蛋白质相互作用后直接鉴定蛋白质的方法。利用这种称为BIA-MS/MS的方法，我们从人胎盘和肝脏的全组织提取物中检测到多种p53相互作用蛋白。肽序列分析显示，与p53的相互作用，以前没有报道的三种蛋白质：细胞周期蛋白依赖性激酶抑制剂p57/Kip 2，丝氨酸/苏氨酸蛋白磷酸酶PPIC，和血红蛋白。使用我们的系统，明确的序列信息可以在毫微微到皮摩尔的水平后，重复恢复过程五次。此外，结合和解离常数很容易通过动力学分析确定。该系统为以简单但高度特异性和灵敏度的方式分析复杂的生物材料提供了强大的工具。
Electrospray tandem mass spectrometry (ESI-MS/MS) was combined with biomolecular interaction analysis (BIA) to develop a method of direct protein identification after real-time analysis of protein-protein interactions. Using this,,method, called BIA-MS/MS, we detected multiple p53-interacting proteins in whole tissue extracts from human placenta and liver. Peptide sequencing revealed three proteins whose interaction with p53 had not been previously reported: a cyclin-dependent kinase inhibitor p57/Kip2, a serine/threonine protein phosphatase PPIC, and hemoglobin. Using our system, unambiguous sequence information can be obtained at the femto- to picomole level after repeating the recovery procedure five times. Furthermore, the association and dissociation constants are easily determined by kinetic analysis. This system provides a powerful tool for analyzing complex biological materials in a simple but highly specific and sensitive manner.