免疫共沉淀联合质谱技术筛选柔嫩艾美耳球虫钙依赖蛋白激酶3互作蛋白

IDENTIFICATION OF ETCDPK 3 INTERACTING PROTEINS BY CO-IMMUNOPRE CIPITATION IN COMBINATION WITH MASS SPECTROMETRY

为探究柔嫩艾美耳球虫钙依赖蛋白激酶3(EtCDPK3)在钙离子信号通路中是如何将信号传递给下游效应分子,进一步促进虫体入侵宿主细胞的机制,以柔嫩艾美耳球虫子孢子蛋白及其单克隆抗体和多克隆抗体为实验材料,利用免疫共沉淀技术筛选EtCDPK3的互作蛋白并进行质谱鉴定分析,使用Maxquant软件对获得的原始数据进行分析。SDS-PAGE结果显示,与阴性对照组相比,2个样品组出现差异条带;免疫共沉淀洗脱液质谱鉴定分析结果显示,获得6个可能与EtCDPK3相互作用的相关蛋白。以上实验结果为进一步深入研究EtCDPK3在柔嫩艾美耳球虫入侵宿主细胞过程中的作用奠定基础。

In order to study the mechanism of Eimeria tenella calcium protein-dependent kinase 3 (EtCDPK3) for signal transition to downstream effector molecules during the invasion of host cells,interacting proteins of EtCDPK3 were identified from the proteins of sporozoites by co-immunoprecipitation using monoclonal and polyclonal antibodies.The resulting products were examined by mass spectrometry and data were analyzed using Maxquant.Different protein bands from the two antibody groups were visualized in SDS-PAGE compared with the negative control group.Results showed that six potential EtCDPK3 interacting proteins were observed by preliminary screening.These results provided the foundation for further research on the role of EtCDPK3 in the invasion of host cells byE.tenella.