苯并[a]芘作用下ADP-核糖基化修饰蛋白分析方法的建立  

Establishment a method for identification of the poly(ADP-ribose)binding proteins induced by benzo(a) pyrene

作　　者：高玮[1] 黄海燕[2] 蔡剑锋[2] 李绚[2] 刘银品 刘建军[2] 庄志雄[2] 陈雯[1] 

机构地区：[1]中山大学公共卫生学院,广州510080 [2]深圳市疾病预防控制中心

出　　处：《卫生研究》2015年第2期196-200,共5页Journal of Hygiene Research

基　　金：国家自然科学基金(No.81001261,81370080);深圳市科技研发资金项目(No.JCYJ20130329103949642);深圳市科技计划项目(No.201302238);深圳市未来产业专项资金(No.ZDSYS20140509101335476)

摘　　要：目的建立苯并[a]芘(B[a]P)作用下ADP-核糖基化修饰蛋白分析方法。方法使用免疫共沉淀技术收集可能发生ADP-核糖基化修饰的蛋白,同步使用高效液相色谱技术和双向电泳技术对蛋白进行分离,再进行MALDI-TOF-MS/MS鉴定,比较两种分离方法鉴定蛋白的差异,并将鉴定蛋白的氨基酸序列与文献报道的ADP-核糖基化修饰蛋白的保守序列进行比对。结果通过高效液相色谱分离结合质谱鉴定找出3个蛋白,双向电泳分离结合质谱鉴定找出12个蛋白,双向电泳分离结合质谱鉴定方法存在明显优势;所鉴定蛋白存在与文献报道的ADP-核糖基化修饰蛋白相似的保守序列。结论免疫共沉淀蛋白双向电泳分离结合质谱鉴定方法可用于分析ADP-核糖基化修饰蛋白,并发现ADP-核糖基化蛋白存在相对保守的结合序列。Objective To establish a method for identification of the poly（ ADP-ribose）binding proteins induced by benzo（ a） pyrene. Methods Poly（ ADP-ribose） binding protein were screened by immunoprecipitation assay and further separated by high performance liquid chromatography（ HPLC） and two dimensional electrophoresis,then identified by MALDI-TOFMS / MS. The proteins sequence were identified by two methods and compared the common binding motif with literature reports. Results Three poly（ ADP-ribose） binding proteins were identified by MALDI-TOF-MS / MS combined with immunoprecipitation assay and HPLC,and twelve poly（ ADP-ribose） binding proteins were identified by MALDI-TOF-MS / MS combined with immunoprecipitation assay and two dimensional electrophoresis. Most of them have a common binding motif which was consistent with the reported. Conclusion Combined the immunoprecipitation assay and two dimensional electrophoresis with MALDI-TOF-MS / MScould be used to analyze the poly（ ADP-ribose） binding proteins,and these proteins have a common conserved binding motif.

关 键 词：ADP-核糖基化 苯并[A]芘 免疫共沉淀 高效液相色谱 双向电泳 

分 类 号：Q513.2[生物学—生物化学]