拟南芥多聚ADP-核糖聚合酶Ⅱ(PARP2)活性的测定  

Enzymatic assay of Arabidopsis poly(ADP-ribose) polymerase Ⅱ

作　　者：吴巧[1] 吴殷[1] 葛晓春[1] 

机构地区：[1]复旦大学生命科学学院生物化学系,上海200438

出　　处：《安徽农业大学学报》2015年第6期904-909,共6页Journal of Anhui Agricultural University

基　　金：国家自然科学基金(31070232;31170169)资助

摘　　要：多聚ADP-核糖聚合酶PARP[poly(ADP-ribose)polymerase]催化的多聚ADP-核糖化反应是一种重要的蛋白质翻译后修饰手段,在DNA修复、染色质结构重塑、基因转录、细胞周期和细胞凋亡等过程中发挥重要作用。目前,动物中的PARP研究较为深入,但植物中进展缓慢。本实验利用PCR技术从拟南芥中得到PARP2基因,利用原核系统表达并纯化出PARP2蛋白,体外测定了PARP2的酶活。SDS-PAGE和Western blotting结果表明,PARP2能够催化自身多聚ADP-核糖化修饰,这一发现为继续研究植物多聚ADP-核糖化聚合酶及多聚ADP-核糖化修饰开拓了新方向。Poly （ADP-ribosyl） ation （PARylation）, catalyzed by poly （ADP-ribose） polymerases （PARPs）, is an important posttranslational modification of proteins. They play important roles in DNA repair, remodeling of chro- matin structure, gene transcription, cell cycle and cell death regulation. PARPs were well studied in animals but less understood in plants. The cDNA fragment of PARP2 was obtained by PCR from Arabidopsis and cloned into pro- karyotic expression vector. The recombinant protein TRX-PARP2 was expressed and purified for enzymatic assay in vitro. TRX-PARP2 protein band shifted upwardly in a time course on SDS-PAGE gel. Anti-PAR antibody was used to confmn the result in a western blotting assay. These data demonstrated that Arabidopsis PARP2 has auto-modification activity in vitro, which shed light on the further study of PARP2 and PARylation in plants.

关 键 词：拟南芥 PARP2 PAR 活性测定 

分 类 号：Q786[生物学—分子生物学]