摘要
泛素化作为真核生物细胞内普遍存在且极为关键的蛋白质翻译后修饰,可以使被修饰的蛋白质发生降解,在植物生长发育、胁迫应答等方面发挥着重要作用。泛素化降解需要通过E3泛素连接酶特异性识别靶蛋白并对其进行修饰,因此E3连接酶在泛素化途径中起着决定性的作用。文章通过克隆番茄SlSL1基因编码序列、构建原核重组表达质粒,并利用诱导表达、亲和吸附纯化SlSL1重组蛋白,以此进行SlSL1的体外泛素化活性检测。SDS-PAGE电泳和考马斯亮蓝染色结果显示,由所构建番茄SlSL1基因重组质粒纯化获得SlSL1蛋白;体外泛素化检测显示,SlSL1蛋白形成多聚化泛素条带,具有E3泛素连接酶活性,表明SlSL1可能在番茄的泛素化调控途径中发挥功能。
Ubiquitination is a ubiquitous and critical post-translational protein modification in eukaryotic cells,which can degrade the modified proteins and play an important role in plant growth,development and stresses response.The ubiquitination requires E3 ubiquitin ligases to specifically recognize and modify the target proteins,so E3 ligases play a decisive role in the ubiquitin pathway.In this paper,tomato SlSL1 gene was cloned for prokaryotic expression vector construction and the SlSL1 recombinant protein was purified by induction expression and affinity adsorption,then the ubiquitination activity of SlSL1 was detected in vitro.SDS-PAGE and Coomassie blue staining showed that SlSL1 protein was purified from SlSL1 gene recombinant plasmid.Analysis of ubiquitination indicated the E3 ligase activity of SlSL1 by the presence of polyubiquitin chains,suggesting its regulatory function through ubiquitination pathway in tomato.
作者
徐焕焕
张芳毓
刘茜
栾能能
王颖格
牛向丽
XU Huanhuan;ZHANG Fangyu;LIU Qian;LUAN Nengneng;WANG Yingge;NIU Xiangli(School of Food and Biological Engineering,Hefei University of Technology,Hefei 230601,China)
出处
《合肥工业大学学报（自然科学版）》
CAS
北大核心
2022年第8期1130-1134,共5页
Journal of Hefei University of Technology：Natural Science
基金
国家自然科学基金资助项目(31671266)
合肥工业大学大学生创新创业训练计划资助项目(s202110359376)。
关键词
番茄
E3泛素连接酶
蛋白纯化
体外泛素化
tomato
E3 ubiquitin ligase
protein purification
in vitro ubiquitination
