摘要
SHP-2是一种非跨膜型蛋白酪氨酸磷酸酶。将其N末端结构域C-SH2克隆至原核表达载体pGEX-2T中,命名为pGEX-2T-C-SH2。该融合蛋白在大肠杆菌中以可溶形式表达于菌体上清液中,经GST亲和层析柱和FPLC分子筛柱纯化后,目的蛋白纯度可达98%。质谱分析表明,目的蛋白相对分子量与理论值基本相同。在最适表达条件下,1 L的M9培养基能够高效表达并获得12 mg C-SH2蛋白,NMR结构和活性位点的分析表明,目的蛋白能正确折叠并呈规则的空间结构。
SHP-2 is a non-transmembrane and protein tyrosine phosphatase, its N terminus domain C- SH2 is cloned into pGEX-2T vector and named as pGEX-2T-C-SH2. The fusion protein is expressed in Escherichia coli and as the supernatant of lysate. The purity of target protein is about 98% after GST- column and FPLC purification. The MS data confirms the molecular weight of the protein, which is very close to the theoretical value. Under the optimized conditions for the purification of C-SH2, the yield of the purified protein is estimated to be 12 mg from 1 L of M9 minimal media. NMR analysis indicates that the protein is properly folded, and existed in a well-ordered structure.
出处
《浙江理工大学学报（自然科学版）》
2009年第4期602-607,共6页
Journal of Zhejiang Sci-Tech University(Natural Sciences)
