【摘要】 利用高效表达载体及人工合成的寡核苷酸接头,构建了一个新的人rIL-4表达克隆pBMhIL4,在pL启动子的控制下,于大肠杆菌中表达完整的人rIL-4蛋白分子。表达产物以包涵体形式存在于大肠杆菌胞浆,SDS-PAGE分析表明表达的人rIL-4约占总菌体蛋白的5～10%,分子量约为15KD。抽提复性后具有较高的生物学活性,以其TCGF样活性检测每升菌液含人IL-4 1×10~6 U,改良的3.5 M盐酸胍加0.25%Triton X100裂解包涵体法具有更高的回收率和生物学活性。更多还原

【Abstract】 Utilizing higher expressing vector and a synthetic oligonucleotide linker,a new expressing clone, pBMhIL 4, has been constructed in our laboratory. The intact human rIL-4 protein molecule was expressed under the control of pL promoter in E. coli. The gene encoding protein was expressed in the plasma of E. coli in the form of inclusion bodies, the expressed IL-4 constituting about 5-10% of the total cellular proteins by SDS-PAGE analysis. The molecular weight of human rIL-4 was about 15KD. After extraction and renaturation, the yieldedsoluble human rIL-4 exhibited biological activity. 1×10~6 units of human IL-4 were produced from 1 liter of bacteria extracts assassed by the assay of its TCGF activity. Using 3.5M Guanidine hydrochloride and 0.25% Triton X100 to lyze inclusion bodies resulted in higher yield and higher biological activity.更多还原