重组人成纤维细胞生长因子-21的表达及纯化工艺的优化

Optimization of Procedure for Expression and Purification of Recombinant Human Fibroblast Growth Factor-21

目的优化重组人成纤维细胞生长因子-21(SUMO-rhFGF-21)融合表达工程菌的表达条件及目的蛋白纯化工艺。方法对工程菌的诱导温度、时间及诱导剂浓度进行优化,并进行罐发酵,对菌体裂解液进行离子交换层析、Ni离子亲和层析及分子筛层析等。纯化产物经SDS-PAGE和HPLC鉴定纯度。结果在诱导温度为37℃,诱导时间为4h,IPTG浓度为0.5mmol/L时,目的蛋白的表达量最高,达20.5%;罐发酵收菌量可达66g/L,目的蛋白的表达量达20.2%;纯化后的rhFGF-21纯度达96%以上。结论优化了rhFGF-21的表达条件及纯化工艺,为rhFGF-21用于新药开发奠定了基础。

Objective To optimize the condition for fermentation of recombinant E. coli with fusion gene of SUMO and human fibroblast growth factor-2l （SUMO-rhFGF-21） as well as the procedure for purification of target protein. Methods The temperature and time for induction as well as concentration of inducer of recombinant E. coil was optimized. The recombinant E. coli was ferment- ed under the optimal condition, and the lysate of bactefa was purified by ion exchange, nickel ion affinity and molecular sieve chromatography. The purified target protein was determined for purity by SDS-PAGE and HPLC. Results The optimal temperature and time for induction of recombinant E. coil were 37℃ and 4 h respectively, while the optimal concentration of IPTG as an inducer was 0. 5 mmol/L. The expression level of target protein reached a peak value of 20. 5% under the optimal condition. The yield of bacteria after fermentation reached 66 g/L, and the expression level of target protein was 20. 2%. After purification, the purity of target protein was more than 96%. Conclusion The procedure for expression and purification of rhFGF-21 was optimized, which laid a foundation of developing rhFGF-21 as a novel drug.