关键词: 人神经生长因子, 大肠杆菌, 可溶表达, 生物学活性

Abstract: The goal of this work is to express the recombinant human nerve growth factor（rhβNGF）in Escherichia coli in soluble form,to separate and purify the expressed products,and to determine the biological activity. First,hβNGF gene was amplified and then inserted into expression vector pMAL-c2X,then E. coli expression system of hβNGF-MBP was constructed and induced for expression. Further,the MBP in purified expressed products was cleaved by Factor Xa enzyme,after identified by Western blot,the biological activity was examined by TF-1 method. The results showed that enzyme digestion and sequencing confirmed that the recombinant plasmid pMAL-c2X-hβNGF was constructed correctly,hβNGF-MBP was secretory expressed under 2℃,180 r/min and 0.5 mmol/L IPTG induction. MBP tag in hβNGF-MBP was removed by Factor Xa digestion,and the purified hβNGF via SDS-PAGE was about 13 kD with the purity over 95%. The protein was identified as hβNGF by Western blot. The biological activity test showed that the specific activity was about 1×10⁶ U/mg. Overall,recombinant hβNGF was successfully expressed in E. coli in soluble form with high biological activity.

Key words: human nerve growth factor, Escherichia coli, soluble expression, biological activity