抗HIV-1gp120单链抗体的原核表达与初步纯化

Expression of Single Chain Antibody to HTV - 1 gpl20 and Preliminary Pu-rification of Expressed Product

目的 在原核细胞表达抗HIV-1gp120单链抗体基因,纯化后检测其活性。方法 将单链抗体基因插入pET28原核表达载体进行诱导表达,对包涵体进行变性复性处理后,利用Ni-NTA金属螯合层析柱对表达的蛋白进行初步纯化,纯化后的单链抗体与gp120标准抗原膜条反应。结果 表达产物主要以包涵体形式存在,最高表达量占菌体总蛋白量的51％,金属螯合层析一步纯化后纯度超过85％。纯化后的单链抗体可以特异地识别gp120标准抗原。结论 在原核细胞表达的单链抗体,复性纯化后具有生物学活性。

Objective To express anti - HIV - 1 gp120 ScFv gene in E. coli, purify expressed product and detect the biological activity of it. Methods The anti-HIV - 1 gp120 ScFv gene was inserted into vector pET28, then transformed to E. coli for expression. The expressed product was degenerated and renatu-ralized, and purified by Ni - NTA affinity chromatography. The purified ScFv was identified by reaction with standard gp120 antigen. Results The expressed product existed mainly in the forms of inclusion bodies and contained 51% of total protein. After being purified by Ni - NTA chromatography, the purity of it reached above 85% . The purified ScFv recognized standard gp120 antigen specifically. Conclusion ScFv with biological activity was highly expressed in E. coli.