马尔堡病毒糖蛋白GP1亚基的真核表达与纯化

Eukaryotic Expression and Purification of Marburg Virus Envelope GP1 Protein

目的:用真核表达系统可溶性表达重组马尔堡病毒糖蛋白GP1亚基并纯化。方法:从糖蛋白(GP)全长基因上扩增GP1基因序列,克隆至真核表达载体pcDNA3.4,在哺乳动物表达系统中进行表达,镍柱亲和层析纯化目的蛋白,ELISA检测重组蛋白与特异抗体的免疫反应性。结果:GP1在哺乳动物表达系统中获得可溶性表达,纯化得到的目的蛋白与特异抗体具有良好的结合活性,推测其构象与天然马尔堡病毒GP1具有相似性。结论:真核表达了马尔堡病毒糖蛋白GP1亚基,且目的蛋白具有良好的抗原性,可用于GP特异抗体筛选、疫苗效果评价等研究。

Objective: To construct an expressing vector of Marburg virus(MARV) envelope GP1 protein, and express recombinant GP1 protein in mammalian expression system. Methods: The GP1 sequence was amplified from full-length gene of MARV glycoprotein(GP) and cloned into pc DNA3.4 vector. After sequencing, the recombinant pc DNA3.4-GP1 was transfected into Expi293 cells. The protein of interest was purified through Ni affinity chromatography and identified by Western blot and ELISA assays. Results: We obtained recombinant GP1 protein from the supernatant of transfected Expi293 cells. Purified GP1 protein can bind anti-MARV GP specific monoclonal antibodies. Conclusion: GP1 protein has been expressed in mammalian expression system, and can be used to screen specific antibodies and evaluate vaccine efficacy.