新型疫苗佐剂Ov-ASP-1佐剂活性功能区的设计、表达及生物学特性研究

目的 获得保留Ov-ASP-1佐剂活性的功能区。方法 用前期制备的Ov-ASP-1单克隆抗体对Ov-ASP-1的9个肽进行特异性结合,根据肽的结合情况设计功能区ASPPRM,并进行密码子优化、构建原核表达载体后在大肠杆菌中表达。表达产物经 Western印迹鉴定后的用镍柱纯化蛋白,复性完全后对获得的ASPPRM活性蛋白进行生物学特性分析。结果 ASPPRM以包涵体形式表达,分子量为17 kD。将其和OVA共同免疫小鼠后ASPPRM组抗OVA的IgG抗体显著高于OVA组。结论 本研究获得了保留佐剂活性的ASPPRM蛋白,为后续ASPPRM体内的免疫增效作用及Ov-ASP-1佐剂活性功能域的探索奠定基础。

Expression on the functional domain of Ov-ASP-1 protein in Eschervchia coli and its adjuvanticity in vivo

We aimed to find and express the functional domain of Ov-ASP-1, which has similar adjuvanticity to Ov-ASP-1. Through the binding results of the monoclonal antibodies (mAb) specific to the Ov-ASP-1 and peptides of Ov-ASP-1, which was prepared previously, we designed a functional domain ASPPRM. Then, ASPPRM protein was expressed in E. coli, purified by affinity chromatography with Ni-charged resin and refolded by gradient dialysis to soluble form. The purified ASPPRM was intramuscularly immunized to BALB/c mice to evaluate its biologicalactivity. The functional domain ASPPRM was expressed in E. coli in the form of inclusion bodies. ASPPRM enhanced the immunogenicity of OVA in BALB/c mice. The ASPPRM protein was obtained, which had similar adjuvanticity to Ov-ASP-1. This research may lay a foundation for the following work on the adjuvanticity of ASPPRM and the functional domain of Ov-ASP-1.