CTB/CS3融合蛋白与GM1结合能力和免疫原性的分析

免疫霍乱毒素B亚单位（CTB）或肠毒素大肠杆菌（ETEC）定居因子CS3可使人体对ETEC的侵染有保护作用.为探索研制ETEC双组分亚单位疫苗的可行性，利用大肠杆菌诱导表达系统表达了CTB与CS3的融合蛋白（CTB/CS3）.蛋白质印迹结果表明，诱导表达的29 ku蛋白具有CTB和CS3蛋白双重抗原性.经Ni-NTA亲和层析纯化获得重组蛋白CTB/CS3，复性的重组蛋白可以部分形成五聚体并保留了与神经节苷脂GM1的结合能力.动物实验表明，融合蛋白CTB/CS3具有CTB和CS3蛋白的双重免疫原性，同时，CTB的免疫载体作用提高了CS3的免疫强度.

GM1-binding Ability and Immunogenicity of CTB/CS3 Fusion Protein Expressed in E.coli

Enterotoxigenic Escherichia coli (ETEC) is a major pathogen that evokes acute diarrhea among children worldwide and travelers to developing countries. However, there is no ideal vaccine against it yet. In an effort to develop a subunit vaccine for ETEC, a translational fusion with cholera toxin B subunit (CTB) upstream of CS3 was constructed. The fusion protein synthesized in E.coli had a molecular mass of 29 ku, as expected and retained the antigenicity of both CTB and CS3 as confirmed by Western blot analysis with the polyclonal anti-CTX rabbit serum and the monoclonal anti-CS3 mouse serum, respectively. The 6×His-tagged CTB/CS3 protein was purified by Ni-NTA affinity chromatography followed by renaturation. A fraction of the fusion protein could form pentamers and these pentamers retained the ability to bind GM1-ganglioside. Mice immunized by intraperitoneal injection with the fusion protein produced anti-CTB and anti-CS3 serum IgG and secretory IgA. Furthermore, it was shown that fusion to CTB increased the systemic and mucosal immune responses against CS3 to some extent.