GE Fei-fei, QIU Ya-feng, YANG Yao-wu and CHEN Fu-yan. Compavison of JEV E-Hsp70 and E-Bindingdomain Fusion Protein on Immune Responses in Mice[J]. Virologica Sinica, 2005, 20(4): 357-367.
Citation: GE Fei-fei, QIU Ya-feng, YANG Yao-wu, CHEN Fu-yan. Compavison of JEV E-Hsp70 and E-Bindingdomain Fusion Protein on Immune Responses in Mice .VIROLOGICA SINICA, 2005, 20(4) : 357-367.

乙脑病毒E-Hsp70与E-肽连接区对小鼠特异性免疫的比较

  • 在本实验室已构建的原核表达载体(含乙脑疫苗株SA14-14-2株E蛋白基因主要抗原片段)的基础上用巴斯德毕赤酵母系统表达,该片段长1113bp,编码371个氨基酸残基,研究酵母表达的该乙脑病毒(Japaneseencephali-tisvirus)E蛋白主要抗原片段与结核杆菌热休克蛋白70(Hsp70)的融合蛋白以及该抗原肽与Hsp70上的一个功能域-肽连接区(Peptidebindingdomain,以下简称BD)融合形成的蛋白,用这三种蛋白分别免疫BALB/c小鼠,以酵母单独表达的E蛋白主要抗原片段免疫作为对照,比较它们对小鼠抗E蛋白主要抗原片段特异性的细胞免疫和体液免疫的影响。采用腹腔内注射蛋白的方法免疫小鼠,主要从IL-2的mRNA水平,淋巴细胞的增殖和抗体滴度这三个方面进行比较,最后我们得出E-BD融合蛋白在免疫效果方面比E-Hsp70略好一些,所以在本试验中肽连接区是完全可以代替Hsp70独立行使其佐剂功能。

Compavison of JEV E-Hsp70 and E-Bindingdomain Fusion Protein on Immune Responses in Mice

  • Further investigation of this interesting adjuvant-free carrier effect is necessary to ascertain whether peptide-binding portion alone can replace the whole M.tuberculosis Hsp70 acting as carrier. In our study, we selected a portion of E protein with higher index of antigenic determinants dependent on analysis of computer software and con-structed two chimeric vectors of pPICZα-E-Hsp70 and pPICZα-E-BD. Vectors were transformed into yeast X-33 by electroporation. Expression of fusion protein in yeast was induced by the addition of methanol every 24 hours and analysed by SDS-PAGE and Western blotting. We produced E-Hsp70 fusion protein at a yield of 33 mg per litter of culture and E-Bindingdomain fusion protein at a yield of 97 mg per litter of culture in methylotrophic yeast Pichia pastoris.with specific antigenicitys. To compare the immune response induced by E-Hsp70 fusion protein with that induced by E-bindingdomain, mice were immunized i.p. on day 0 and day 21. Mice immunized with E-bindingdomain had highe

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    Compavison of JEV E-Hsp70 and E-Bindingdomain Fusion Protein on Immune Responses in Mice

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    Abstract: Further investigation of this interesting adjuvant-free carrier effect is necessary to ascertain whether peptide-binding portion alone can replace the whole M.tuberculosis Hsp70 acting as carrier. In our study, we selected a portion of E protein with higher index of antigenic determinants dependent on analysis of computer software and con-structed two chimeric vectors of pPICZα-E-Hsp70 and pPICZα-E-BD. Vectors were transformed into yeast X-33 by electroporation. Expression of fusion protein in yeast was induced by the addition of methanol every 24 hours and analysed by SDS-PAGE and Western blotting. We produced E-Hsp70 fusion protein at a yield of 33 mg per litter of culture and E-Bindingdomain fusion protein at a yield of 97 mg per litter of culture in methylotrophic yeast Pichia pastoris.with specific antigenicitys. To compare the immune response induced by E-Hsp70 fusion protein with that induced by E-bindingdomain, mice were immunized i.p. on day 0 and day 21. Mice immunized with E-bindingdomain had highe

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