Objectives

Develop a Cell Surface Display system in Saccharomyces cerevisiae, based on the construction of an expression cassette for pYES2 plasmid.

Results

The construction of an expression cassette containing the α-factor signal peptide and the C-terminal portion of the α-agglutinin protein was made and its sequence inserted into a plasmid named pYES2/gDαAgglutinin. The construction allows surface display of bovine herpesvirus type 5 (BoHV-5) glycoprotein D (gD) on S. cerevisiae BY4741 strain. Recombinant protein expression was confirmed by dot blot, and indirect immunofluorescence using monoclonal anti-histidine antibodies and polyclonal antibodies from mice experimentally vaccinated with a recombinant gD.

Conclusions

These results demonstrate that the approach and plasmid used represent not only an effective system for immobilizing proteins on the yeast cell surface, as well as a platform for immunobiologicals development.

中文翻译：

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酿酒酵母中酵母表面展示的表达盒和质粒构建

目标

基于 pYES2 质粒表达盒的构建，在酿酒酵母中开发细胞表面展示系统。

结果

构建包含α-因子信号肽和α-凝集素蛋白C-末端部分的表达盒，并将其序列插入名为pYES2/gDα凝集素的质粒中。该构造允许在酿酒酵母BY4741 菌株上表面展示 5 型牛疱疹病毒 (BoHV-5) 糖蛋白 D (gD) 。重组蛋白表达通过斑点印迹法和间接免疫荧光法使用单克隆抗组氨酸抗体和多克隆抗体证实，这些抗体来自用重组 gD 进行实验接种的小鼠。

结论

这些结果表明，所使用的方法和质粒不仅代表了将蛋白质固定在酵母细胞表面的有效系统，而且代表了免疫生物学开发的平台。