酵母呈现SARS冠状病毒S蛋白受体结合区及其鉴定

Display of receptor-binding domain of SARS-CoV spike protein on the yeast cell surface

目的获得能在酵母表面正确折叠的SARS病毒S蛋白的受体结合区。方法通过PCR技术从pVAX1/S质粒扩增出编码S蛋白318至520氨基酸残基的cDNA,酶切后克隆到酵母表面呈现载体pYD1,构建pYD1-RBD重组质粒,转化酵母细胞,流式细胞仪检测所呈现的RBD片段。结果通过流式细胞仪检测到酵母细胞表面有RBD片段的表达,并且此表面呈现的RBD片段与针对RBD不同表位的2株单抗均能结合,SARS病毒免疫的小鼠血清抗体也能与该片段结合。结论酵母表面呈现的RBD保持了天然RBD分子的构象,为基于RBD的药物筛选和疫苗设计提供物质基础。

Objective To display receptor-binding domain （RBD） of SARS-CoV spike protein on the surface of yeast cell. Methods The DNA sequence coding RBD was amplified from pVAX1/S plasmid by PCR, then subcloned into the yeast surface display vector pYD1. The recombinant vector was transformed into yeast ceils EBY100. Flow cytometric analysis was carried out to evaluate direct binding of anti-RBD/anti-SARS CoV antibodies onto surface of yeast cell displaying RBD. Results RBD displayed on the surface of yeast cell could be recognized by two mAbs against RBD different epitopes and antibodies from SARS-CoV immtmized mouse serum. Conclusion RBD of SARS-CoV S protein can be displayed on the surface of yeast cell with efficient folding, which provide basis for screening anti-SARS-CoV agents and developing RBD-based vaccine.