一个能诱生戊型肝炎病毒中和抗体的ORF2编码蛋白短片段

目的：比较戊型肝炎病毒（HEV）第4基因型ORF2编码蛋白片段pN472-C617（aa472～617）和pN477-C613（aa477～613）的免疫原性，找到能诱生HEV中和抗体的ORF2编码蛋白的更短片段。方法：表达和纯化pN472-C617和pN477-C613，分别免疫BALB/c小鼠，以间接ELISA检测免疫血清的抗体效价，并以基于PCR的体外中和试验检测免疫血清的中和活性。结果：pN472-C617和pN477-C613可在大肠杆菌高效可溶性表达，纯化后的重组蛋白能在小鼠体内诱导出高效价的抗体。基于PCR的体外中和试验显示pN472-C617免疫血清可有效中和HEV，阻断其在敏感细胞表面的吸附和细胞内复制；而两端仅比pN472-C617各短5个氨基酸的pN477-C613的免疫血清不具有中和HEV的活性。结论：重组蛋白pN472-C617具有良好的抗原性和诱生中和抗体的能力，是目前文献报道中含有HEV中和抗原表位的最短ORF2编码蛋白片段，可作为重组亚单位候选疫苗用于HEV疫苗的开发。

A short, neutralizing antibody-inducible, ORF2-encoded protein of hepatitis E virus

Objective:To find a short, neutralizing antibody-inducible, ORF2-encoded protein by means of comparing the immunogenicity of pN472-C617 and pN477-C613 which represent amino acids 472-617 and 477-613 of HEV ORF2-encoded protein of hepatitis E virus(HEV) genotype 4, respectively.Methods:The two recombinant proteins were expressed, purified, and then used to immunize BALB/c mice. Anti-HEV titers in the immune sera were detected by ELISA. Anti-HEV neutralizing activity was tested by a PCR-based in vitro neutralization assay.Results:Both of the two recombinant proteins were efficiently expressed in E.coli in soluble forms. The purified proteins induced mice to develop high levels of anti-HEV specific antibodies. However, only the immune sera obtained from the mice immunized with pN472-C617 showed the neutralizing activity to the homologous HEV strain by preventing the virus from absorption on PLC/PRF/5 cells surfaces and replication in the cells. The immune sera against pN477-C613, which was truncated five amino acids from both N- and C-terminal of pN472-C617, had no HEV neutralizing activity.Conclusion:The pN472-C617 is the shortest neutralizing antibody-inducible ORF2-encoded protein of HEV reported in literatures so far. It may be considered as a potential candidate for a novel HEV subunit vaccine in our future study.