HBeAg与CD81分子结合的研究

目的 :研究HBeAg与CD81分子在细胞内、外的相互作用。方法 :应用RT PCR技术 ,从HepG2细胞中扩增CD81全基因 ,并构建重组真核表达载体。将其与pGBKT7 eAg共转染营养缺陷型酵母细胞 ,观察生长情况。应用网织红细胞裂解体外翻译及免疫共沉淀试验 ,进一步验证CD81分子与HBeAg的结合。结果 :经EcoRI和BamHI酶切和DNA序列测定鉴定表明 ,构建的CD81基因的重组表达载体正确。共转染后的酵母细胞在营养缺陷的培养基中生长良好。体外免疫共沉淀试验证实 ,CD81与HBeAg出现沉淀带。结论 :HBeAg与CD81分子在细胞内、外均可结合 ,推测CD81在HBV的致病过程中起着重要的作用

Study on binding of HBeAg to CD81

AIM: To investigate the interaction between HBeAg and CD81. METHODS: The CD81 gene was amplified by RT-PCR from HepG2 cells. The recombinant expression vector pGADT7-CD81 was constructed by routine molecular biological method. The auxotroph yeast cells were cotransfected with pGADT7-CD81 and pGBKT7-eAg and plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing x-alpha-gal. The binding of HBeAg to CD81 was also detected in vitro translation and coimmunoprecipitation test. RESULTS: The recombinant expression vector was constructed and confirmed by restriction enzyme (EcoR I and BamH I) digestion and DNA sequencing analysis. The positive yeast clones could grow and from blue colonies on SD/-Trp-Leu-His-Ade/x-alpha-gal medium. The result of immunocoprecipitation showed that HBeAg could bind to CD81. CONCLUSION: HBeAg can bind to CD81, suggesting that CD81 plays an important role in the pathogenesis of HBV.