CAJ | 학술논문

目的探讨HBV X蛋白与细胞色素c氧化酶Ⅲ(COXⅢ)之间的结合作用.方法采用交合实验验证HBV X蛋白和COXⅢ在酵母细胞内的结合作用,利用离体结合实验证实两者在体外的结合作用,采用免疫共沉淀实验证实两者在哺乳动物细胞中的结合作用.结果携带COXⅢ基因的酵母与携带X基因的酵母交合后发生反应,β-半乳糖苷酶(LacZ)活性检测菌落呈现蓝色,离体结合实验Western印迹中出现特异性结合活性反应条带,提示X蛋白和COXⅢ在体外存在直接相互作用,免疫共沉淀实验在相对分子质量为17 000处出现蛋白条带,提示在哺乳动物细胞水平仍能检测到X蛋白和COXⅢ的特异结合.结论 COXⅢ与X蛋白在原核细胞和哺乳动物细胞中均存在特异性结合,HBV可能通过此反应干扰线粒体呼吸链功能及能量代谢过程.
목적 탐토HBV X단백여세포색소c양화매Ⅲ(COXⅢ)지간적결합작용.방법 채용교합실험험증HBV X단백화COXⅢ재효모세포내적결합작용,이용리체결합실험증실량자재체외적결합작용,채용면역공침정실험증실량자재포유동물세포중적결합작용.결과 휴대COXⅢ기인적효모여휴대X기인적효모교합후발생반응,β-반유당감매(LacZ)활성검측균락정현람색,리체결합실험Western인적중출현특이성결합활성반응조대,제시X단백화COXⅢ재체외존재직접상호작용,면역공침정실험재상대분자질량위17 000처출현단백조대,제시재포유동물세포수평잉능검측도X단백화COXⅢ적특이결합.결론 COXⅢ여X단백재원핵세포화포유동물세포중균존재특이성결합,HBV가능통과차반응간우선립체호흡련공능급능량대사과정.
Objective To explore the interaction between hepatitis B virus(HBV) X protein and cytochrome C oxidase Ⅲ (COXⅢ).Methods Mating experiment and in vitro binding assays were utilized tO verify the interaction between HBV X protein and COX Ⅲ in vivo and in vitro respectively in yeast cells.Coimmunoprecipitation was employed to demonstrated the binding between these tWO proteins in mammalian cells.Results In mating experiment,the resulting diploids containing both pAS2-1-X and pACT2-COX Ⅲ plasmids turned blue in the β-gal assay,the result of Western blot displayed a positive reactive strap which indicated X protein could interact directly with COX Ⅲ in vitro binding assays.Interaction between these two proteins in mammalian cells was further confirmed by coimmunoprecipitation,as a protein strap about 17 000 was detected in Western blot.Conclusions It is speculated the specific interaction may exist between HBV X protein and COX Ⅲ in both prokaryotie and mammalian cells.X protein may impair the mitochondrial respiration chain and energy metabolism through its association with COX Ⅲ.