与兔出血症病毒VP60蛋白相互作用蛋白的初步鉴定

Identification of the cellular protein interacted with vp60 protein of rabbit hemorrhagic disease virus

为获得与兔出血症病毒(RHDV)衣壳蛋白(VP60)相互作用的细胞蛋白,本研究以pMD-VP60重组质粒为模板扩增RHDV的VP60基因,克隆于表达载体pGEX-6p-1中,构建了重组表达质粒pGEX-VP60,并转化大肠杆菌BL21(DE3)中,经IPTG诱导表达GST重组蛋白(rVP60),western blot分析表明该蛋白具有与VP60单克隆抗体(MAb)良好的反应原性。以亲和层析纯化的rVP60进行pull-down实验,从兔的肝脏细胞膜蛋白中获得与VP60相互作用的蛋白,多肽质量指纹图谱分析表明,其相互作用的蛋白为ATP合成酶β亚基,提示该蛋白可能与RHDV的感染相关。

To identify the cellular protein interacted with VP60 of rabbit hemorrhagic disease virus （RHDV）, VP60 gene was amplified from pMD-VP60 by PCR and ligated to expression vector pGEX-6p-1. Recombinant protein （rVP60） was expressed in recombinant E.coli BE21 （DE3） induced with IPTG. Western blot showed that rVP60 was recognized by anti-VP60 monoclonal antibody. Furthermore, a protein was pull-down with rVP60 from cellular membrane protein lysate, which was identified as ATP synthase beta-subunit by mass spectrometry. Therefore, the ATP synthase beta-subunit could be involved in RHDV infection.