摘要:目的 探讨Grp78/Bip在戊型肝炎病毒(HEV)衣壳蛋白进入宿主细胞过程中的作用。方法 采用pull-down和免疫共沉淀技术进一步验证p239与Grp78/Bip的相互作用;采用激光共聚焦显微镜技术检测p239与细胞膜表面Grp78/Bip共定位情况;采用原核表达纯化的Grp78/Bip蛋白封闭p239的Grp78/Bip结合位点,检测其阻断p239吸附细胞的效果。结果 p239与Grp78/Bip可以直接结合,而且这种结合是可逆的生理性结合;p239与Grp78/Bip在细胞膜上存在部分共定位;Grp78/Bip能部分阻断p239对肝细胞的吸附。结论 Grp78/Bip参与并介导HEV衣壳蛋白对宿主细胞的吸附。Objective To further investigate the interaction between recombinant hepatitis E virus (HEV) capsid protein p239 and Grp78/Bip and the role of Grp78/Bip in HEV penetration.Methods We utilized pull-down, immunoprecipitation and antibody blocking assays to examine the interaction between p239 and Grp78/Bip. Confocal microscopy was used to investigate the co-localization of these two proteins. Purified Grp78/Bip was used to block the attachment of p239 to host cells.Results p239 directly bound to Grp78/Bip and this binding was sensitive to ATP. Furthermore, antibody blocking results demonstrate that this interaction was indeed conformation-dependent. A partial co-localization of p239 and Grp/Bip was observed on the plasma membrane of HepG2 by confocal microscopy. Pre-incubation of Grp78/Bip with p239 significantly blocked the attachment of p239 to HepG2 cells.Conclusion Grp78/Bip participates in the attachment and/or entry of the HEV capsid protein to host cells. These results further contribute to the understanding of the entry mechanism of the hepatitis E virus after infection. if(document.getElementById('ChDivSummary').innerHTML!=""){CutSpan('ChDivSummary',500);DisplaySpanDiv('ChDivSummary');ClearSummaryOnLoad('SummaryLinkChID','SummaryLinkEnID');}else{CutSpan('EnDivSummary',1000);DisplaySpanDiv('EnDivSummary');ClearSummaryOnLoad('SummaryLinkEnID','SummaryLinkChID');}
Abstract：Objective To further investigate the interaction between recombinant hepatitis E virus (HEV) capsid protein p239 and Grp78/Bip and the role of Grp78/Bip in HEV penetration.Methods We utilized pull-down, immunoprecipitation and antibody blocking assays to examine the interaction between p239 and Grp78/Bip. Confocal microscopy was used to investigate the co-localization of these two proteins. Purified Grp78/Bip was used to block the attachment of p239 to host cells.Results p239 directly bound to Grp78/Bip and this binding was sensitive to ATP. Furthermore, antibody blocking results demonstrate that this interaction was indeed conformation-dependent. A partial co-localization of p239 and Grp/Bip was observed on the plasma membrane of HepG2 by confocal microscopy. Pre-incubation of Grp78/Bip with p239 significantly blocked the attachment of p239 to HepG2 cells.Conclusion Grp78/Bip participates in the attachment and/or entry of the HEV capsid protein to host cells. These results further contribute to the understanding of the entry mechanism of the hepatitis E virus after infection. if(document.getElementById('ChDivSummary').innerHTML!=""){CutSpan('ChDivSummary',500);DisplaySpanDiv('ChDivSummary');ClearSummaryOnLoad('SummaryLinkChID','SummaryLinkEnID');}else{CutSpan('EnDivSummary',1000);DisplaySpanDiv('EnDivSummary');ClearSummaryOnLoad('SummaryLinkEnID','SummaryLinkChID');}
