Griffin Stephen D C, Harvey Ruth, Clarke Dean S, Barclay Wendy S, Harris Mark, Rowlands David J
Astbury Centre of Molecular Biology, School of Biochemistry and Microbiology, University of Leeds, Leeds LS2 9JT, UK.
Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK.
J Gen Virol. 2004 Feb;85(Pt 2):451-461. doi: 10.1099/vir.0.19634-0.
We previously identified the function of the hepatitis C virus (HCV) p7 protein as an ion channel in artificial lipid bilayers and demonstrated that this in vitro activity is inhibited by amantadine. Here we show that the ion channel activity of HCV p7 expressed in mammalian cells can substitute for that of influenza virus M2 in a cell-based assay. This was also the case for the p7 from the related virus, bovine viral diarrhoea virus (BVDV). Moreover, amantadine was shown to abrogate HCV p7 function in this assay at a concentration that specifically inhibits M2. Mutation of a conserved basic loop located between the two predicted trans-membrane alpha helices rendered HCV p7 non-functional as an ion channel. The intracellular localization of p7 was unaffected by this mutation and was found to overlap significantly with membranes associated with mitochondria. Demonstration of p7 ion channel activity in cellular membranes and its inhibition by amantadine affirm the protein as a target for future anti-viral chemotherapy.
我们之前已确定丙型肝炎病毒（HCV）p7蛋白在人工脂质双分子层中作为离子通道的功能，并证明金刚烷胺可抑制这种体外活性。在此我们表明，在基于细胞的检测中，哺乳动物细胞中表达的HCV p7的离子通道活性可替代流感病毒M2的离子通道活性。相关病毒牛病毒性腹泻病毒（BVDV）的p7也是如此。此外，在该检测中，金刚烷胺在特异性抑制M2的浓度下可消除HCV p7的功能。位于两个预测的跨膜α螺旋之间的保守碱性环发生突变，使HCV p7作为离子通道失去功能。p7的细胞内定位不受此突变影响，且发现与线粒体相关膜有显著重叠。细胞膜中p7离子通道活性的证明及其被金刚烷胺抑制，证实该蛋白是未来抗病毒化疗的一个靶点。
