title: Two different conformations in hepatitis C virus p7 protein account for proton transport and dye release.
abstract: The p7 protein from the hepatitis C virus (HCV) is a 63 amino acid long polypeptide that is essential for replication, and is involved in protein trafficking and proton transport. Therefore, p7 is a possible target for antivirals. The consensus model for the channel formed by p7 protein is a hexameric or heptameric oligomer of α-helical hairpin monomers, each having two transmembrane domains, TM1 and TM2, where the N-terminal TM1 would face the lumen of this channel. A reported high-throughput functional assay to search for p7 channel inhibitors is based on carboxyfluorescein (CF) release from liposomes after p7 addition. However, the rationale for the dual ability of p7 to serve as an ion or proton channel in the infected cell, and to permeabilize membranes to large molecules like CF is not clear. We have recreated both activities in vitro, examining the conformation present in these assays using infrared spectroscopy. Our results indicate that an α-helical form of p7, which can transport protons, is not able to elicit CF release. In contrast, membrane permeabilization to CF is observed when p7 contains a high percentage of β-structure, or when using a C-terminal fragment of p7, encompassing TM2. We propose that the reported inhibitory effect of some small compounds, e.g., rimantadine, on both CF release and proton transport can be explained via binding to the membrane-inserted C-terminal half of p7, increasing its rigidity, in a similar way to the influenza A M2-rimantadine interaction.
authors.affiliation: School of Biological Sciences, Nanyang Technological University, Singapore, Singapore.
authors.forename: Siok Wan
authors.initials: SW
authors.lastname: Gan
authors.affiliation: School of Biological Sciences, Nanyang Technological University, Singapore, Singapore.
authors.forename: Wahyu
authors.initials: W
authors.lastname: Surya
authors.affiliation: School of Biological Sciences, Nanyang Technological University, Singapore, Singapore.
authors.forename: Ardcharaporn
authors.initials: A
authors.lastname: Vararattanavech
authors.affiliation: School of Biological Sciences, Nanyang Technological University, Singapore, Singapore.
authors.forename: Jaume
authors.initials: J
authors.lastname: Torres
mesh_terms: D001692:Biological Transport; D004579:Electron Transport; D005456:Fluorescent Dyes / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism; D013058:Mass Spectrometry; D011487:Protein Conformation; D054730:Protein Interaction Domains and Motifs; D017494:Proton Pumps / Q000737:chemistry / Q000378:metabolism; D011522:Protons; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000086082:Viroporin Proteins
chemical_list: D005456:Fluorescent Dyes; D008051:Lipid Bilayers; D017494:Proton Pumps; D011522:Protons; D011994:Recombinant Proteins; D014764:Viral Proteins; C088369:p7 protein, Hepatitis C virus; D000086082:Viroporin Proteins