Evidence for the formation of a heptameric ion channel complex by the hepatitis C virus p7 protein in vitro.
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Clarke D, Griffin S, Beales L, Gelais CS, Burgess S, Harris M, Rowlands D
Evidence for the formation of a heptameric ion channel complex by the hepatitis C virus p7 protein in vitro.
J Biol Chem. 2006 Dec 1;281(48):37057-68. Epub 2006 Oct 10.
- PubMed ID
- 17032656 [ View in PubMed]
- Abstract
The p7 protein of hepatitis C virus functions as an ion channel both in vitro and in cell-based assays and is inhibited by amantadine, long alkyl chain imino-sugar derivatives, and amiloride compounds. Future drug design will be greatly aided by information on the stoichiometry and high resolution structure of p7 ion channel complexes. Here, we have refined a bacterial expression system for p7 based on a glutathione S-transferase fusion methodology that circumvents the inherent problems of hydrophobic protein purification and the limitations of chemical synthesis. Rotational averaging and harmonic analysis of transmission electron micrographs of glutathione S-transferase-FLAG-p7 fusion proteins in liposomes revealed a heptameric stoichiometry. The oligomerization of p7 protein was then confirmed by SDS-PAGE and mass spectrometry analysis of pure, concentrated FLAG-p7. The same protein was also confirmed to function as an ion channel in suspended lipid bilayers and was inhibited by amantadine. These data validate this system as a means of generating high resolution structural information on the p7 ion channel complex.