Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
2020286 | Protein Expression and Purification | 2015 | 6 Pages |
Abstract
Non-structural protein 2 (NS2) of the hepatitis C virus (HCV) is an integral membrane protein that contains a cysteine protease and that plays a central organizing role in assembly of infectious progeny virions. While the crystal structure of the protease domain has been solved, the NS2 full-length form remains biochemically and structurally uncharacterized because recombinant NS2 could not be prepared in sufficient quantities from cell-based systems. We show here that functional NS2 in the context of the NS2-NS3pro precursor protein, ensuring NS2-NS3 cleavage, can be efficiently expressed by using a wheat germ cell-free expression system. In this same system, we subsequently successfully produce and purify milligram amounts of a detergent-solubilized form of full-length NS2 exhibiting the expected secondary structure content. Furthermore, immuno-electron microscopy analyses of reconstituted proteoliposomes demonstrate NS2 association with model membranes.
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Authors
Marie-Laure Fogeron, David Paul, Vlastimil Jirasko, Roland Montserret, Denis Lacabanne, Jennifer Molle, Aurélie Badillo, Célia Boukadida, Sonia Georgeault, Philippe Roingeard, Annette Martin, Ralf Bartenschlager, François Penin, Anja Böckmann,