Hepatitis C Virus E2 Protein Ectodomain Is Essential for Assembly of Infectious Virions
Figure 2
Oxidation kinetics and DTT-resistance analysis of HCV proteins E1 and E2. (a) S6.1 cells transfected with JFH1 or JFH1/Con1E2 RNA were pulse labeled with [35S] methionine and cysteine for 20 min and chased for different time periods, from 30 min to 6 hrs. PNSs were immunoprecipitated with a conformational monoclonal anti-E2 antibody (CBH-2 and CBH-5 for genotypes 2a and 1b, resp.) and analyzed on SDS-PAGE. Chase periods are reported above the lanes. Samples were analyzed on 10% SDS-PAGE under nonreducing (n.red) and reducing (red) conditions (DTT 200 mM). (b) S6.1 cells transfected with JFH1 or JFH1/Con1E2 RNA were pulse labeled and chased for the indicated time periods, in duplicate. One of the dishes in each pair was chased for an additional 5 min in the presence of 5 mM DTT (+) in the culture media before proceeding with lysis. PNSs were immunoprecipitated with CBH2 or CBH5 for genotype 2a and 1b E2 proteins, respectively. Upper panel, E2 protein. Lower panel, E1 protein. The samples were analyzed on 10% SDS PAGE under reducing and nonreducing conditions. Symbols refer to different form of E1 and E2: ox for oxidized; red for reduced; pr for protein precursor; NT for not transfected. Position on gel of prestained Molecular Weight marker (Amersham) is reported on the right side.
