FIG. 4.

Examination of WT and combined deletion and insertion mutant S proteins expressed in HuH-7 cells. Each deletion and insertion consisted of a five-amino-acid-residue deletion and insertion of Lys-Leu (A) or Gly-Ala (B) residues as described in the Materials and Methods section. Six days after transfection of 0.5 × 10⁶ HuH-7 cells with 2 μg of p123 DNA or its derivatives, transfected cells were labeled with 200 μCi of [³⁵S]Cys-[³⁵S]Met for 24 h. After labeling, cell lysates (C) and supernatants (S) were immunoprecipitated with anti-S antibodies. One half of each immunoprecipitate was analyzed as described in the legend for Fig. 2. The migration positions of glycosylated (gp27) and non-glycosylated (p24) S proteins are indicated. EN, env-negative.