FIG. 6.

Effect of trypsin on the cleavage site mutants. HeLa cell monolayers were cotransfected with plasmids encoding the SV41 HN protein and each cleavage site mutant. After 22 h, the cells were treated or mock treated with 5 μg/ml of trypsin at 37°C for 15 min and biotinylated with NHS-LC-biotin. Then, the cell lysates were subjected to immunoprecipitation with MAb 1D1, and the immunoprecipitates were separated by SDS-PAGE under reducing conditions, followed by electroblotting onto a PVDF membrane. The biotinylated proteins on the membrane were visualized as described in the legend to Fig. 3C.