FIG. 6.

Translation and processing of HCV polyprotein in Krebs-2 S10 as effected by the indicated concentrations of compound A, a hexapeptide-like competitive inhibitor of HCV NS3 protease (51). (A) Compound A, when used in a 0.25 to 1,000 nM concentration range, does not inhibit HCV RNA translation. Reaction mixtures containing the indicated concentrations of compound A were programmed with HCV RNA under standard conditions (serial dilutions of compound A were made up in 0.1% dimethyl sulfoxide, and 1 μl of each dilution was added to a 20-μl reaction mixture). Portions (1 μl) of each sample were assayed for TCA-insoluble radioactivity. (B) Compound A affects NS3 protease-mediated, but not signal peptidase-mediated, processing of the HCV polyprotein. Portions of the reaction mixtures in panel A were subjected to SDS-15% PAGE analysis and autoradiography. To improve detection of the high-molecular-weight products, the exposure to the gel of the film used for scanning the upper part of the figure was reduced three times compared to that used for printing the lower part of the figure. HCV-specific proteins whose appearance is sensitive and resistant to compound A are indicated at the left and right, respectively.