AN AMINO-TERMINAL DOMAIN OF THE HEPATITIS-C VIRUS NS3 PROTEASE IS ESSENTIAL FOR INTERACTION WITH NS4A

Hepatitis C virus (HCV) genomic RNA is translated into a large polypro tein that is processed into structural and nonstructural proteins. Pro cessing at the N termini of several nonstructural proteins requires se quences contained in both NS3 and NS4A. NS3 contains a serine protease , whereas the function of NS4A in proteolysis is yet to be determined. By using the vaccinia virus-T7 hybrid expression system to transientl y express HCV polypeptides in HeLa cells, we studied the effect of sev eral N-terminal and C-terminal deletions of HCV NS3 on the processing activity at all the downstream cleavage sites. In this way, we have de lineated the minimal domain of NS3 required for the serine protease ac tivity associated with this protein. In addition, we demonstrate the f ormation of a stable complex between NS3 and NS4A: analysis of the del etion mutants reveals a region at the N terminus of NS3 that is necess ary for both complex formation and modulation of the proteolytic activ ity by NS4A but npt for the NS4A-independent serine protease activity of NS3.