method for reproducing in vitro the proteolytic activity of hcv protease ns3 virus.

摘要

PCT No. PCT/IT95/00018 Sec. 371 Date Aug. 23, 1996 Sec. 102(e) Date Aug. 23, 1996 PCT Filed Feb. 14, 1995 PCT Pub. No. WO95/22985 PCT Pub. Date Aug. 31, 1995This is a method for reproducing in vitro the serine protease activity associated with the HCV NS3 protein, that comprises using both of the sequences contained in NS3 and the sequences contained in NS4A. This method takes advantage of the ability of the HCV NS4A protein, or sequences contained therein, to act as a cofactor of the serine protease activity or more generally of the enzymatic activities associated with NS3. Optimal serine protease activity is obtained when NS4A is present in a molar ratio of at least 1:1 with NS3. NS3 and NS4A can also be incorporated in the reaction mixture as NS3-NS4A precursor, as this precursor will generate, by means of an autoproteolytic event, equimolar amounts of NS3 and NS4A. It is also possible to mutate the cleavage site between NS3 and NS4A in a procursor, so that NS4A remains covalently The sequences that do not influence the proteolytic activity of NS3 can subsequently be removed from protease activity.