EXPRESSION OF HIGHLY-ACTIVE RECOMBINANT NS3 PROTEASE DOMAIN OF HEPATITIS-C VIRUS IN ESCHERICHIA-COLI

The serine protease domain of HCV comprising amino acids 1027-1218 (De lta NS3) was expressed in E. coli with a His tag at its N-terminal end , The protease was purified to apparent homogeneity by a single step a ffinity chromatography resulting in high yields (similar to 3 mg/l of cultured cells). The Delta NS3 efficiently cleaves a 17-mer peptide co rresponding to the NS5A-NS5B junction with k(cat)/K-m = 160 X 10 min(- 1) mu M(-1) in the presence of NS4A peptide. Our Delta NS3 represents the minimal domain possessing highly active protease of NS3 constructe d so far. The Delta NS3 protein also efficiently processed a longer su bstrate corresponding to NS5A/5B junction (2203-2506 amino acids) that was synthesized by in vitro transcription and translation system.