Assessment of substrate specificity of hepatitis G virus NS3 protease by agenetic method

The RNA genome of hepatitis G virus (HGV) encodes a large polyprotein that is processed to mature proteins by viral-encoded proteases. The HGV NS3 pro tease is responsible for the cleavage of the HGV polyprotein at four differ ent locations. No conserved sequence motif has been identified for the clea vage sites of the NS3 protease. To determine the substrate specificity of t he NS3 protease, amino acid sequences cleaved by the NS3 protease were obta ined from randomized sequence libraries by using a screening method referre d to as GASP (Genetic Assay for Site-specific Proteolysis). Based on statis tical analyses of the obtained cleavable sequences, a consensus substrate s equence was deduced: Gln-Glu-Thr-Leu-Val I Ser, with the scissile bond loca ted between Val and Ser. The relevance of this peptide as a cleavable subst rate was further supported by molecular modeling of the NS3 protease. Our r esult would provide an insight on the molecular activity of the NS3 proteas e and may be useful for the design of substrate-based inhibitors. (C) 2001 Academic Press.