AN ENZYME-LINKED-IMMUNOSORBENT-ASSAY FOR DETECTING PROTEOLYTIC ACTIVITY OF HEPATITIS-C VIRUS PROTEINASE

An ELISA method for the quantitation in vitro of HCV serine proteinase activity was developed. A peptide substrate, sp-Ile-Val-Pro-Cys-Ser-M et-Ser-Tyr-Thr-Trp-Thr-Lys (biotin) -OH (Sub-1), was hydrolyzed by a r ecombinant NS3 proteinase fused with maltose binding protein (MBP-NS3) into a product with a free amino moiety at the N-terminus. The produc t was immobilized, and the amino moiety was analyzed by digoxigenin la beling followed by immunological reaction with anti digoxigenin-alkali ne phosphatase conjugate and then the colorimeteric reaction. This met hod is suited for the high throughput screening of inhibitors, and the screening can be accelerated by automatic operation. (C) 1997 Academi c Press.