A novel mechanism to ensure terminal initiation by hepatitis C virus NS5B polymerase

Hepatitis C virus (HCV) nonstructural protein 5B (NS5B) RNA-dependent RNA polymerase (RdRp) has acquired a unique beta -hairpin in the thumb subdomain which protrudes toward the active site, We report here that this beta -hairpin plays an important role in positioning the 3 ' terminus of the viral RNA genome for correct initiation of replication The presence of this beta -hairpin interferes with polymerase binding to preannealed double-stranded RNA (dsRNA) molecules and allows only the single-stranded 3 ' terminus of an RNA template to bind productively to the active site, We propose that this beta -hairpin may serve as a gate which prevents the 3 ' terminus of the template RNA from slipping through the active site and ensures initiation of replication from the terminus of the genome. This hypothesis is supported by the ability of a beta -hairpin deletion mutant that utilizes dsRNA substrates and initiates RNA synthesis internally. The proposed terminal initiation mechanism may represent a novel replication strategy adopted by HCV and related viruses. (C) 2001 Academic Press.