Multiple interactions within the hepatitis C virus RNA polymerase repress primer-dependent RNA synthesis

The hepatitis C virus (HCV) RNA-dependent RNA polymerase (RdRp) initiates RNA synthesis in vivo by a de novo mechanism. In vitro, however, the HCV RdRp can initiate de novo or extend from a primed template. A novel beta-loop near the RdRp active site was previously found to prevent the use of primed templates. We found that, in addition to the beta-loop, the C-terminal tail of the HCV RdRp and the de novo initiation GTP are required to exclude the use of primed-templates. GTP binding to the NTPi site of the HCV RdRp orchestrates the participation of other structures. The interactions of the beta-loop, C-terminal tail, and GTP provide an elegant solution to ensure de novo initiation of HCV RNA synthesis.