Abstract
A number of hepatitis C virus (HCV) proteins, including NS5B, the RNA-dependent RNA polymerase, were detected in membrane fractions from Huh7 cells containing autonomously replicating HCV RNA replicons. These membrane fractions were used in a cell-free system for the analysis of HCV RNA replication. Initial characterization revealed a reaction in which the production of replicon RNA increased over time at temperatures ranging from 25 to 40 degrees C. Heparin sensitivity and nucleotide starvation experiments suggested that de novo initiation was occurring in this system. Both Mn2+ and Mg2+ cations could be used in the reaction; however, concentrations of Mn2+ greater than 1 mM were inhibitory. Compounds shown to inhibit recombinant NS3 and NS5B activity in vitro were found to inhibit RNA synthesis in the cell-free system. This system should be useful for biochemical analysis of HCV RNA synthesis by a multisubunit membrane-associated replicase and for evaluating potential antiviral agents identified in biochemical or cell-based screens.
Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
