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Updated: May 29, 2026

A Protocol for Analyzing Hepatitis C Virus Replication
Published on: June 26, 2014
Single-base pair unwinding and asynchronous RNA release by the hepatitis C virus NS3 helicase
Wei Cheng1, Srikesh G Arunajadai, Jeffrey R Moffitt
1Department of Pharmaceutical Sciences, College of Pharmacy, University of Michigan, Ann Arbor, MI 48109, USA. chengwe@umich.edu
Abstract:
Nonhexameric helicases use adenosine triphosphate (ATP) to unzip base pairs in double-stranded nucleic acids (dsNAs). Studies have suggested that these helicases unzip dsNAs in single-base pair increments, consuming one ATP molecule per base pair, but direct evidence for this mechanism is lacking. We used optical tweezers to follow the unwinding of double-stranded RNA by the hepatitis C virus NS3 helicase. Single-base pair steps by NS3 were observed, along with nascent nucleotide release that was asynchronous with base pair opening. Asynchronous release of nascent nucleotides rationalizes various observations of its dsNA unwinding and may be used to coordinate the translocation speed of NS3 along the RNA during viral replication.
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