Abstract
As one of the essential enzymes for viral genome replication, the hepatitis C virus NS3 helicase is one of the best characterized RNA helicases to date in understanding the mechanistic cycles in a helicase-catalyzed strand separation reaction. Recently, single-molecule studies on NS3, in particular the use of optical tweezers with sub-base pair spatial resolution, have allowed people to examine the potential elementary steps of NS3 in unwinding the double-stranded RNA fueled by ATP binding and hydrolysis. In this chapter, I detail the essential technical elements involved in conducting a high-resolution optical tweezers study of NS3 helicase, starting from the purification of the recombinant helicase protein from E. coli to setting up a high-resolution single-molecule experiment using optical tweezers.
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Acknowledgments
This work was supported by NSF CAREER Award CHE1149670 to WC, NIH Director’s New Innovator Award 1DP2OD008693 to WC, and also in part by Research Grant No. 5-FY10-490 to WC from the March of Dimes Foundation.
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Cheng, W. (2015). Mechanisms of HCV NS3 Helicase Monitored by Optical Tweezers. In: Boudvillain, M. (eds) RNA Remodeling Proteins. Methods in Molecular Biology, vol 1259. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2214-7_15
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DOI: https://doi.org/10.1007/978-1-4939-2214-7_15
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