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Science 27 July 2007:
Vol. 317. no. 5837, pp. 513 - 516
DOI: 10.1126/science.1144130
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Reports

Spring-Loaded Mechanism of DNA Unwinding by Hepatitis C Virus NS3 Helicase

Sua Myong,1,2* Michael M. Bruno,3,4 Anna M. Pyle,3,4 Taekjip Ha1,2,4

NS3, an essential helicase for replication of hepatitis C virus, is a model enzyme for investigating helicase function. Using single-molecule fluorescence analysis, we showed that NS3 unwinds DNA in discrete steps of about three base pairs (bp). Dwell time analysis indicated that about three hidden steps are required before a 3-bp step is taken. Taking into account the available structural data, we propose a spring-loaded mechanism in which several steps of one nucleotide per adenosine triphosphate molecule accumulate tension on the protein-DNA complex, which is relieved periodically via a burst of 3-bp unwinding. NS3 appears to shelter the displaced strand during unwinding, and, upon encountering a barrier or after unwinding >18 bp, it snaps or slips backward rapidly and repeats unwinding many times in succession. Such repetitive unwinding behavior over a short stretch of duplex may help to keep secondary structures resolved during viral genome replication.

1 Physics Department, University of Illinois, 1110 West Green Street, Urbana, IL 61801, USA.
2 Institute for Genomic Biology, University of Illinois, 1206 West Gregory Drive, Urbana, IL 61801, USA.
3 Molecular Biophysics and Biochemistry, Yale University, 266 Whitney Avenue, Room 334A, Bass Building, New Haven, CT 06511, USA.
4 Howard Hughes Medical Institute.

* To whom correspondence should be addressed. E-mail: smyong{at}uiuc.edu

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THIS ARTICLE HAS BEEN CITED BY OTHER ARTICLES:
Hepatitis C Virus NS3 Helicase Forms Oligomeric Structures That Exhibit Optimal DNA Unwinding Activity in Vitro.
B. Sikora, Y. Chen, C. F. Lichti, M. K. Harrison, T. A. Jennings, Y. Tang, A. J. Tackett, J. B. Jordan, J. Sakon, C. E. Cameron, et al. (2008)
J. Biol. Chem. 283, 11516-11525
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Conformational changes of a Swi2/Snf2 ATPase during its mechano-chemical cycle.
R. Lewis, H. Durr, K.-P. Hopfner, and J. Michaelis (2008)
Nucleic Acids Res. 36, 1881-1890
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Real-time observation of bacteriophage T4 gp41 helicase reveals an unwinding mechanism.
T. Lionnet, M. M. Spiering, S. J. Benkovic, D. Bensimon, and V. Croquette (2007)
PNAS 104, 19790-19795
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The Serine Protease Domain of Hepatitis C Viral NS3 Activates RNA Helicase Activity by Promoting the Binding of RNA Substrate.
R. K. F. Beran, V. Serebrov, and A. M. Pyle (2007)
J. Biol. Chem. 282, 34913-34920
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MCM Forked Substrate Specificity Involves Dynamic Interaction with the 5'-Tail.
E. Rothenberg, M. A. Trakselis, S. D. Bell, and T. Ha (2007)
J. Biol. Chem. 282, 34229-34234
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Science. ISSN 0036-8075 (print), 1095-9203 (online)