Single-Molecule Studies Reveal Dynamics of DNA Unwinding by the Ring-Shaped T7 Helicase

Helicases are molecular motors that separate DNA strands for efficient replication of genomes. We probed the kinetics of individual ring-shaped T7 helicase molecules as they unwound double-stranded DNA (dsDNA) or translocated on single-stranded DNA (ssDNA). A distinctive DNA sequence dependence was...

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Bibliographic Details
Published in:Cell 2007-06, Vol.129 (7), p.1299-1309
Main Authors: Johnson, Daniel S., Bai, Lu, Smith, Benjamin Y., Patel, Smita S., Wang, Michelle D.
Format: Article
Language:English
Subjects:
Bacteriophage T7 - genetics
Biomechanical Phenomena
DNA
DNA - biosynthesis
DNA - genetics
DNA Helicases - genetics
DNA Replication - genetics
DNA, Single-Stranded - genetics
DNA, Single-Stranded - metabolism
Mechanotransduction, Cellular - genetics
Models, Molecular
Molecular Motor Proteins - genetics
Molecular Structure
PROTEINS
Stress, Mechanical
Time Factors
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Summary:Helicases are molecular motors that separate DNA strands for efficient replication of genomes. We probed the kinetics of individual ring-shaped T7 helicase molecules as they unwound double-stranded DNA (dsDNA) or translocated on single-stranded DNA (ssDNA). A distinctive DNA sequence dependence was observed in the unwinding rate that correlated with the local DNA unzipping energy landscape. The unwinding rate increased ∼10-fold (approaching the ssDNA translocation rate) when a destabilizing force on the DNA fork junction was increased from 5 to 11 pN. These observations reveal a fundamental difference between the mechanisms of ring-shaped and nonring-shaped helicases. The observed force-velocity and sequence dependence are not consistent with a simple passive unwinding model. However, an active unwinding model fully supports the data even though the helicase on its own does not unwind at its optimal rate. This work offers insights into possible ways helicase activity is enhanced by associated proteins.
ISSN:0092-8674
1097-4172
DOI:10.1016/j.cell.2007.04.038