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Nucleic Acids Research 2005 33(7):2099-2105; doi:10.1093/nar/gki512
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Published online 11 April 2005

© The Author 2005. Published by Oxford University Press. All rights reserved
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions{at}oupjournals.org

Article

Torque-limited RecA polymerization on dsDNA

Thijn van der Heijden, John van Noort, Hendrikje van Leest, Roland Kanaar1,2, Claire Wyman1,2, Nynke Dekker and Cees Dekker*

Kavli Institute of Nanoscience, Delft University of Technology Lorentzweg 1, 2628 CJ Delft, The Netherlands 1Department of Cell Biology and Genetics, Erasmus Medical Center PO Box 1738, 3000 DR Rotterdam, The Netherlands 2Department of Radiation Oncology, Erasmus Medical Center PO Box 1738, 3000 DR Rotterdam, The Netherlands

*To whom correspondence should be addressed. Tel: +31 15 2786094; Fax: +31 15 2781202; Email: dekker{at}mb.tn.tudelft.nl

Received February 10, 2005. Revised March 28, 2005. Accepted March 28, 2005.

The assembly of RecA onto a torsionally constrained double-stranded DNA molecule was followed in real time using magnetic tweezers. Formation of a RecA–DNA filament on the DNA tether was stalled owing to different physical processes depending on the applied stretching force. For forces up to 3.6 pN, the reaction stalled owing to the formation of positive plectonemes in the remaining DNA molecule. Release of these plectonemes by rotation of the magnets led to full coverage of the DNA molecule by RecA. At stretching forces larger than 3.6 pN, the twist induced during filament formation caused the reaction to stall before positive supercoils were generated. We deduce a maximum built-up torsion of 10.1 ± 0.7 kbT. In vivo this built-up torsion may be used to favor regression of a stalled replication fork or to free the chromosomal DNA in E.coli from its condensing proteins.

Present address: John van Noort, Department of Biophysics, Huygens Laboratory, Leiden University, 2300 RA Leiden, The Netherlands


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