Nucleic Acids Research Advance Access originally published online on February 21, 2008
Nucleic Acids Research 2008 36(7):2301-2310; doi:10.1093/nar/gkn035
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Nucleic Acids Research, 2008, Vol. 36, No. 7 2301-2310
© 2008 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Nucleic Acid Enzymes |
Single-molecule observations of topotecan-mediated TopIB activity at a unique DNA sequence
1Kavli Institute of Nanoscience, Faculty of Applied Sciences, Delft University of Technology, Lorentzweg 1, 2628 CJ Delft, The Netherlands, 2Department of Molecular Pharmacology, St. Jude Children's Research Hospital, 332 N. Lauderdale, Memphis, TN 38105, USA, 3BioQuant, Soft Matter and Biological Physics, University of Heidelberg, Im Neuenheimer Feld 267, 69120 Heidelberg, Germany, 4Laboratoire ‘Régulation et dynamique des génomes’ UMR 5153 CNRS-Muséum National d’Histoire Naturelle USM0503 and 5INSERM UR565; 43 rue Cuvier, 75231 Paris cedex 05, France
* To whom correspondence should be addressed. Tel: +31 15 2783219; Fax: +31 15 2781202; Email: n.h.dekker{at}tudelft.nl
Received December 18, 2007. Revised January 16, 2008. Accepted January 21, 2008.
The rate of DNA supercoil removal by human topoisomerase IB (TopIB) is slowed down by the presence of the camptothecin class of antitumor drugs. By preventing religation, these drugs also prolong the lifetime of the covalent TopIB–DNA complex. Here, we use magnetic tweezers to measure the rate of supercoil removal by drug-bound TopIB at a single DNA sequence in real time. This is accomplished by covalently linking camptothecins to a triple helix-forming oligonucleotide that binds at one location on the DNA molecule monitored. Surprisingly, we find that the DNA dynamics with the TopIB–drug interaction restricted to a single DNA sequence are indistinguishable from the dynamics observed when the TopIB–drug interaction takes place at multiple sites. Specifically, the DNA sequence does not affect the instantaneous supercoil removal rate or the degree to which camptothecins increase the lifetime of the covalent complex. Our data suggest that sequence-dependent dynamics need not to be taken into account in efforts to develop novel camptothecins.
Present address: Daniel A. Koster, Departments of Physics of Complex Systems and Molecular Cell Biology, Weizmann Institute of Science, Rehovot, 76100 Israel