Nucleic Acids Research Advance Access originally published online on April 9, 2009
Nucleic Acids Research 2009 37(11):3580-3587; doi:10.1093/nar/gkp208
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Nucleic Acids Research, 2009, Vol. 37, No. 11 3580-3587
© 2009 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.
Molecular Biology |
mwr Xer site-specific recombination is hypersensitive to DNA supercoiling
1Bionanotechnology IRC Department of Physics, 2Division of Molecular Genetics, Department of Biochemistry, University of Oxford, Oxford OX1 3QU, UK, 3Center for Applied Biotechnology Studies, Department of Biological Science, College of Natural Science and Mathematics, California State University Fullerton, Fullerton, CA 92834-6850, USA and 4Division of Molecular Genetics, Faculty of Biomedical and Life Sciences, University of Glasgow, Bower Building, University Ave, Glasgow G12 8QQ, Scotland, UK
*To whom correspondence should be addressed. Tel: +44 1865 272269; Fax: +44 1865 272400; Email: s.trigueros1{at}physics.ox.ac.uk
Received October 29, 2008. Revised February 24, 2009. Accepted March 11, 2009.
The multiresistance plasmid pJHCMW1, first identified in a Klebsiella pneumoniae strain isolated from a neonate with meningitis, includes a Xer recombination site, mwr, with unique characteristics. Efficiency of resolution of mwr-containing plasmid dimers is strongly dependent on the osmotic pressure of the growth medium. An increase in supercoiling density of plasmid DNA was observed as the osmotic pressure of the growth culture decreased. Reporter plasmids containing directly repeated mwr, or the related cer sites were used to test if DNA topological changes were correlated with significant changes in efficiency of Xer recombination. Quantification of Holliday junctions showed that while recombination at cer was efficient at all levels of negative supercoiling, recombination at mwr became markedly less efficient as the level of supercoiling was reduced. These results support a model in which modifications at the level of supercoiling density caused by changes in the osmotic pressure of the culture medium affects resolution of mwr-containing plasmid dimers, a property that separates mwr from other Xer recombination target sites.