Nucleic Acids Research Advance Access published online on December 19, 2006
Nucleic Acids Research, doi:10.1093/nar/gkl1078
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Nucleic Acid Enzymes |
Spermidine biases the resolution of Holliday junctions by phage 
integrase
Department of Biology and Center for Microbial Sciences, San Diego State University San Diego, CA 92182-4614, USA
*To whom correspondence should be addressed. Tel: +1 619 594 4490; Fax: +1 619 594 5676; Email: asegall{at}sunstroke.sdsu.edu
Received August 16, 2006. Revised November 21, 2006. Accepted November 21, 2006.
Holliday junctions are a central intermediate in diverse pathways of DNA repair and recombination. The isomerization of a junction determines the directionality of the recombination event. Previous studies have shown that the identity of the central sequence of the junction may favor one of the two isomers, in turn controlling the direction of the pathway. Here we demonstrate that, in the absence of DNA sequence-mediated isomer preference, polycations are the major contributor to biasing strand cleavage during junction resolution. In the case of wild-type phage 
excision junctions, spermidine plays the dominant role in controlling the isomerization state of the junction and increases the rate of junction resolution. Spermidine also counteracts the sequence-imposed bias on resolution. The spermidine-induced bias is seen equally on supercoiled and linear excisive recombination junction intermediates, and thus is not just an artefact of in vitro recombination conditions. The contribution of spermidine requires the presence of accessory factors, and results in the repositioning of Int's core-binding domains on junctions, perhaps due to DNA-spermidine–protein interactions, or by influencing DNA conformation in the core region. Our results lead us to propose that spermidine together with accessory factors promotes the formation of the second junction isomer. We propose that this rearrangement triggers the activation of the second pair of Int active sites necessary to resolve Holliday junctions during phage Int-mediated recombination.
Present address: Geoffrey D. Cassell, Nativis Inc., 10975 N. Torrey Pines Rd. Suite 150, La Jolla, CA 92037, USA
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