Kinetic stability of intermolecular DNA quadruplexes

¹ University of Southampton

^* To whom correspondence should be addressed. E-mail: k.r.fox{at}soton.ac.uk.

Submitted on February 14, 2005
Revised on March 19, 2005
Accepted on 13 April 2005

	Abstract

Fluorescently labelled oligodeoxyribonucleotides containing a single tract of four successive guanines have been used to study the thermodynamic and kinetic properties of short intermolecular DNA quadruplexes. When these assemble to form intermolecular quadruplexes the fluorophores are in close proximity and the fluorescence is quenched. On raising the temperature these complexes dissociate and there is a large increase in fluorescence. These complexes are exceptionally stable in potassium containing buffers, and possess Tm values that are too high to measure. Tm values were determined in sodium containing buffers for which the rate of reannealing is extremely slow; the melting profiles are effectively irreversible and the apparent melting temperatures are dependent on the rates of heating. The dissociation kinetics of these complexes were estimated by rapidly increasing the temperature and following the time dependent changes in fluorescence. From these data we have estimated the half-lives of these quadruplexes at 37°C. Addition of a T to the unlabelled end of the oligonucleotide increases quadruplex stability. In contrast addition of a T between the fluorophore and the oligonucleotide leads to a decrease in stability.

Key Words: DNA melting, G-quartet, dissociation kinetics, quadruplex

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