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Studies on the stability of multistranded DNA

Studies on the stability of multistranded DNA
Studies on the stability of multistranded DNA

A novel technique, using fluorescently-labelled oliogonucleotides, has been developed to measure the thermal stability of quadruplex-forming DNA sequences. This has been used to compare the stability of human (TTAGGG)n and Oxytricha (G4T4)n telomeric repeats and to examine their kinetic properties. These sequences are stabilised more by potassium than sodium and the Oxytricha sequence is the most stable. Both sequences produce hysteresis between the melting and annealing profiles, especially at higher rates of temperature change, indicative of slow dynamics of folding and unfolding. These dynamics were studied by temperature-jump experiments, which confirmed that the kinetics of these telomeric repeats is very slow. Both sequences show slower dynamics in potassium than sodium, though the kinetic pathway is complicated, requiring two exponentials for its complete description. These kinetics are much slower for the Oxytricha  repeat, which displays a t1/2 of about 30 minutes even at the melting temperature.

The interaction of a series of substituted anthraquinones with quadruplex DNA was studied by fluorescence melting experiment and bandshift analysis. Structure-activity relationships for these compounds were derived by comparing closely related ligands.  These studies showed that the propionamide sidechains are an important determinant of the activity and showed that (i) 2,6- substitution of the anthraquinone ring is better than 2,7- substitution; (ii) attachment via the nitrogen rather than carbon of the amide of the propionamide sidechains is often superior; (iii) longer sidechains are better especially if they contain positive charges and (iv) bulky groups in the sidechains are generally unfavourable.

The final part of this thesis describes the (unsuccessful) development of a novel SELEX methodology (REPSA) for identifying strong triplex binding sites from a pool of potential target sites.

University of Southampton
Brown, Nicholas Matthew
Brown, Nicholas Matthew

Brown, Nicholas Matthew (2003) Studies on the stability of multistranded DNA. University of Southampton, Doctoral Thesis.

Record type: Thesis (Doctoral)

Abstract

A novel technique, using fluorescently-labelled oliogonucleotides, has been developed to measure the thermal stability of quadruplex-forming DNA sequences. This has been used to compare the stability of human (TTAGGG)n and Oxytricha (G4T4)n telomeric repeats and to examine their kinetic properties. These sequences are stabilised more by potassium than sodium and the Oxytricha sequence is the most stable. Both sequences produce hysteresis between the melting and annealing profiles, especially at higher rates of temperature change, indicative of slow dynamics of folding and unfolding. These dynamics were studied by temperature-jump experiments, which confirmed that the kinetics of these telomeric repeats is very slow. Both sequences show slower dynamics in potassium than sodium, though the kinetic pathway is complicated, requiring two exponentials for its complete description. These kinetics are much slower for the Oxytricha  repeat, which displays a t1/2 of about 30 minutes even at the melting temperature.

The interaction of a series of substituted anthraquinones with quadruplex DNA was studied by fluorescence melting experiment and bandshift analysis. Structure-activity relationships for these compounds were derived by comparing closely related ligands.  These studies showed that the propionamide sidechains are an important determinant of the activity and showed that (i) 2,6- substitution of the anthraquinone ring is better than 2,7- substitution; (ii) attachment via the nitrogen rather than carbon of the amide of the propionamide sidechains is often superior; (iii) longer sidechains are better especially if they contain positive charges and (iv) bulky groups in the sidechains are generally unfavourable.

The final part of this thesis describes the (unsuccessful) development of a novel SELEX methodology (REPSA) for identifying strong triplex binding sites from a pool of potential target sites.

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Published date: 2003

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Local EPrints ID: 465297
URI: http://eprints.soton.ac.uk/id/eprint/465297
PURE UUID: 56c74fcd-c1d8-43e0-8495-fd6aaab1baf3

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Date deposited: 05 Jul 2022 00:35
Last modified: 05 Jul 2022 04:50

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Author: Nicholas Matthew Brown

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