Kinetic studies on the formation of DNA triplexes containing the nucleoside analogue 2'-O-(2-aminoethyl)-5-(3-amino-1-propynyl)uridine |
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Authors: | Rusling David A Broughton-Head Victoria J Tuck Alex Khairallah Hannah Osborne Sadie D Brown Tom Fox Keith R |
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Institution: | School of Biological Sciences, University of Southampton, Bassett Crescent East, Southampton, SO16 7PX, UK. k.r.fox@soton.ac.uk. |
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Abstract: | We have examined the kinetics of triple helix formation of oligonucleotides that contain the nucleotide analogue 2'-O-(2-aminoethyl)-5-(3-amino-1-propynyl)uridine (bis-amino-U, BAU), which forms very stable base triplets with AT. Triplex stability is determined by both the number and location of the modifications. BAU-containing oligonucleotides generate triplexes with extremely slow kinetics, as evidenced by 14 degrees C hysteresis between annealing and melting profiles even when heated at a rate as slow as 0.2 degrees C min(-1). The association kinetics were measured by analysis of the hysteresis profiles, temperature-jump relaxation and DNase I footprinting. We find that the slow kinetics are largely due to the decreased rate of dissociation; BAU modification has little effect on the association reaction. The sequence selectivity is also due to the slower dissociation of BAU from AT than other base pairs. |
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