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.
- Base Pair Mismatch
- Base Sequence
- Deoxyribonuclease I/chemistry
- Peptide Mapping
- Time Factors
- Transition Temperature
- Uridine/analogs & derivatives