Cooperative Oligonucleotide-Directed Triple Helix Formation at Adjacent DNA Sites

The changes in binding free energies due to cooperativity for oligodeoxyribonucleotides bound to adjacent sites by triple helix formation have been determined by quantitative affinity cleavage titrations. A 20-fold enhancement in equilibrium association constant is realized for an 11 mer pyrimidine oligonucleotide binding in the presence of a neighboring bound site at 24 °C and pH 7.0 (25 mM Tris-OAc, 10 mM NaCl, 1 mM spermine). This corresponds to an increase in binding free energy of 1.8 ± 0.2 kcal mol⁻¹. This cooperativity is not observed when the two binding sites are separated by one base pair.