TRIPLE-HELIX FORMATION BY AN OLIGONUCLEOTIDE CONTAINING ONE (DA)12 AND 2 (DT)12 SEQUENCES BRIDGED BY 2 HEXAETHYLENE GLYCOL CHAINS

The triple-helix formation by the oligonucleotide (dA)12-x-(dT)12-x-(dT)12, where x is a hexaethylene glycol group, was investigated by thermal denaturation analysis and circular dichroism spectroscopy. Thermal denaturation analysis showed that this single-stranded oligonucleotide is able to fold back on itself twice to give a triple helix at low temperature. Upon an increase in the temperature, two cooperative transitions were observed: formation of a double-stranded structure with a dangling x-(dT)12 extremity, then formation of a single-stranded coil structure. Due to the intramolecular character of the transition, the triplex is much more stable than that formed by the reference mixture (dA)12 + 2(dT)12. In 0.1 M NaCl, the triplex-to-coil transition occurred at about 30-degrees-C whereas the duplex-to-coil was at about 60-degrees-C. Upon an increase in the salt, the increase of temperature corresponding to the triplex-to-duplex transition was larger than that of the duplex-to-coil transition. MgCl2 showed higher efficiency than NaCl to promote triplex or duplex formation. The thermodynamic parameters DELTA-H and DELTA-S were determined at various ionic strengths for both transitions. Both the enthalpy change and entropy change associated with triplex-to-duplex transition (Hoogsteen base pairing) were smaller than those associated to the duplex-to-coil transition (Watson-Crick base pairing). When the ionic strength increased, the parameters -DELTA-H and -DELTA-S showed a very small decrease for the duplex-to-coil transition whereas a strong increase was observed with the triplex-to-duplex transition. A total of 3.5 +/- 0.3 sodium ions are thermodynamically released in the triplex-to-duplex transition, whereas this number is 2.3 +/- 0.3 in the duplex-to-coil transition. CD spectroscopy demonstrated that the conformation of the triplex structure of (dA)12-x-(dT)12-x-(dT)12 was essentially identical to that of the reference mixture (dA)12 + 2(dT)12. These results open up new prospects for the use of this type of oligonucleotide in studies of triple helix properties.