Synthesis of 1'-phenazine-tethered psicofuranosyl oligonucleotides: The thermal stability and fluorescence properties of their duplexes and triplexes

Article abstract of DOI:10.1016/S0040-4020(98)00237-3

The synthesis of modified oligonucleotides (ODNs), tethered with phenazine (pzn) at Cl' of 1-(3'-deoxypsicofuranosyl)uracil, the thermal stability, and fluorescence properties of their duplexes and triplexes are described. The key intermediates, the psicofuranosylracil derivatives 7 and 8 with phenazine 1'-tethered through a linker composed of a phosphate and two methylene groups, were synthesized from 1-(3'-deoxypsicofuranosyl)uracil 5(5 → 6 (26%) → 7 (62%) or 8 (70%)]. Compound 7 was converted into the corresponding 6'-O-DMTr protected 4'-phosphoramidite block 13 in three steps [7 → 9 (83%) → 11 (83%) → 13 (83%)] whereas compound 8 was used in the preparation of the modified solid support 14 in four steps 8 → 10 (28%) → 12 (70%) → 14]. Modified 9-mer ODNs 28-31 and 18-mer ODNs 22-25 were then assembled in a usual manner using automated solid-phase DNA synthesis protocol. The phenazine-tethered 9-mers (28-31) were tested for their ability to form stable duplexes with target DNA-strands (19-20). The phenazine- tethered 18-mers (22-25) were tested for their ability to form scale triplexes with 24-mer and 29-mer duplex targets (15-16 and 17-18). Triplexes consisting of modified ODNs with pzn at the 5'-terminal (as in 24) and at 3',5'-terminals (as in 25) were more stable than the unmodified parent triplex. No triplex was found to have formed with modified ODNs with pzn attached at 3'- or at the middle of the strand at neutral pH (7.3), but triplex formation was observed at acidic pH (6.0) although they were less stable than the unmodified parent triplex. The same trend was observed for duplexes. The fluorescence intensity of pzn in the modified triplexes was enhanced and blue-shifted by -13 nm relative to the single strand. In contrast, the changes in fluorescence intensities of pzm in the modified duplexes were relatively less compared to the triplexes. The fluorescence intensity increased proportionally as the thermal stabilities of the triplexes increased. A comparison of the fluorescent intensity changes (ΔF) shows that the fluorophore in duplexes (ΔF?-1.2 to +1.5) experiences relatively minor change in the microenvironment compared to that of the triplexes (ΔF? 1.5 to 4.5). Nevertheless, in both cases the phenazine residue most probably interacts with the neighbouring nucleobases as a weak exterior binder.