Synthesis of phenazine-tethered arabino and xylofuranosyl oligonucleotide conjugates: The thermal stability and fluorescence properties of their duplexes (DNA-DNA & DNA-RNA) & triplexes

Abstract

The synthesis of phenazine (Pzn) tethered ara-U and xylo-U incorporated oligonucleotides, and their properties as DNA-DNA, DNA-RNA duplexes and as triplexes are reported. 2′- O-Pzn-tethered ara-U amidite ( 11) and Pzn-linked amidite block ( 4) as well as CPG solid supports functionalised with 2′- O-Pzn-tethered ara-U ( 10a – 10c) and 3′- O-Pzn-tethered xylo-U succinates ( 22a – 22c) were used in the solid-phase DNA synthesis to prepare modified 9mers 25 – 29, 31, 41 – 45 (for DNA-DNA & DNA-DRA duplex studies) and 18mers 35 – 40, 46 – 50 (for the DNA triplex studies). Thermal melting experiments with the resulting duplexes and triplexes showed that all oligo-DNAs, except for the middle-modified ones ( 31 & 40), have enhanced affinity to the DNA and RNA targets ( 23& 52) as well as for the DNA duplex target 32·33. Pzn-tethered ara-U block was more efficient at the 3′-terminal of oligonucleotides than at the 5′-end in the duplexes and especially in triplexes, where it provided a dramatic improvement in the stability ( ΔT m = 16.1°C). Employment of Pzn-tethered ara-U block at the 3′-end together with the Pzn-tethered dT block at the 5′-end of the oligo-DNAs ( 29, 38) provided the best duplex and triplex stabilisation to give a highest ΔT m of 14.4°C for DNA-DNA duplexes, ΔT m of 11.7°C for DNA-RNA duplexes and ΔT m of 19.6°C for triplexes. All DNA-DNA and DNA-RNA duplexes as well as DNA triplexes, formed by the oligos modified with 2′- O-Pzn-tethered ara-U blocks showed geater stability than those formed by the oligos modified with xylo-analogs with the same length of the linker arms. For both ara- and xylo-configurations, the best DNA-DNA & DNA-RNA duplex stabilisation was provided by the short ethyl linkers, and increasing the length of the linker led to considerable destabilisation of the duplexes. In case of triplexes, longer linker arms were required to obtain better stabilisation. Hexyl linker provided the highest triplex stabilisation for the oligonucleotides modified with Pzn-tethered ara-U block ( ΔT m = 16.5°C) and butyl linker was found to be most suitable for the oligo-DNAs modified with Pzn-tehtered xylo-U block ( ΔT m = 12.3°C). Fluorescence studies showed that Pzn behaves as a weak exterior binder upon DNA-DNA or DNA-RNA duplex or DNA triplex formation which accounts for moderate changes in the fluorescent properties of the Pzn moiety (ΔF for DNA-DNA and DNA-RNA duplexes = ±0.2, ΔF for triplexes = 1.4 – 2.5). Employment of Pzn at both 3′ and 5′ ends of the oligonucleotides (as in 27, 29, 38 & 39) provided the greatest duplex and triplex stabilisation so far, and led to more effective interaction between the Pzn moieties and the double and triple helixes (ΔF for DNA-DNA and DNA-RNA duplexes = 4, ΔF for triplexes = 5).