Abstract
Natural oligonucleotides have many rotatable single bonds, and thus their structures are inherently flexible. Structural flexibility leads to an entropic loss when unwound oligonucleotides form a duplex with single-stranded DNA or RNA. An effective approach to reduce such entropic loss in the duplex-formation is the conformational restriction of the flexible phosphodiester linkage and/or sugar moiety. We here report the synthesis and biophysical properties of a novel artificial nucleic acid bearing an oxanorbornane scaffold (OxNorNA), where the adamant oxanorbornane was expected to rigidify the structures of both the linkage and sugar parts of nucleic acid. OxNorNA phosphoramidite with a uracil (U) nucleobase was successfully synthesized over 15 steps from a known sugar-derived cyclopentene. Thereafter, the given phosphoramidite was incorporated into the designed oligonucleotides. Thermal denaturation experiments revealed that oligonucleotides modified with the conformationally restricted OxNorNA-U properly form a duplex with the complementally DNA or RNA strands, although the Tm values of OxNorNA-U-modified oligonucleotides were lower than those of the corresponding natural oligonucleotides. As we had designed, entropic loss during the duplex-formation was reduced by the OxNorNA modification. Moreover, the OxNorNA-U-modified oligonucleotide was confirmed to have extremely high stability against 3′-exonuclease activity, and its stability was even higher than those of the phosphorothioate-modified counterparts (Sp and Rp). With the overall biophysical properties of OxNorNA-U, we expect that OxNorNA could be used for specialized applications, such as conformational fixation and/or bio-stability enhancement of therapeutic oligonucleotides (e.g., aptamers).
Highlights
The structural flexibility of natural oligonucleotides contributes to the formation of various higher-order structures
∆Tm /mod.: the change in Tm value (∆Tm ) per modification compared to the unmodified oligonucleotide. ∆G◦
We evaluated mismatch discrimination of the oxanorbornane scaffold (OxNorNA)-U-modified oligonucleotide
Summary
The structural flexibility of natural oligonucleotides contributes to the formation of various higher-order structures. Chemical modifications that restrict the structures of flexible phosphodiester linkages and/or sugar moieties to those seen in A-form DNA·RNA duplexes. A 0number of conformationally restricted artificial nucleic acids, for which a duplex, and example the 2 ,4is0 -BNA/LNA-modified exhibit a locked high duplex-forming representative. Henessian and co-workers have successfully achieved of both the sugar ringring andsystem the torsion demonstrated that oligonucleotides modified with restrictions tricyclo-DNA, in which a fused rigidifies the torsion angles γ and δ, exhibit increased ssRNA affinity relative to the natural the torsion angles 1γ [13]. The relative(Figure position of the phosphodiester artificial nucleic acid bearing oxanorbornane scaffold. 2artificial nucleic acid bearing an oxanorbornane scaffold (OxNorNA) (Figure 2). Structures of artificial nucleic acid bearing an oxanorbornane scaffold (OxNorNA), isoDNA, 2.
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