Department of Chemistry, University of Turku, Finland.
Chembiochem. 2012 Dec 21;13(18):2690-700. doi: 10.1002/cbic.201200629. Epub 2012 Nov 26.
A 2'-O-methyl-RNA oligonucleotide containing a single free 2'-OH group flanking a branching phosphotriester linkage was prepared as a model for phosphate-branched RNA by using an orthogonally protected dimeric phosphoramidite building block in solid-phase synthesis. The strategy allows the synthesis of phosphate-branched oligonucleotides, the three branches of which may be of any desired sequence. Hydrolytic reactions of the phosphotriester linkages in such oligonucleotides were studied at physiological pH in the presence (and absence) of various complementary oligonucleotides. The fully hybridized oligonucleotide model is an order of magnitude more stable than its single-stranded counterpart, which, in turn, is an order of magnitude more stable than its trinucleoside phosphotriester core lacking any oligonucleotide arms. Furthermore, kinked structures obtained by hybridizing the phosphate-branched oligonucleotide with partially complementary oligonucleotides are three to five times more stable than fully double-stranded ones and only approximately three times less stable than the so-called RNA X structure, which has been postulated to incorporate an RNA phosphotriester linkage. The results indicate that when the intrinsically unstable RNA phosphotriester linkage is embedded in an oligonucleotide of appropriate tertiary structure, its half-life can be at least several hours.
一种含有单个游离 2'-OH 基团的 2'-O-甲基 RNA 寡核苷酸，其侧翼为分支膦酸三酯键，通过固相合成中使用正交保护的二聚体亚磷酰胺构建块，被制备为磷酸酯分支 RNA 的模型。该策略允许合成磷酸酯分支寡核苷酸，其三个分支可以具有任何所需的序列。在生理 pH 下，研究了在存在（和不存在）各种互补寡核苷酸时，此类寡核苷酸中膦酸三酯键的水解反应。完全杂交的寡核苷酸模型比其单链对应物稳定一个数量级，而单链对应物又比其缺少任何寡核苷酸臂的三核苷酸膦酸三酯核心稳定一个数量级。此外，通过使磷酸酯分支寡核苷酸与部分互补寡核苷酸杂交而获得的扭结结构比完全双链结构稳定三到五倍，并且仅比所谓的 RNA X 结构稳定约三倍，该结构已被假定包含 RNA 膦酸三酯键。结果表明，当内在不稳定的 RNA 膦酸三酯键嵌入适当的三级结构的寡核苷酸中时，其半衰期至少可以是几个小时。
