Puffer Barbara, Moroder Holger, Aigner Michaela, Micura Ronald
Institute of Organic Chemistry, Center for Molecular Biosciences Innsbruck (CMBI), Leopold Franzens University, Innrain 52a, 6020 Innsbruck, Austria.
Nucleic Acids Res. 2008 Feb;36(3):970-83. doi: 10.1093/nar/gkm880. Epub 2007 Dec 20.
Site-specifically modified 2'-methylseleno RNA represents a valuable derivative for phasing of X-ray crystallographic data. Several successful applications in three-dimensional structure determination of nucleic acids, such as the Diels-Alder ribozyme, have relied on this modification. Here, we introduce synthetic routes to 2'-methylseleno phosphoramidite building blocks of all four standard nucleosides, adenosine, cytidine, guanosine and uridine, that are tailored for 2'-O-bis(acetoxyethoxy)methyl (ACE) RNA solid-phase synthesis. We additionally report on their incorporation into oligoribonucleotides including deprotection and purification. The methodological expansion of 2'-methylseleno labeling via ACE RNA chemistry is a major step to make Se-RNA generally accessible and to receive broad dissemination of the Se-approach for crystallographic studies on RNA. Thus far, preparation of 2'-methylseleno-modified oligoribonucleotides has been restricted to the 2'-O-[(triisopropylsilyl)oxy]methyl (TOM) and 2'-O-tert-butyldimethylsilyl (TBDMS) RNA synthesis methods.
位点特异性修饰的2'-甲基硒代RNA是用于X射线晶体学数据相位分析的一种有价值的衍生物。在核酸三维结构测定中的一些成功应用，如狄尔斯-阿尔德核酶，都依赖于这种修饰。在此，我们介绍了针对2'-O-双（乙酰氧基乙氧基）甲基（ACE）RNA固相合成定制的四种标准核苷（腺苷、胞苷、鸟苷和尿苷）的2'-甲基硒代亚磷酰胺构建块的合成路线。我们还报告了它们掺入寡核糖核苷酸的情况，包括脱保护和纯化。通过ACE RNA化学方法扩展2'-甲基硒代标记是使硒代RNA普遍可用并使硒方法在RNA晶体学研究中得到广泛传播的重要一步。到目前为止，2'-甲基硒代修饰的寡核糖核苷酸的制备仅限于2'-O-[(三异丙基甲硅烷基)氧基]甲基（TOM）和2'-O-叔丁基二甲基甲硅烷基（TBDMS）RNA合成方法。
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