SYNTHESIS OF 15N LABELED RNA FRAGMENTS

描述：这项提议的长期目标是发展和 展示合成和利用所需的合成策略 特定修饰或标记的RNA片段。这些分子是 特定位置标记的15N和13C可提供独特的 对包括氢在内的局部RNA结构的独立于模型的洞察 粘合、质子化、水化、防止水合和堆积。 15N特殊标记核苷的用途将扩大到 将~(13)C引入嘌呤的C8位和C_2位 腺嘌呤。在一对嘌呤中只有一对与15N发生偶联 将允许区分其他类似的核磁共振共振。 此外，13C可以提供关于基座堆叠的独立信息。 开发了使用特定的15N和13C标记的RNA合成程序 将通过使它们适应亚磷酰胺而变得更广泛 方法。由于许多更简单的RNA基序类似于非正则碱基对， 单个凸起和四个凸起现在已经用15N核磁共振进行了表征， 新的核磁共振工作将主要集中在更复杂的系统上，如内部 环，大发夹环，假结，接吻环，四环受体， 核酶和蛋白质-RNA复合体。用~(15)N核磁共振定义金属 与RNA的离子结合是其结构和功能的关键方面，将是 探索过了。

DESCRIPTION: The long-term goals of this proposal are to develop and demonstrate synthetic strategies necessary for the synthesis and utilization of specifically modified or labeled RNA fragments. Molecules that are site-specifically labeled with 15N and 13C can provide unique and model-independent insight into local RNA structure, including hydrogen bonding, protonation, hydration, protection from hydration and stacking. The utility of 15N specifically labeled nucleosides will be expanded by introducing 13C into the C8 position of both purines and the C2 position of adenine. The resulting coupling with 15N in only one of a pair of purines will allow otherwise similar NMR resonances to be distinguished. Furthermore, the 13C can provide independent information on base stacking. The procedures developed for RNA synthesis using specific 15N and 13C labels will be made more widely available by adapting them to the phosphoramidite method. Since many of the simpler RNA motifs like non-canonical base pairs, single bulges, and tetraloops have now been characterized by 15N NMR, the new NMR work will focus primarily on more complex systems, such as internal loops, large hairpin loops, pseudoknots, kissing loops, tetraloop receptors, ribozymes, and protein-RNA complexes. The use of 15N NMR to define metal ion binding to RNA, a crucial aspect of its structure and function, will be explored.