SOLID STATE NMR STUDIES OF DNA AND RNA DYNAMICS

长期以来，人们一直接受详细的结构信息是 为了充分理解双链DNA 与转录因子相互作用，通常执行完整的 其生物学功能的范围。因此，我们对 DNA的结构随着时间的推移而演变，因为高分辨率 用X射线衍射法和溶液核磁共振法获得DNA的结构模型 显示标准B形式的特定序列变体。它是 合理地问一问，如何同样详细地描述内部 DNA的动力学将改变我们对这些结构变异的看法 因此，我们对DNA如何发挥其生物学功能的理解。 固体核磁共振是探测局域动力学的理想技术 从近溶液条件到结晶状态的生物聚合物 环境。此外，一些新的固体核磁共振技术 近些年来已经发展起来，使得能够定义 具有高准确度和精密度的生物分子结构 样品条件范围广，没有分子量限制。在……里面 在接下来的五年里，我们计划使用固态核磁共振结构和 用动力学技术定义结构与结构之间的关系(S) DNA和RNA中的动力学和功能。具体地说，我们建议 以下五个阶段的计划：1)使用固体核磁共振来探索 含DNA低聚物局部动力学的序列特异性 限制性内切酶、转录因子和 甲基转移酶；2)用固体核磁共振研究动态 DNA甲基化对结构的影响，特别是在富含CpG的DNA中 序列；3)利用固体核磁共振技术研究动力学 以及锤头和发夹的催化位的结构 核酶；4)继续发展和优化固体核磁共振 高分子量核糖核酸的结构解析技术 酸。将这些核磁共振技术扩展到17.6特斯拉 多共振CPMAS探头的构建，从而有助于 完成特定目标1-4；5)合成所有同位素标记 实现特定目标1-4所需的DNA和RNA亚磷酰胺。

It has been long accepted that detailed structural information is necessary in order to fully understand the means by which duplex DNA interacts with transcription factors, and in general performs the full range of its biological functions. Accordingly, our view of the structure of DNA has evolved through the years, as high resolution structural models of DNA obtained by X-ray diffraction and solution NMR display sequence specific variations from the canonical B form. It is reasonable to ask how an equally detailed picture of the internal dynamics of DNA would modify our view of these structural variations and thus our understanding of how DNA performs its biological functions. Solid state NMR is an ideal technique for probing the localized dynamics of biopolymers from near solution conditions to the crystalline environment. In addition, a number of novel solid state NMR techniques have been developed in recent years that enable the definition of biomolecular structure with high accuracy and precision over the same broad range of sample conditions, with no molecular weight limit. In the next five years we plan to use solid state NMR structural and dynamical techniques to define the relationship(s) between structure, dynamics and function in DNA and RNA. Specifically we propose the following five stage program: 1) Use solid state NMR to explore the sequence specificity of localized dynamics in DNA oligomers containing binding sites for restriction enzymes, transcription factors and methyltransferases; 2) Use solid state NMR to investigate the dynamic and structural impact of DNA methylation, especially in CpG-rich DNA sequences; 3) Use solid state NMR techniques to investigate the dynamics and structure of the catalytic sites of the hammerhead and hairpin ribozymes; 4) Continue to develop and optimize solid state NMR techniques for structure elucidation in high molecular weight nucleic acids. Extend these NMR techniques to 17.6 Tesla by completing construction of a multi-resonant CPMAS probe, thus aiding in the completion of specific goals 1-4; 5) Synthesize all isotopically labeled DNA and RNA phosphoramidites required to achieve specific goals 1-4.