RNA STRUCTURE DETERMINATION BY DOUBLE QUANTUM COHERENCE ESR

该子项目是利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得主要资金， 因此可以在其他CRISP条目中表示。列出的机构是 中心，不一定是研究者的机构。 RNA研究是全球基因组学项目的基础。它在翻译储存在DNA中的遗传信息并通过核糖体机制表现出来方面起着关键作用。病毒和细菌RNA的研究是RNA研究的重要组成部分，并为疾病控制提供了见解。在RNA测序以及获得3-D结构及其与RNA功能的关系方面有大量的持续努力。在我们最初的努力中，我们应用DQC ESR来测量A型RNA中的端到端距离。 三个RNA与13，16，和26个碱基对的自旋标记的3-碘-乙酰氨基-乙酰基连接到硫尿苷。 从DQC评估了5 '-5'距离，得到5-5'连接的氮氧化合物上的电子自旋之间的距离分别为40、55和72“，这与提出的结构非常一致。描述双量子滤波重聚焦初级回波（DQF-RPE）对26 bp RNA的应用的简短通信在J. Am.与S.Butcher正在进行的合作，准备将联合收割机NOE和DQC约束结合起来，以改善结构细化。 这将包括一个30 kDa的RNA同源二聚体与螺旋包装基序称为四环受体的研究。 该基序在许多不同核酶的三级结构形成中起重要作用。 第二种感兴趣的RNA结构来源于参与翻译移码的HIV-1茎环结构域（Staple和Butcher，NAR 2003，PDB代码1 PJY）。 这种RNA结构对于病毒复制是必不可少的，但是在未知结构的较大（45个核苷酸）RNA的情况下发生。结构中的不确定性是两个螺旋之间的关系，这两个螺旋被3个核苷酸的嘌呤“铰链”区分开。根据分子模型，分子的长度约为71 º。然而，该距离是未知的，并且如果铰链区域如预期的那样在两个螺旋之间引起弯曲，则该距离可能小于71。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Studies of RNA are fundamental for the global genomics project. It plays a key role in translating genetic information stored in DNA and manifesting it via the ribosome machinery. The study of viral and bacterial RNA is an important component of RNA research and offers insights into disease control. There is a large, ongoing effort in RNA sequencing as well as efforts to obtain 3-D structures and their relation to RNA function. In our initial effort, we applied DQC ESR to measure end-to-end distances in A-form RNA. Three RNAs with 13, 16, and 26 base pairs were spin-labeled by 3-iodo-acetamido-proxyl attached to thiouridine. The 5'-5' distances were evaluated from DQC, yielding the distances between electron spins on 5-5' attached nitroxides of 40, 55, and 72¿, respectively, which agrees well with the proposed structures. A short communication describing the application of double-quantum filtered refocused primary echo (DQF-RPE) to 26 b.p. RNA is in press in J. Am. Chem. Soc. The ongoing collaboration with S.Butcher, is poised to combine NOE and DQC constraints to improve structure refinement. This will include the study of a 30 kDa RNA homodimer with a helical packing motif known as the tetraloop receptor. This motif plays an important role in tertiary structure formation in many different ribozymes. The second RNA structure of interest is derived from HIV-1 stem-loop domain involved in translational frameshifting (Staple and Butcher, NAR 2003, pdb code 1PJY). This RNA structure is essential for viral replication, but occurs in the context of a larger (45 nucleotide) RNA of unknown structure. The uncertainty in the structure is the relationship between the two helices, which are separated by a 3 nucleotide purine "hinge" region. The length of the molecule, based on molecular modeling, is approximately 71¿. However, this distance is unknown and may be less than 71¿ if the hinge region causes a bend between the two helices, as expected.