Improvement of the NMR structural quality for RNA and DNA

核磁共振波谱（NMR）对DNA和RNA的表征非常重要，包括各种方法，提供对这些生物分子的结构，功能，相互作用和构象动力学的基本见解。尽管核磁共振在这一领域做出了杰出的贡献，但对于核酸来说，一些基本的工具和方法（通常用于蛋白质）仍然缺失，需要进一步发展或微调。值得注意的是，用于核酸结构测定的力场参数化仍然缺乏。因此，推进这一领域的发展对于阐明更精确、高质量、高分辨率的核磁共振结构是必不可少的。这种方法需要用表征良好的RNA和DNA参比系统进行验证。我们的目标是改进现有的方法和协议，并提供高质量的数据来优化力场参数化。在这方面，我们已经利用丰富的结构数据阐明了两种RNA发夹模型系统（四联体UUCG和CUUG）的结构，并结合了在水中的优化纯化潜力。UUCG结构目前被认为是通过与其他方法和经验数据交叉验证进行力场优化和评估的黄金标准。我们目前正在研究额外的四足虫（GCAA， GAAG）的结构和动力学。此外，我们从目前对RNA病毒（Covid-19和西尼罗河病毒）基因组元素以及其他小DNA和RNA基序和重复序列的广泛研究中提供了额外合适的模型靶点。为了真实地描述RNA结构的整体特性，我们研究了CUUG四环系统，尽管它具有很高的热稳定性，但它具有大量的动力学特性。因此，我们最近将核磁共振与分子动力学模拟相结合，以探测潜在的构象景观并提供合适的结构集合。在这个应用程序中，我们将包括更大的RNA和DNA分子，g -四重结构和RNA/DNA三重结构。为了确定它们的结构，我们将收集更多的全局、远程结构、取向和动态约束数据，并利用三维结构预测和同源建模来评估核酸力场并改进工具和方案。此外，我们的目标是包括最重要的和发生的RNA和DNA修饰以及几种离子，并计划采用和开发构象集合的结构计算方法。这些新的和改进的参数和方法将在已建立的软件中提供给结构核磁共振社区，并通过在我们现有的核磁共振结构确定门户网站服务中实现。此外，我们将把结果发表在合适的开源数据库、期刊和平台上。

Nuclear Magnetic Resonance spectroscopy (NMR) is very important for the characterization of DNA and RNA, encompassing a variety of methods that offer essential insight on the structure, function, interactions and conformational dynamics of these biomolecules. Despite the eminent contribution of NMR in this field, some essential tools and approaches (which are routinely available for proteins) are still missing for nucleic acids and require further development or fine-tuning. Notably, the parametrization of forcefields used for the structure determinations of nucleic acids still lacks behind. Advancing the development in this area is therefore indispensable to elucidate more accurate and high-quality well resolved NMR structures. Such approach requires verification with well characterized RNA and DNA reference systems. Our objective is to improve the currently established methods and protocols and provide high quality data to optimize the forcefield parametrization. In this respect, we already elucidated the structure of two RNA hairpin model systems (the tetraloops UUCG and CUUG) using a wealth of structural data and incorporated optimized potentials for refinement in water. The UUCG structure is currently considered to be the gold standard for forcefield optimizations and assessment by cross-validation with other methods and empirical data. We currently investigate the structure and dynamics of additional tetraloops (GCAA, GAAG). Moreover, we provide additional suitable model targets from our current extensive investigations of elements from the genome of RNA viruses (Covid-19 and West Nile Virus) and other small DNA and RNA motifs and repeats. In our endeavor to describe the ensemble characteristic of RNA structures realistically, we studied the CUUG tetraloop system, which turned out to feature substantial dynamics despite its high thermal stability. Thus, we have recently combined NMR with molecular dynamic simulations to probe the underlying conformational landscape and provide suitable structural ensembles. In this application, we will include larger RNA and DNA molecules, G-Quadruplexes, and RNA/DNA triplexes. For their structure determination, we will gather additional global, long-range structural, orientational and dynamic restraint data and utilize 3D structure prediction and homology modeling for evaluation of the nucleic acid forcefield and improvement of tools and protocols. Moreover, we aim to include the most important and occurring RNA and DNA modifications as well as several ions and plan to employ and develop approaches for the structure calculation of conformational ensembles. These new and improved parameters and methods will be made available for the structural NMR community in established software and through implementation in our existing web-portal service for NMR structure determination. In addition, we will publish the results in suitable open source databases, journals and platforms.