Marchanka Alexander, Carlomagno Teresa
Centre for Biomolecular Drug Research (BMWZ) and Institute of Organic Chemistry, Leibniz University Hannover, Hannover, Germany.
Centre for Biomolecular Drug Research (BMWZ) and Institute of Organic Chemistry, Leibniz University Hannover, Hannover, Germany; Helmholtz Centre for Infection Research, Group of NMR-based Structural Chemistry, Braunschweig, Germany.
Methods Enzymol. 2019;615:333-371. doi: 10.1016/bs.mie.2018.08.029. Epub 2018 Dec 7.
RNA structure is essential to understand RNA function and regulation in cellular processes. RNA acts either in isolation or as part of a complex with proteins. Both isolated and protein-complexed RNA represent a challenge for structural biology, due to the complexity of its conformational space and intrinsic dynamics. NMR, with its capability to cope with dynamic structures, is the optimal technique to study RNA conformation, ideally in solution. However, RNA-protein complexes are often very large, which limits the application of solution-state NMR. In this chapter we describe the methodology that we developed to determine the structure of RNA by solid-state NMR. Solid-state NMR is not limited by large molecular weights and can be optimally applied to study both the RNA and the protein components of large RNA-protein complexes. We review the methods for resonance assignments, collection of RNA-specific distance restraints, as well as detection of protein-RNA interfaces.
RNA结构对于理解细胞过程中RNA的功能和调控至关重要。RNA既可以单独发挥作用，也可以作为与蛋白质复合物的一部分。由于其构象空间的复杂性和内在动力学，孤立的RNA和与蛋白质复合的RNA对结构生物学来说都是一项挑战。核磁共振（NMR）能够处理动态结构，是研究RNA构象的理想技术，最好是在溶液中进行。然而，RNA-蛋白质复合物通常非常大，这限制了溶液态NMR的应用。在本章中，我们描述了我们开发的通过固态NMR确定RNA结构的方法。固态NMR不受大分子重量的限制，可以最佳地应用于研究大型RNA-蛋白质复合物的RNA和蛋白质成分。我们回顾了共振归属方法、RNA特异性距离约束的收集以及蛋白质-RNA界面的检测。
