Department of Biochemistry and Biophysics, Texas A&M University, College Station, Texas 77843, USA.
Center for Phage Technology, College Station, Texas 77843, USA.
RNA. 2020 Dec;26(12):1755-1766. doi: 10.1261/rna.071100.119. Epub 2020 Aug 21.
Ribonucleic acids (RNAs) play essential roles in living cells. Many of them fold into defined three-dimensional (3D) structures to perform functions. Recent advances in single-particle cryo-electron microscopy (cryo-EM) have enabled structure determinations of RNA to atomic resolutions. However, most RNA molecules are structurally flexible, limiting the resolution of their structures solved by cryo-EM. In modeling these molecules, several computational methods are limited by the requirement of massive computational resources and/or the low efficiency in exploring large-scale structural variations. Here we use hierarchical natural move Monte Carlo (HNMMC), which takes advantage of collective motions for groups of nucleic acid residues, to refine RNA structures into their cryo-EM maps, preserving atomic details in the models. After validating the method on a simulated density map of tRNA, we applied it to objectively obtain the model of the folding intermediate for the specificity domain of ribonuclease P from and refine a flexible ribosomal RNA (rRNA) expansion segment from the () ribosome in different conformational states. Finally, we used HNMMC to model atomic details and flexibility for two distinct conformations of the complete genomic RNA (gRNA) inside MS2, a single-stranded RNA virus, revealing multiple pathways for its capsid assembly.
核糖核酸 (RNA) 在活细胞中发挥着重要作用。其中许多 RNA 能够折叠成特定的三维 (3D) 结构，从而发挥其功能。近年来，单颗粒冷冻电子显微镜 (cryo-EM) 的发展使得 RNA 结构的测定能够达到原子分辨率。然而，大多数 RNA 分子结构具有柔韧性，这限制了 cryo-EM 解析其结构的分辨率。在对这些分子进行建模时，几种计算方法受到大量计算资源的要求和/或在探索大规模结构变化方面效率低下的限制。在这里，我们使用分层自然移动 Monte Carlo (HNMMC) 方法，该方法利用核酸残基的集体运动，将 RNA 结构细化到其 cryo-EM 图谱中，在模型中保留原子细节。在对 tRNA 的模拟密度图进行验证后，我们将其应用于客观获得核糖核酸酶 P 特异性结构域折叠中间体的模型，并细化来自不同构象状态的 （）核糖体中的柔性核糖体 RNA (rRNA) 扩展片段。最后，我们使用 HNMMC 对 MS2 内部完整基因组 RNA (gRNA) 的两种不同构象进行建模，以揭示其衣壳组装的多种途径，MS2 是一种单链 RNA 病毒。
