State Key Laboratory of Analytical Chemistry for Life Sciences, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing, China.
Chemistry and Biomedicine Innovation Center (ChemBIC), Nanjing University, Nanjing, China.
Nat Commun. 2021 Jun 7;12(1):3368. doi: 10.1038/s41467-021-23764-y.
Folding of RNA can produce elaborate tertiary structures, corresponding to their diverse roles in the regulation of biological activities. Direct observation of RNA structures at high resolution in their native form however remains a challenge. The large vestibule and the narrow constriction of a Mycobacterium smegmatis porin A (MspA) suggests a sensing mode called nanopore trapping/translocation, which clearly distinguishes between microRNA, small interfering RNA (siRNA), transfer RNA (tRNA) and 5 S ribosomal RNA (rRNA). To further profit from the acquired event characteristics, a custom machine learning algorithm is developed. Events from measurements with a mixture of RNA analytes can be automatically classified, reporting a general accuracy of ~93.4%. tRNAs, which possess a unique tertiary structure, report a highly distinguishable sensing feature, different from all other RNA types tested in this study. With this strategy, tRNAs from different sources are measured and a high structural conservation across different species is observed in single molecule.
RNA 的折叠可以产生精细的三级结构，这与其在调节生物活性方面的多种功能相对应。然而，在其天然状态下以高分辨率直接观察 RNA 结构仍然是一个挑战。巨大的前庭和分枝杆菌 smegmatis 孔蛋白 A（MspA）的狭窄收缩提示了一种称为纳米孔捕获/易位的传感模式，这种模式可以清楚地区分 microRNA、小干扰 RNA（siRNA）、转移 RNA（tRNA）和 5S 核糖体 RNA（rRNA）。为了进一步利用所获得的事件特征，开发了一种定制的机器学习算法。来自混合 RNA 分析物测量的事件可以自动分类，报告的总体准确性约为 93.4%。具有独特三级结构的 tRNA 报告了一个高度可区分的传感特征，与本研究中测试的所有其他 RNA 类型都不同。通过这种策略，从不同来源测量 tRNA，并在单分子中观察到不同物种之间的高度结构保守性。
