Probing RNA Conformations Using a Polymer-Electrolyte Solid-State Nanopore.
Probing RNA Conformations Using a Polymer-Electrolyte Solid-State Nanopore.
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DOI：
10.1021/acsnano.2c08312
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发表时间：
2022-12-27
期刊：
影响因子：
17.1
通讯作者：
Actis, Paolo
中科院分区：
文献类型：
--
作者：
Chau, Chalmers;Marcuccio, Fabio;Soulias, Dimitrios;Edwards, Martin Andrew;Tuplin, Andrew;Radford, Sheena E.;Hewitt, Eric;Actis, Paolo
纳米孔系统已经成为具有单分子分辨率的生物分子复合物分析的领先平台。生物分子（如RNA）的构象高度依赖于电解质组成，但固态纳米孔系统通常需要高盐浓度来操作，从而排除了在生理相关条件下对大分子构象的分析。在这里，我们报告了基于溶解在50%w/v聚（乙二醇）（PEG）中的碱金属卤化物盐的聚合物-电解质固态纳米孔系统的实施，以增强我们的系统的性能。我们表明，聚合物电解质浴支配的分析物的易位动力学与浴中使用的盐的物理性质。这使我们能够将CsBr鉴定为补充PEG以产生最大信号增强的最佳盐。利用聚合物电解质的影响，我们探测了生理相关条件下基孔肯雅病毒（CHIKV）RNA基因组片段的构象。我们的系统能够指纹CHIKV RNA片段的长度范围从10300到102000 nt，并随后区分共转录折叠和天然重折叠的102000 nt CHIKV RNA之间的构象。我们设想，聚合物-电解质固态纳米孔系统将进一步实现在生理相关条件下对单个生物分子的结构和构象分析。
Nanopore systems have emerged as a leading platform for the analysis of biomolecular complexes with single-molecule resolution. The conformation of biomolecules, such as RNA, is highly dependent on the electrolyte composition, but solid-state nanopore systems often require high salt concentration to operate, precluding analysis of macromolecular conformations under physiologically relevant conditions. Here, we report the implementation of a polymer–electrolyte solid-state nanopore system based on alkali metal halide salts dissolved in 50% w/v poly(ethylene) glycol (PEG) to augment the performance of our system. We show that polymer–electrolyte bath governs the translocation dynamics of the analyte which correlates with the physical properties of the salt used in the bath. This allowed us to identify CsBr as the optimal salt to complement PEG to generate the largest signal enhancement. Harnessing the effects of the polymer–electrolyte, we probed the conformations of the Chikungunya virus (CHIKV) RNA genome fragments under physiologically relevant conditions. Our system was able to fingerprint CHIKV RNA fragments ranging from ∼300 to ∼2000 nt length and subsequently distinguish conformations between the co-transcriptionally folded and the natively refolded ∼2000 nt CHIKV RNA. We envision that the polymer–electrolyte solid-state nanopore system will further enable structural and conformational analyses of individual biomolecules under physiologically relevant conditions.
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影响因子：
10.8
作者：
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通讯作者：
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影响因子：
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