Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan.
Department of Chemistry, Graduate School of Science, Kyoto University, Kyoto, Japan.
Methods Mol Biol. 2021;2323:221-232. doi: 10.1007/978-1-0716-1499-0_16.
RNA-protein (RNP) complexes are promising biomaterials for the fields of nanotechnology and synthetic biology. Protein-responsive RNA sequences (RNP motifs) can be integrated into various RNAs, such as messenger RNA, short-hairpin RNA, and synthetic RNA nanoobjects for a variety of purposes. Direct observation of RNP interaction in solution at high resolution is important in the design and construction of RNP-mediated nanostructures. Here we describe a method to construct and visualize RNP nanostructures that precisely arrange a target protein on the RNA scaffold with nanometer scale. High-speed AFM (HS-AFM) images of RNP nanostructures show that the folding of RNP complexes of defined sizes can be directly visualized at single RNP resolution in solution.
RNA-蛋白质（RNP）复合物是纳米技术和合成生物学领域有前途的生物材料。蛋白质反应性 RNA 序列（RNP 基序）可以整合到各种 RNA 中，如信使 RNA、短发夹 RNA 和合成 RNA 纳米物体，用于各种目的。在高分辨率下直接观察溶液中的 RNP 相互作用对于设计和构建 RNP 介导的纳米结构非常重要。在这里，我们描述了一种构建和可视化 RNP 纳米结构的方法，该方法可以将目标蛋白质精确地排列在 RNA 支架上，达到纳米级尺度。RNP 纳米结构的高速原子力显微镜（HS-AFM）图像表明，在溶液中可以以单个 RNP 的分辨率直接可视化确定大小的 RNP 复合物的折叠。
