Tripepi Andrea, Shakoor Huma, Klapetek Petr
Central European Institute of Technology, Masaryk University, Brno, Czech Republic.
National Center of Biomolecular Research, Brno, Czech Republic.
Curr Protoc. 2025 Jan;5(1):e70084. doi: 10.1002/cpz1.70084.
Atomic force microscopy (AFM) has recently received increasing interest in molecular biology. This technique allows quick and reliable detection of biomolecules. However, studying RNA-protein complexes using AFM poses significant challenges. Here, we describe a simple and reliable method to visualize positively charged proteins bound to RNA that does not require metallic cations. This method allowed us to effectively detect and visualize Staufen-RNA complexes by height or logarithmic stiffness. The study of the mechanical properties is particularly important in the case of protein-coated RNA complexes, where RNA cannot be detected by height channel. In any case, it is necessary to compare AFM data with the data derived from other techniques like nuclear magnetic resonance, X-ray crystallography, cryogenic electron microscopy, and small-angle X-ray scattering. © 2025 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Preparation and visualization of RNA-protein complex.
原子力显微镜（AFM）最近在分子生物学领域受到越来越多的关注。这项技术能够快速且可靠地检测生物分子。然而，使用AFM研究RNA-蛋白质复合物面临重大挑战。在此，我们描述了一种简单可靠的方法，用于可视化与RNA结合的带正电荷蛋白质，该方法无需金属阳离子。此方法使我们能够通过高度或对数刚度有效地检测和可视化Staufen-RNA复合物。在蛋白质包被的RNA复合物中，由于无法通过高度通道检测到RNA，因此对其力学性质的研究尤为重要。无论如何，有必要将AFM数据与来自其他技术（如核磁共振、X射线晶体学、低温电子显微镜和小角X射线散射）的数据进行比较。© 2025作者。由Wiley Periodicals LLC出版的《当前方案》。基本方案：RNA-蛋白质复合物的制备与可视化。
