Quantitative Assessment of Tip Effects in Single-Molecule High-Speed Atomic Force Microscopy Using DNA Origami Substrates.,Angewandte Chemie International Edition

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Quantitative Assessment of Tip Effects in Single-Molecule High-Speed Atomic Force Microscopy Using DNA Origami Substrates.
Angewandte Chemie International Edition ( IF 16.1 ) Pub Date : 2020-06-02 , DOI: 10.1002/anie.202005884
Charlotte Kielar _{1,

2} , Siqi Zhu ₁ , Guido Grundmeier ₁ , Adrian Keller ₁

Affiliation

High‐speed atomic force microscopy (HS‐AFM) is widely employed in the investigation of dynamic biomolecular processes at a single‐molecule level. However, it remains an open and somewhat controversial question, how these processes are affected by the rapidly scanned AFM tip. While tip effects are commonly believed to be of minor importance in strongly binding systems, weaker interactions may significantly be disturbed. Herein, we quantitatively assess the role of tip effects in a strongly binding system using a DNA origami‐based single‐molecule assay. Despite its femtomolar dissociation constant, we find that HS‐AFM imaging can disrupt monodentate binding of streptavidin (SAv) to biotin (Bt) even under gentle scanning conditions. To a lesser extent, this is also observed for the much stronger bidentate SAv–Bt complex. The presented DNA origami‐based assay can be universally employed to quantify tip effects in strongly and weakly binding systems and to optimize the experimental settings for their reliable HS‐AFM imaging.

中文翻译：

使用 DNA 折纸底物在单分子高速原子力显微镜中定量评估尖端效应。

高速原子力显微镜（HS-AFM）广泛用于研究单分子水平的动态生物分子过程。然而，快速扫描的 AFM 针尖如何影响这些过程仍然是一个开放且有些争议的问题。虽然通常认为尖端效应在强结合系统中无关紧要，但较弱的相互作用可能会受到严重干扰。在此，我们使用基于 DNA 折纸的单分子测定定量评估尖端效应在强结合系统中的作用。尽管其飞摩尔解离常数，但我们发现即使在温和的扫描条件下，HS-AFM 成像也会破坏链霉亲和素（SAv）与生物素（Bt）的单齿结合。在较小程度上，这也观察到更强的双齿 SAv-Bt 复合物。所提出的基于 DNA 折纸的测定法可以普遍用于量化强结合和弱结合系统中的尖端效应，并优化其可靠的 HS-AFM 成像的实验设置。

更新日期：2020-08-10

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