Department of Biochemistry, University of Zurich, Winterthurerstrasse 190, 8057 Zurich, Switzerland.
Aix Marseille Univ, CNRS, Centrale Marseille, Institut Fresnel, 13013 Marseille, France.
J Am Chem Soc. 2022 Jan 12;144(1):52-56. doi: 10.1021/jacs.1c09387. Epub 2021 Dec 31.
Single-molecule Förster resonance energy transfer (FRET) is a versatile technique for probing the structure and dynamics of biomolecules even in heterogeneous ensembles. However, because of the limited fluorescence brightness per molecule and the relatively long fluorescence lifetimes, probing ultrafast structural dynamics in the nanosecond time scale has thus far been very challenging. Here, we demonstrate that nanophotonic fluorescence enhancement in zero-mode waveguides enables measurements of previously inaccessible low-nanosecond dynamics by dramatically improving time resolution and reduces data acquisition times by more than an order of magnitude. As a prototypical example, we use this approach to probe the dynamics of a short intrinsically disordered peptide that were previously inaccessible with single-molecule FRET measurements. We show that we are now able to detect the low-nanosecond correlations in this peptide, and we obtain a detailed interpretation of the underlying distance distributions and dynamics in conjunction with all-atom molecular dynamics simulations, which agree remarkably well with the experiments. We expect this combined approach to be widely applicable to the investigation of very rapid biomolecular dynamics.
单分子Förster 共振能量转移（FRET）是一种非常有用的技术，即使在异质体系中也可以用来探测生物分子的结构和动态。然而，由于每个分子的荧光亮度有限，并且荧光寿命相对较长，因此迄今为止探测纳秒时间尺度的超快结构动态一直是非常具有挑战性的。在这里，我们证明零模波导中的纳米光子荧光增强可通过显著提高时间分辨率并将数据采集时间减少一个数量级以上来测量以前无法测量的低纳秒动态。作为一个典型的例子，我们使用这种方法来探测以前无法用单分子 FRET 测量的短天然无序肽的动力学。我们表明，我们现在能够检测到这种肽中的低纳秒相关性，并且我们结合原子分子动力学模拟获得了对基础距离分布和动力学的详细解释，这与实验结果非常吻合。我们预计这种组合方法将广泛适用于研究非常快速的生物分子动力学。
