Ecole Polytechnique Fédérale de Lausanne (EPFL), Laboratory of Molecular Nanodynamics, CH-1015, Lausanne, Switzerland.
Nat Commun. 2023 Sep 13;14(1):5649. doi: 10.1038/s41467-023-41444-x.
Observing proteins as they perform their tasks has largely remained elusive, which has left our understanding of protein function fundamentally incomplete. To enable such observations, we have recently proposed a technique that improves the time resolution of cryo-electron microscopy (cryo-EM) to microseconds. Here, we demonstrate that microsecond time-resolved cryo-EM enables observations of fast protein dynamics. We use our approach to elucidate the mechanics of the capsid of cowpea chlorotic mottle virus (CCMV), whose large-amplitude motions play a crucial role in the viral life cycle. We observe that a pH jump causes the extended configuration of the capsid to contract on the microsecond timescale. While this is a concerted process, the motions of the capsid proteins involve different timescales, leading to a curved reaction path. It is difficult to conceive how such a detailed picture of the dynamics could have been obtained with any other method, which highlights the potential of our technique. Crucially, our experiments pave the way for microsecond time-resolved cryo-EM to be applied to a broad range of protein dynamics that previously could not have been observed. This promises to fundamentally advance our understanding of protein function.
观察蛋白质执行其任务的情况在很大程度上仍然难以实现，这使得我们对蛋白质功能的理解从根本上不完整。为了实现这种观察，我们最近提出了一种将冷冻电子显微镜（cryo-EM）的时间分辨率提高到微秒级的技术。在这里，我们证明了微秒时间分辨的 cryo-EM 能够观察到快速的蛋白质动力学。我们使用这种方法来阐明豇豆花叶病毒（CCMV）衣壳的力学性质，其大幅度的运动在病毒生命周期中起着至关重要的作用。我们观察到 pH 值的跃变导致衣壳的扩展构象在微秒时间尺度上收缩。虽然这是一个协同的过程，但衣壳蛋白的运动涉及不同的时间尺度，导致反应路径弯曲。很难想象，如果使用任何其他方法，如何能够获得这样一个详细的动力学图像，这凸显了我们技术的潜力。至关重要的是，我们的实验为微秒时间分辨的 cryo-EM 应用于以前无法观察到的广泛的蛋白质动力学铺平了道路。这有望从根本上推进我们对蛋白质功能的理解。
