Department of Chemistry, Duke University, Durham, North Carolina 27708, United States.
J Phys Chem B. 2024 Jun 13;128(23):5590-5600. doi: 10.1021/acs.jpcb.4c01710. Epub 2024 May 29.
A viral infection process covers a large range of spatiotemporal scales. Tracking the viral infection process with fluorescent labels over long durations while maintaining a fast sampling rate requires bright and highly photostable labels. StayGold is a recently identified green fluorescent protein that has a greater photostability and higher signal intensity under identical illumination conditions compared to existing fluorescence protein variants. Here, StayGold protein fusions were used to generate virus-like particles (StayGold-VLPs) to achieve hour-long 3D single-virus tracking (SVT) with 1000 localizations per second (kHz sampling rate) in live cells. The expanded photon budget from StayGold protein fusions prolonged the tracking duration, facilitating a comprehensive study of viral trafficking dynamics with high temporal resolution over long time scales. The development of StayGold-VLPs presents a simple and general VLP labeling strategy for better performance in SVT, enabling exponentially more information to be collected from single trajectories and allowing for the future possibility of observing the entire life cycle of a single virus.
病毒感染过程涵盖了很大的时空尺度。要想在长时间内用荧光标记跟踪病毒感染过程，同时保持快速的采样率，就需要明亮且高度稳定的标记。StayGold 是一种新发现的绿色荧光蛋白，与现有的荧光蛋白变体相比，在相同的照明条件下，它具有更高的稳定性和更高的信号强度。在这里，将 StayGold 蛋白融合用于生成病毒样颗粒（StayGold-VLPs），以实现在活细胞中以每秒 1000 个局部化的速度进行长达 1 小时的 3D 单病毒跟踪（SVT）。StayGold 蛋白融合扩展了光子预算，延长了跟踪时间，从而可以在长时间尺度上以高时间分辨率全面研究病毒运输动力学。StayGold-VLPs 的开发为 SVT 提供了一种简单而通用的 VLPs 标记策略，从而可以从单个轨迹中收集更多的信息，并为将来观察单个病毒的整个生命周期提供了可能。
