格罗尔
低温电子显微
微流控
分辨率(逻辑)
折叠(DSP实现)
材料科学
生物系统
毫秒
时间分辨率
纳米技术
化学
物理
计算机科学
光学
生物
生物化学
工程类
大肠杆菌
人工智能
天文
电气工程
基因
作者
Stefania Torino,Mugdha Dhurandhar,Annelore Stroobants,Raf Claessens,Rouslan G. Efremov
出处
期刊:Nature Methods
[Springer Nature]
日期:2023-08-17
卷期号:20 (9): 1400-1408
被引量:4
标识
DOI:10.1038/s41592-023-01967-z
摘要
Single-particle cryogenic electron microscopy (cryo-EM) allows reconstruction of high-resolution structures of proteins in different conformations. Protein function often involves transient functional conformations, which can be resolved using time-resolved cryo-EM (trEM). In trEM, reactions are arrested after a defined delay time by rapid vitrification of protein solution on the EM grid. Despite the increasing interest in trEM among the cryo-EM community, making trEM samples with a time resolution below 100 ms remains challenging. Here we report the design and the realization of a time-resolved cryo-plunger that combines a droplet-based microfluidic mixer with a laser-induced generator of microjets that allows rapid reaction initiation and plunge-freezing of cryo-EM grids. Using this approach, a time resolution of 5 ms was achieved and the protein density map was reconstructed to a resolution of 2.1 Å. trEM experiments on GroEL:GroES chaperonin complex resolved the kinetics of the complex formation and visualized putative short-lived conformations of GroEL–ATP complex. This article describes a time-resolved cryo-plunger that combines a droplet-based microfluidic mixer with a laser-induced generator of microjets that allows rapid reaction initiation and plunge-freezing of cryo-EM grids. A time resolution of 5 ms was achieved using this approach.
科研通智能强力驱动
Strongly Powered by AbleSci AI