Structural dynamics: review of time‐resolved cryo‐EM

The structural determination of biological macromolecules has been transformative for understanding biochemical mechanisms and developing therapeutics. However, the ultimate goal of characterizing how structural dynamics underpin biochemical processes has been difficult. This is largely due to significant technical challenges that hinder data collection and analysis on the native timescales of macromolecular dynamics. Single‐particle cryo‐EM provides a powerful platform to approach this challenge, since samples can be frozen faster than the single‐turnover timescales of most biochemical reactions. In order to enable time‐resolved analysis, significant innovations in the handling and preparation of cryo‐EM samples have been implemented, bringing us closer to the goal of the direct observation of protein dynamics in the milliseconds to seconds range. Here, the current state of time‐resolved cryo‐EM is reviewed and the most promising future research directions are discussed. Time‐resolved cryo‐EM is an emerging technique in structural biology that allows the user to capture structural states which would otherwise be too transient for standard methods. There has been a resurgence in technical advancements in this field in the last five years and this review provides a summary of the technical highlights.