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Light-coupled cryo-plunger for time-resolved cryo-EM.
Journal of Structural Biology ( IF 3.0 ) Pub Date : 2020-09-17 , DOI: 10.1016/j.jsb.2020.107624
Nate Yoder 1 , Farzad Jalali-Yazdi 1 , Sigrid Noreng 1 , Alexandra Houser 1 , Isabelle Baconguis 1 , Eric Gouaux 2
Affiliation  

Proteins are dynamic molecules that can undergo rapid conformational rearrangements in response to stimuli. These structural changes are often critical to protein function, and thus elucidating time-dependent conformational landscapes has been a long-standing goal of structural biology. To harness the power of single particle cryo-EM methods to enable ‘time-resolved’ structure determination, we have developed a light-coupled cryo-plunger that pairs flash-photolysis of caged ligands with rapid sample vitrification. The ‘flash-plunger’ consists of a high-power ultraviolet LED coupled with focusing optics and a motorized linear actuator, enabling the user to immobilize protein targets in vitreous ice within a programmable time window – as short as tens of milliseconds – after stimulus delivery. The flash-plunger is a simple, inexpensive and flexible tool to explore short-lived conformational states previously unobtainable by conventional sample preparation methods.



中文翻译:

用于时间分辨冷冻电镜的光耦合冷冻柱塞。

蛋白质是动态分子,可以响应刺激进行快速的构象重排。这些结构变化通常对蛋白质功能至关重要,因此阐明时间依赖性构象景观一直是结构生物学的长期目标。为了利用单粒子冷冻 EM 方法的力量来实现“时间分辨”结构测定,我们开发了一种光耦合冷冻柱塞,可将笼配体的闪光光解与快速样品玻璃化相结合。“闪光柱塞”由高功率紫外线 LED 与聚焦光学器件和电动线性致动器组成,使用户能够在刺激传递后的可编程时间窗口内(短至数十毫秒)将蛋白质目标固定在玻璃冰中. 闪光柱塞是一个简单的,

更新日期:2020-10-02
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