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利用挥发性醇类的气相扩散对大分子晶体进行高效冷冻保护

Efficient cryoprotection of macromolecular crystals using vapor diffusion of volatile alcohols.

作者信息

Farley Christopher, Juers Douglas H

机构信息

Department of Physics, Whitman College, Walla Walla, WA 99362, United States.

Department of Physics, Whitman College, Walla Walla, WA 99362, United States; Program in Biochemistry, Biophysics and Molecular Biology, Whitman College, Walla Walla, WA 99362, United States.

出版信息

J Struct Biol. 2014 Nov;188(2):102-6. doi: 10.1016/j.jsb.2014.09.011. Epub 2014 Oct 5.

Abstract

Macromolecular X-ray crystallography, usually done at cryogenic temperature to limit radiation damage, often requires liquid cryoprotective soaking that can be labor intensive and damaging to crystals. Here we describe a method for cryoprotection that uses vapor diffusion of volatile cryoprotective agents into loop-mounted crystals. The crystal is mounted into a vial containing a small volume of an alcohol-based cryosolution. After a short incubation with the looped crystal sitting in the cryosolution vapor, the crystal is transferred directly from the vial into the cooling medium. Effective for several different protein crystals, the approach obviates the need for liquid soaking and opens up a heretofore underutilized class of cryoprotective agents for macromolecular crystallography.

摘要

大分子X射线晶体学通常在低温下进行以限制辐射损伤,这往往需要进行液体冷冻保护浸泡,此过程可能 labor intensive 且对晶体有损害。在此,我们描述一种冷冻保护方法,该方法利用挥发性冷冻保护剂的蒸汽扩散到用环挂载的晶体中。将晶体安装到一个装有少量醇基冷冻溶液的小瓶中。在用环挂载的晶体在冷冻溶液蒸汽中短暂孵育后,将晶体直接从小瓶转移到冷却介质中。该方法对几种不同的蛋白质晶体均有效,避免了液体浸泡的需要,并为大分子晶体学开辟了一类迄今未充分利用的冷冻保护剂。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8ada/4252874/43685b42e235/nihms633346f1.jpg

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