High-resolution cryo-EM using beam-image shift at 200 keV.
High-resolution cryo-EM using beam-image shift at 200 keV.
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DOI：
10.1107/s2052252520013482
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发表时间：
2020-11-01
期刊：
影响因子：
3.9
通讯作者：
Cianfrocco MA
中科院分区：
文献类型：
--
作者：
Cash JN;Kearns S;Li Y;Cianfrocco MA
在这项工作中，研究表明，通过计算校正可以克服 200 keV 光束图像偏移冷冻电镜引起的显着显微镜像差，从而将醛缩酶的结构从 4.9 Å 分辨率提高到 2.8 Å。单颗粒冷冻电子显微镜 (cryo-EM) 数据收集的最新进展利用光束图像移位来提高吞吐量。尽管在 300 keV 冷冻电镜仪器上实现，但仍不清楚光束图像偏移数据收集对 200 keV 仪器数据质量的影响程度以及像差可通过计算校正的程度。为了测试这一点，使用光束图像位移在 200 keV 下收集了醛缩酶的冷冻电镜数据集并进行了分析。该分析表明，仪器光束倾斜和粒子运动最初将分辨率限制为 4.9 Å。在RELION中经过粒子抛光和迭代像差校正后，可以获得2.8 Å分辨率的结构。该分析表明，显微镜像差的软件校正可以显着提高 200 keV 的分辨率。
In this work, it is shown that significant microscope aberrations caused by beam-image-shift cryo-EM at 200 keV can be overcome using computational correction, improving a structure of aldolase from 4.9 to 2.8 Å resolution. Recent advances in single-particle cryo-electron microscopy (cryo-EM) data collection utilize beam-image shift to improve throughput. Despite implementation on 300 keV cryo-EM instruments, it remains unknown how well beam-image-shift data collection affects data quality on 200 keV instruments and the extent to which aberrations can be computationally corrected. To test this, a cryo-EM data set for aldolase was collected at 200 keV using beam-image shift and analyzed. This analysis shows that the instrument beam tilt and particle motion initially limited the resolution to 4.9 Å. After particle polishing and iterative rounds of aberration correction in RELION, a 2.8 Å resolution structure could be obtained. This analysis demonstrates that software correction of microscope aberrations can provide a significant improvement in resolution at 200 keV.