Bromberg Raquel, Guo Yirui, Borek Dominika, Otwinowski Zbyszek
Department of Biophysics, The University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
IUCrJ. 2020 Mar 26;7(Pt 3):445-452. doi: 10.1107/S2052252520002444. eCollection 2020 May 1.
Here, an analysis is performed of how uncorrected antisymmetric aberrations, such as coma and trefoil, affect cryo-EM single-particle reconstruction (SPR) results, and an analytical formula quantifying information loss owing to their presence is inferred that explains why Fourier-shell coefficient-based statistics may report significantly overestimated resolution if these aberrations are not fully corrected. The analysis is validated with reference-based aberration refinement for two cryo-EM SPR data sets acquired with a 200 kV microscope in the presence of coma exceeding 40 µm, and 2.3 and 2.7 Å reconstructions for 144 and 173 kDa particles, respectively, were obtained. The results provide a description of an efficient approach for assessing information loss in cryo-EM SPR data acquired in the presence of higher order aberrations, and address inconsistent guidelines regarding the level of aberrations that is acceptable in cryo-EM SPR experiments.
在此，我们分析了未校正的反对称像差（如彗差和三叶草像差）如何影响冷冻电镜单颗粒重构（SPR）结果，并推导了一个量化由于这些像差的存在而导致信息损失的解析公式，该公式解释了如果这些像差未得到充分校正，基于傅里叶壳层相关系数的统计数据为何可能报告分辨率被显著高估的情况。我们通过基于参考的像差精修对在200 kV显微镜下采集的两个冷冻电镜SPR数据集进行了验证，其中存在超过40 µm的彗差，分别获得了144 kDa和173 kDa颗粒的2.3 Å和2.7 Å重构结果。这些结果描述了一种评估在存在高阶像差情况下采集的冷冻电镜SPR数据中信息损失的有效方法，并解决了关于冷冻电镜SPR实验中可接受像差水平的不一致指南问题。
