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bioRxiv · 10.1101/798280

High-resolution cryo-EM reconstructions in the presence of substantial aberrations

Bromberg, R.·Guo, Y.·Borek, D.·Otwinowski, Z.
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Abstract

The beam-image shift method accelerates data acquisition in cryo-EM single particle reconstruction (cryo-EM SPR) by fast repositioning of the imaging area, but at the cost of more severe and complex optical aberrations.\n\nWe analyze here how uncorrected anti-symmetric aberrations, such as coma and trefoil, affect cryo-EM SPR results, and then infer an analytical formula quantifying information loss due to their presence that explains why Fourier-shell coefficient (FSC)-based statistics may report significantly overestimated resolution if these aberrations are not fully corrected. We validate our analysis with reference-based aberration refinement for two cryo-EM SPR datasets acquired with a 200 kV microscope in the presence of coma exceeding 40 {micro}m, and obtained 2.3 and 2.7 [A] reconstructions for 144 and 173 kDa particles, respectively.\n\nOur 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 acceptable in cryo-EM SPR experiments.

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Bromberg, R., Guo, Y., Borek, D., Otwinowski, Z.. 2019-10-08. High-resolution cryo-EM reconstructions in the presence of substantial aberrations. https://doi.org/10.1101/798280

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