4.2 Article

Application of super-resolution and correlative double sampling in cryo-electron microscopy

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FARADAY DISCUSSIONS
卷 240, 期 -, 页码 261-276

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ROYAL SOC CHEMISTRY
DOI: 10.1039/d2fd00049k

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  1. Wellcome Trust
  2. MRC
  3. BBSRC

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Developments in cryo-electron tomography and sub-tomogram averaging show exciting potential in achieving near-atomic resolution structures in protein determination. The application of super-resolution mode in single particle analysis and cryo-ET STA data collection yield different resolution benefits, with considerations needed for specific experimental conditions.
Developments in cryo-EM have allowed atomic or near-atomic resolution structure determination to become routine in single particle analysis (SPA). However, near-atomic resolution structures determined using cryo-electron tomography and sub-tomogram averaging (cryo-ET STA) are much less routine. In this paper, we show that collecting cryo-ET STA data using the same conditions as SPA, with both correlated double sampling (CDS) and the super-resolution mode, allowed apoferritin to be reconstructed out to the physical Nyquist frequency of the images. Even with just two tilt series, STA yields an apoferritin map at 2.9 angstrom resolution. These results highlight the exciting potential of cryo-ET STA in the future of protein structure determination. While processing SPA data recorded in super-resolution mode may yield structures surpassing the physical Nyquist limit, processing cryo-ET STA data in the super-resolution mode gave no additional resolution benefit. We further show that collecting SPA data in the super-resolution mode, with CDS activated, reduces the estimated B-factor, leading to a reduction in the number of particles required to reach a target resolution without compromising the data size on disk and the area imaged in SerialEM. However, collecting SPA data in CDS does reduce throughput, given that a similar resolution structure, with a slightly larger B-factor, is achievable with optimised parameters for speed in EPU (without CDS).

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