Electron counting and beam-induced motion correction enable near-atomic-resolution single-particle cryo-EM
Xueming Li,Paul Mooney,Shawn Q. Zheng,Shawn Q. Zheng,Christopher R. Booth,Michael B. Braunfeld,Michael B. Braunfeld,Sander Gubbens,David A. Agard,David A. Agard,Yifan Cheng +10 more
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TLDR
This approach determined a 3.3-Å-resolution structure of an ∼700-kDa protein with D7 symmetry, the Thermoplasma acidophilum 20S proteasome, showing clear side-chain density and greatly enhances image quality and data acquisition efficiency.Abstract:
In recent work with large high-symmetry viruses, single-particle electron cryomicroscopy (cryo-EM) has achieved the determination of near-atomic-resolution structures by allowing direct fitting of atomic models into experimental density maps. However, achieving this goal with smaller particles of lower symmetry remains challenging. Using a newly developed single electron-counting detector, we confirmed that electron beam-induced motion substantially degrades resolution, and we showed that the combination of rapid readout and nearly noiseless electron counting allow image blurring to be corrected to subpixel accuracy, restoring intrinsic image information to high resolution (Thon rings visible to ∼3 A). Using this approach, we determined a 3.3-A-resolution structure of an ∼700-kDa protein with D7 symmetry, the Thermoplasma acidophilum 20S proteasome, showing clear side-chain density. Our method greatly enhances image quality and data acquisition efficiency-key bottlenecks in applying near-atomic-resolution cryo-EM to a broad range of protein samples.read more
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Single particle electron cryomicroscopy: trends, issues and future perspective.
TL;DR: In this article, a review of the state-of-the-art single particle image micrographs of macromolecular complexes is presented, showing illustrative examples of single particle images of large and small macromolescular complexes.
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DeepPicker: A deep learning approach for fully automated particle picking in cryo-EM.
Feng Wang,Huichao Gong,Gaochao liu,Meijing Li,Chuangye Yan,Tian Xia,Xueming Li,Jianyang Zeng +7 more
TL;DR: Tests on the recently-published cryo-EM data of three complexes have demonstrated that the deep learning based scheme, DeepPicker, can successfully accomplish the human-level particle picking process and identify a sufficient number of particles that are comparable to those picked manually by human experts.
Journal ArticleDOI
The structure of the yeast mitochondrial ribosome
TL;DR: The structure of the 75-component yeast mitoribosome is determined using electron cryomicroscopy and provides insights into the evolution and species-specific specialization of mitochondrial translation.
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In Situ Cryo-Electron Tomography: A Post-Reductionist Approach to Structural Biology.
TL;DR: A detailed computational workflow for in situ structural analysis is outlined and two recent studies are described to illustrate how this workflow can be adapted to examine both known and unknown cellular complexes.
Journal ArticleDOI
Molecular mechanism of APC/C activation by mitotic phosphorylation
Suyang Zhang,Leifu Chang,Claudio Alfieri,Zhiguo Zhang,Jing Yang,Sarah L. Maslen,Mark Skehel,David Barford +7 more
TL;DR: The results reveal the mechanism for the regulation of mitotic APC/C by phosphorylation and provide a rationale for the development of selective inhibitors of this state.
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Prevention of overfitting in cryo-EM structure determination
Sjors H.W. Scheres,Shaoxia Chen +1 more
TL;DR: Analysis of simulated data with realistic signal-to-noise ratios indicates that the accuracy of the orientation determination is not affected by the exclusion of high-frequency terms, nor by the use of a model that is reconstructed from only half of the particles, as expected.