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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Damage-free Vibrational Spectroscopy of Biological Materials in the Electron Microscope
Peter Rez,Toshihiro Aoki,Katia March,Dvir Gur,Ondrej L. Krivanek,Niklas Dellby,Tracy C. Lovejoy,Sharon G. Wolf,Hagai Cohen +8 more
TL;DR: The potential of aloof spectroscopy is demonstrated, which opens up the possibility of non-damaging compositional analyses of organic functional groups, including non-crystalline biological materials, at a spatial resolution of ∼10 nm, simultaneously combined with imaging in the electron microscope.
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Direct Observation of Wet Biological Samples by Graphene Liquid Cell Transmission Electron Microscopy.
Jungwon Park,Hyesung Park,Peter Ercius,Adrian F. Pegoraro,Chen Xu,Jin Woong Kim,Sang Hoon Han,David A. Weitz +7 more
TL;DR: A graphene liquid cell (GLC) is introduced using multilayer graphene sheets to reliably encapsulate and preserve biological samples in a liquid for TEM observation and is used to directly observe the structure of influenza viruses in their native buffer solution at room temperature.
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Electron cryotomography of vitrified cells with a Volta phase plate
TL;DR: After 3D reconstruction, tomograms acquired with the combination of a VPP and an energy filter showed structural features in neuronal processes with outstanding clarity, and it is expected that VPP will become a standard element of the electron cryotomography workflow.
Journal ArticleDOI
Binding of ISRIB reveals a regulatory site in the nucleotide exchange factor eIF2B
Alisa Zyryanova,Felix Weis,Alexandre Faille,Akeel Abo Alard,Ana Crespillo-Casado,Yusuke Sekine,Heather P. Harding,Felicity Allen,Leopold Parts,Christophe Fromont,Peter Fischer,Alan J. Warren,David Ron +12 more
TL;DR: Findings point to a site in eIF2B that can be exploited by ISRIB to regulate translation, and this work describes a 4.1-angstrom-resolution cryo–electron microscopy structure of human eif2B with an ISRIb molecule bound at the interface between the β and δ regulatory subunits.
Journal ArticleDOI
Structures of the human mitochondrial ribosome in native states of assembly.
Alan Brown,Sorbhi Rathore,Dari Kimanius,Shintaro Aibara,Xiao Chen Bai,Xiao Chen Bai,Joanna Rorbach,Joanna Rorbach,Alexey Amunts,Alexey Amunts,Venki Ramakrishnan +10 more
TL;DR: Comparison of the structures reveals insights into the timing of rRNA folding and protein incorporation during the final steps of ribosomal maturation and the evolutionary adaptations that are required to preserve biogenesis after the structural diversification of mitoribosomes.
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