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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EMRinger: side chain-directed model and map validation for 3D cryo-electron microscopy
Benjamin A Barad,Nathaniel Echols,Ray Yu-Ruei Wang,Yifan Cheng,Frank DiMaio,Paul D. Adams,Paul D. Adams,James S. Fraser +7 more
TL;DR: EMRinger, a tool that assesses the precise fitting of an atomic model into the map during refinement and shows how radiation damage alters scattering from negatively charged amino acids is reported.
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How cryo-EM is revolutionizing structural biology
TL;DR: The recent advances in electron detection and image processing are reviewed and the exciting new opportunities that they offer to structural biology research are illustrated.
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Measuring the optimal exposure for single particle cryo-EM using a 2.6 Å reconstruction of rotavirus VP6
Tim Grant,Nikolaus Grigorieff +1 more
TL;DR: A method of using optimal exposure values to filter movie frames, yielding images with improved contrast that lead to higher resolution reconstructions that should benefit cryo-EM work on all types of samples, especially those of relatively low-molecular mass.
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Cryo-EM Structure of a Fully Glycosylated Soluble Cleaved HIV-1 Envelope Trimer
Dmitry Lyumkis,Jean-Philippe Julien,Natalia de Val,Albert Cupo,Clinton S. Potter,Per Johan Klasse,Dennis R. Burton,Rogier W. Sanders,Rogier W. Sanders,John P. Moore,Bridget Carragher,Ian A. Wilson,Andrew B. Ward,Andrew B. Ward +13 more
TL;DR: A cryo–electron microscopy reconstruction and structural model of a cleaved, soluble Env trimer in complex with a CD4 binding site bnAb, PGV04 is presented, which reveals the spatial arrangement of Env components, including the V1/V2, V3, HR1, and HR2 domains, as well as shielding glycans.
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Cryo-EM structures of MERS-CoV and SARS-CoV spike glycoproteins reveal the dynamic receptor binding domains
Yuan Yuan,Duanfang Cao,Yanfang Zhang,Jun Ma,Jianxun Qi,Qihui Wang,Guangwen Lu,Ying Wu,Jinghua Yan,Yi Shi,Xinzheng Zhang,George F. Gao +11 more
TL;DR: High-resolution structures of the trimeric MERS- coV and SARS-CoV S proteins in its pre-fusion conformation are presented by single particle cryo-electron microscopy, demonstrating an inherently flexible RBD readily recognized by the receptor.
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