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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Cryo-EM structures of the mammalian endo-lysosomal TRPML1 channel elucidate the combined regulation mechanism
TL;DR: It is proposed TRPML1 is regulated by pH, Ca2+, and phosphoinositides in a combined manner so as to accommodate the dynamic endocytosis process.
Journal ArticleDOI
Structural changes of tailless bacteriophage ΦX174 during penetration of bacterial cell walls.
Yingyuan Sun,Aaron P. Roznowski,Joshua M. Tokuda,Thomas Klose,Alex Mauney,Lois Pollack,Bentley A. Fane,Michael G. Rossmann +7 more
TL;DR: The structural response of ΦX174 is investigated and it is shown that the phage binds to LPS through one of its pentameric spikes, and that DNA ejection through preformed tubes consisting of viral H proteins is caused.
Journal ArticleDOI
Single-particle cryo-EM using alignment by classification (ABC): the structure of Lumbricus terrestris haemoglobin
Pavel Afanasyev,Pavel Afanasyev,C. Seer-Linnemayr,Raimond B. G. Ravelli,Rishi Matadeen,S. de Carlo,B. Alewijnse,B. Alewijnse,Rodrigo Villares Portugal,Navraj S. Pannu,Michael Schatz,M. van Heel,M. van Heel +12 more
TL;DR: An efficient and fast pipeline is presented for obtaining near-atomic resolution structures from large single-particle cryo-EM data sets because the approach is virtually reference-free and is therefore less prone to the perils of reference bias.
Journal ArticleDOI
IgG Antibody 3D Structures and Dynamics.
TL;DR: IPET, as a particle-by-particle methodology for 3D structural characterization, has shown advantages in studying structural variety and conformational changes of antibodies, providing direct imaging data for biomolecular engineering to improve development and clinical application of synthetic antibodies.
Journal ArticleDOI
Double-stranded RNA virus outer shell assembly by bona fide domain-swapping.
Zhaoyang Sun,Kamel El Omari,X. Sun,Serban L. Ilca,Abhay Kotecha,David I. Stuart,Minna M. Poranen,Juha T. Huiskonen,Juha T. Huiskonen +8 more
TL;DR: The structure of the φ6 double-shelled particle provides a prime exemplar of bona fide domain-swapping and is extended from the level of monomeric subunits and multimers to closed spherical shells, to hypothesize a mechanism by which closed protein shells may arise in evolution.
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