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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MotionCor2: Anisotropic Correction of Beam-Induced Motion for Improved Cryo-Electron Microscopy
Shawn Q. Zheng,Eugene Palovcak,Jean Paul Armache,Kliment A. Verba,Yifan Cheng,David A. Agard +5 more
TL;DR: MotionCor2 software corrects for beam-induced sample motion, improving the resolution of cryo-EM reconstructions.
Journal ArticleDOI
New tools for automated high-resolution cryo-EM structure determination in RELION-3.
Jasenko Zivanov,Takanori Nakane,Björn O. Forsberg,Dari Kimanius,Wim J. H. Hagen,Erik Lindahl,Erik Lindahl,Sjors H.W. Scheres +7 more
TL;DR: CPU-based vector acceleration has been added in addition to GPU support, which provides flexibility in use of resources and avoids memory limitations in the third major release of RELION.
Journal ArticleDOI
Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix
Dorothee Liebschner,Pavel V. Afonine,Matthew L. Baker,Gábor Bunkóczi,Vincent B. Chen,Tristan I. Croll,Bradley J. Hintze,Li-Wei Hung,Swati Jain,Airlie J. McCoy,Nigel W. Moriarty,Robert D. Oeffner,Billy K. Poon,Michael G. Prisant,Randy J. Read,Jane S. Richardson,David S. Richardson,Sammito,Oleg V. Sobolev,Duncan H. Stockwell,Thomas C. Terwilliger,Alexandre Urzhumtsev,Alexandre Urzhumtsev,Lizbeth L. Videau,Carmen J. Williams,Paul D. Adams,Paul D. Adams +26 more
TL;DR: Recent developments in the Phenix software package are described in the context of macromolecular structure determination using X-rays, neutrons and electrons.
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Gctf: Real-time CTF determination and correction
TL;DR: The main target of Gctf is to maximize the cross-correlation of a simulated CTF with the logarithmic amplitude spectra of observed micrographs after background subtraction to improve CTF parameters of all particles for subsequent image processing.
Journal ArticleDOI
Structure of the TRPV1 ion channel determined by electron cryo-microscopy
TL;DR: In this article, a high-resolution electron cryo-microscopy structure of the rat transient receptor potential (TRP) channel in its closed state is presented; the overall structure of this ion channel is found to share some common features with voltage-gated ion channels, although several unique, TRP-specific features are also characterized.
References
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Journal ArticleDOI
A rad-hard CMOS active pixel sensor for electron microscopy
Marco Battaglia,Marco Battaglia,Devis Contarato,Peter Denes,Dionisio Doering,Piero Giubilato,Tae Sung Kim,Serena Mattiazzo,Velimir Radmilovic,Sarah Zalusky,Sarah Zalusky +10 more
TL;DR: In this article, a CMOS pixel test structure able to withstand doses in excess of 1.5 GHz was presented, and the point spread function measured with 300 keV electrons was (8.1 ± 1.6 ) μ m for 10 μ m pixel and (10.9 ± 2.3 ) μm for 20 μ m pixels, respectively, which agrees well with the values of 8.4 and 10.5 μ m predicted by simulation.
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GPU-enabled FREALIGN: Accelerating single particle 3D reconstruction and refinement in Fourier space on graphics processors
TL;DR: To facilitate processing of very large datasets, the computer program, FREALIGN, was modified to execute the computationally most intensive procedures on Graphics Processing Units (GPUs) and the execution speed increased between 10 and 240-fold depending on the task performed by FREalIGN.