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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Architecture of the polyketide synthase module: surprises from electron cryo-microscopy.
TL;DR: The unexpected module architecture revealed a striking evolutionary divergence of the polyketide synthase compared to its metazoan fatty acid synthase homolog, as well as remarkable conformational rearrangements dependent on its biochemical state during the full catalytic cycle.
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Advances in high-resolution cryo-EM of oligomeric enzymes
Janet Vonck,Deryck J. Mills +1 more
TL;DR: Cryo-EM structures can be solved de novo for large complexes that resist crystallisation or structure determination by crystallographic techniques, and many conformations in a sample can be separated computationally.
Journal ArticleDOI
Native structure of a retroviral envelope protein and its conformational change upon interaction with the target cell.
Christiane Riedel,Daven Vasishtan,C. Alistair Siebert,Cathy Whittle,Maik Joerg Lehmann,Walther Mothes,Kay Grünewald +6 more
TL;DR: Insight is provided into the changes occurring in retroviral Env during its initial interaction with the cell, employing murine leukemia virus (MLV) as model system and elucidating the changes in MLV Env upon interaction with a host cell.
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Cryo-Electron Microscopy Structure of the Macrobrachium rosenbergii Nodavirus Capsid at 7 Angstroms Resolution.
TL;DR: The structural analysis supports the assertion that MrNV may belong to a new genus of the Nodaviridae and provides the first structural view of an important pathogen affecting aquaculture industries across the world.
Journal ArticleDOI
RsgA couples the maturation state of the 30S ribosomal decoding center to activation of its GTPase pocket.
Jorge P. López-Alonso,T. Kaminishi,Takeshi Kikuchi,Yuyia Hirata,Idoia Iturrioz,Neha Dhimole,Andreas Schedlbauer,Yoichi Hase,Simon Goto,Daisuke Kurita,Akira Muto,Shu Zhou,Chieko Naoe,Deryck J. Mills,David Gil-Carton,Chie Takemoto,Hyouta Himeno,Paola Fucini,Sean R. Connell +18 more
TL;DR: The results show that RsgA destabilizes the 30S structure, including late binding r-proteins, providing a structural basis for avoiding kinetically trapped assembly intermediates and validates the architecture of the decoding center and facilitates the timely release of Rs gTPase to control the progression of 30S biogenesis.
References
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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.