https://www.biorxiv.org/content/biorxiv/early/2020/02/20/2020.02.19.956581.full.pdf

Structure, Function, and Antigenicity of the SARS-CoV-2 Spike Glycoprotein.

Single-particle electron cryomicroscopy (cryo-EM) is a powerful method for determining the structures of biological macromolecules. With automated microscopes, cryo-EM data can often be obtained in a few days. However, processing cryo-EM image data to reveal heterogeneity in the protein structure and to refine 3D maps to high resolution frequently becomes a severe bottleneck, requiring expert intervention, prior structural knowledge, and weeks of calculations on expensive computer clusters. Here we show that stochastic gradient descent (SGD) and branch-and-bound maximum likelihood optimization algorithms permit the major steps in cryo-EM structure determination to be performed in hours or minutes on an inexpensive desktop computer. Furthermore, SGD with Bayesian marginalization allows ab initio 3D classification, enabling automated analysis and discovery of unexpected structures without bias from a reference map. These algorithms are combined in a user-friendly computer program named cryoSPARC (http://www.cryosparc.com).

cryoSPARC: algorithms for rapid unsupervised cryo-EM structure determination

Angiotensin-converting enzyme 2 (ACE2) is the cellular receptor for severe acute respiratory syndrome-coronavirus (SARS-CoV) and the new coronavirus (SARS-CoV-2) that is causing the serious coronavirus disease 2019 (COVID-19) epidemic. Here, we present cryo-electron microscopy structures of full-length human ACE2 in the presence of the neutral amino acid transporter B0AT1 with or without the receptor binding domain (RBD) of the surface spike glycoprotein (S protein) of SARS-CoV-2, both at an overall resolution of 2.9 angstroms, with a local resolution of 3.5 angstroms at the ACE2-RBD interface. The ACE2-B0AT1 complex is assembled as a dimer of heterodimers, with the collectrin-like domain of ACE2 mediating homodimerization. The RBD is recognized by the extracellular peptidase domain of ACE2 mainly through polar residues. These findings provide important insights into the molecular basis for coronavirus recognition and infection.

Structural basis for the recognition of SARS-CoV-2 by full-length human ACE2.

Here, we describe the third major release of RELION. CPU-based vector acceleration has been added in addition to GPU support, which provides flexibility in use of resources and avoids memory limitations. Reference-free autopicking with Laplacian-of-Gaussian filtering and execution of jobs from python allows non-interactive processing during acquisition, including 2D-classification, de novo model generation and 3D-classification. Per-particle refinement of CTF parameters and correction of estimated beam tilt provides higher resolution reconstructions when particles are at different heights in the ice, and/or coma-free alignment has not been optimal. Ewald sphere curvature correction improves resolution for large particles. We illustrate these developments with publicly available data sets: together with a Bayesian approach to beam-induced motion correction it leads to resolution improvements of 0.2-0.7 A compared to previous RELION versions.

/pdf/new-tools-for-automated-high-resolution-cryo-em-structure-d3sztfqg71.pdf

New tools for automated high-resolution cryo-EM structure determination in RELION-3.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a newly emerged coronavirus that is responsible for the current pandemic of coronavirus disease 2019 (COVID-19), which has resulted in more than 3.7 million infections and 260,000 deaths as of 6 May 20201,2. Vaccine and therapeutic discovery efforts are paramount to curb the pandemic spread of this zoonotic virus. The SARS-CoV-2 spike (S) glycoprotein promotes entry into host cells and is the main target of neutralizing antibodies. Here we describe several monoclonal antibodies that target the S glycoprotein of SARS-CoV-2, which we identified from memory B cells of an individual who was infected with severe acute respiratory syndrome coronavirus (SARS-CoV) in 2003. One antibody (named S309) potently neutralizes SARS-CoV-2 and SARS-CoV pseudoviruses as well as authentic SARS-CoV-2, by engaging the receptor-binding domain of the S glycoprotein. Using cryo-electron microscopy and binding assays, we show that S309 recognizes an epitope containing a glycan that is conserved within the Sarbecovirus subgenus, without competing with receptor attachment. Antibody cocktails that include S309 in combination with other antibodies that we identified further enhanced SARS-CoV-2 neutralization, and may limit the emergence of neutralization-escape mutants. These results pave the way for using S309 and antibody cocktails containing S309 for prophylaxis in individuals at a high risk of exposure or as a post-exposure therapy to limit or treat severe disease.

/pdf/cross-neutralization-of-sars-cov-2-by-a-human-monoclonal-hgjbq7oa3x.pdf

Cross-neutralization of SARS-CoV-2 by a human monoclonal SARS-CoV antibody.

https://journals.iucr.org/m/issues/2017/06/00/kf5004/kf5004.pdf

Cryo-EM reconstruction of the chlororibosome to 3.2 Å resolution within 24 h.

Ligand-gated ion channels are critical mediators of electrochemical signal transduction across evolution. Biophysical and pharmacological development in this family relies on high-quality structural data in multiple, subtly distinct functional states. However, structural data remain limited, particularly for the unliganded or resting state. Here we report cryo-electron microscopy structures of the Gloeobacter violaceus ligand-gated ion channel (GLIC) under resting and activating conditions (neutral and low pH). Parallel models were built either manually or using recently developed density-guided molecular simulations. The moderate resolution of resting-state reconstructions, particularly in the extracellular domain, was improved under activating conditions, enabling the visualization of residues at key subunit interfaces including loops B, C, F, and M2–M3. Combined with molecular dynamics simulations, the cryo-electron microscopy structures at different pH describe a heterogeneous population of closed channels, with activating conditions condensing the closed-channel energy landscape on a pathway towards gating.

https://biorxiv.org/content/10.1101/2020.06.19.161356v1.full.pdf

Characterization of the dynamic resting state of a pentameric ligand-gated ion channel by cryo-electron microscopy and simulations

In the version of this Article originally published, the name of co-author Annemarie Perez Boerema was coded wrongly, resulting in it being incorrect when exported to citation databases. This has been corrected, though no visible changes will be apparent.

/pdf/publisher-correction-structure-of-the-chloroplast-ribosome-4a19oprbh2.pdf

Publisher Correction: Structure of the chloroplast ribosome with chl-RRF and hibernation-promoting factor.

Eukaryotic organisms rely on several mechanisms to inhibit bacterial growth and infection, which mankind has sought to mimic to their specific and more targeted use. Due to the effect of adaptation ...

The antimicrobial effect of dermcidin investigated by computational electrophysiology molecular dynamics simulations

Abstract The pyruvate dehydrogenase complex (PDC) is a central metabolic enzyme in all living cells composed majorly of E1, E2, and E3. Tight coupling of their reactions makes each component essential, so that any loss impacts oxidative metabolism pathologically. E3 retention is mediated by the E3-binding protein (E3BP), which is here resolved within the PDC core from N.crassa , resolved to 3.2Å. Fungal and mammalian E3BP are shown to be orthologs, arguing E3BP as a broadly eukaryotic gene. Fungal E3BP architectures predicted from sequence data and computational models further bridge the evolutionary distance between N.crassa and humans, and suggest discriminants for E3-specificity. This is confirmed by similarities in their respective E3-binding domains, where an interaction previously not described is also predicted. This provides evolutionary parallels for a crucial interaction human metabolism, an interaction specific to fungi that can be targeted, and an example of protein evolution following gene neofunctionalization. 

/pdf/the-structure-and-evolutionary-diversity-of-the-fungal-e3-3qoei23r.pdf

Björn O. Forsberg

Papers

Cryo-EM reconstruction of the chlororibosome to 3.2 Å resolution within 24 h.

Characterization of the dynamic resting state of a pentameric ligand-gated ion channel by cryo-electron microscopy and simulations

Publisher Correction: Structure of the chloroplast ribosome with chl-RRF and hibernation-promoting factor.

The antimicrobial effect of dermcidin investigated by computational electrophysiology molecular dynamics simulations

The structure and evolutionary diversity of the fungal E3-binding protein