17 Jul 2020

Stabilizing the Closed SARS-CoV-2 Spike Trimer

The trimeric spike (S) protein of SARS-CoV-2 is the primary focus of most vaccine design and development efforts. Due to intrinsic instability typical of class I fusion proteins, S tends to prematurely refold to the post-fusion conformation, compromising immunogenic properties and prefusion trimer yields. To support ongoing vaccine development efforts, P.M. Langedijk et al. report the structure-based design of soluble S trimers, with increased yields and stabilities, based on introduction of single point mutations and disulfide-bridges. They identify two regions in the S-protein critical for the protein’s stability: the heptad repeat region 1 of the S2 subunit and subunit domain 1 at the interface with S2. We combined a minimal selection of mostly inter-protomeric mutations to create a stable S-closed variant with a 6.4-fold higher expression than the parental construct while no longer containing a heterologous trimerization domain. The cryo-EM structure reveals a correctly folded, predominantly closed pre-fusion conformation. Highly stable and well producing S protein and the increased understanding of S protein structure will support vaccine development and serological diagnostics.

16 Jul 2020

Molecular architecture of the SARS-CoV-2 virus

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an enveloped virus responsible for the COVID-19 pandemic. Despite recent advances in the structural elucidation of SARS-CoV-2 proteins and the complexes of the spike (S) proteins with the cellular receptor ACE2 or neutralizing antibodies, detailed architecture of the intact virus remains to be unveiled. Here, Li et al. from Zhejiang University School of Medicine, Hangzhou, China, report the molecular assembly of the authentic SARS-CoV-2 virus using cryo-electron tomography (cryo-ET) and subtomogram averaging (STA). Native structures of the S proteins in both pre- and post-fusion conformations were determined to average resolutions of 9-11 Å. Compositions of the N-linked glycans from the native spikes were analyzed by mass spectrometry, which revealed highly similar overall processing states of the native glycans to that of the recombinant glycoprotein glycans. The in-situ architecture of the ribonucleoproteins (RNP) and its higher-order assemblies were revealed. These characterizations have revealed the architecture of the SARS-CoV-2 virus to an unprecedented resolution, and shed lights on how the virus packs its ~30 Kb long single-segmented RNA in the ~80 nm diameter lumen. Overall, the results unveiled the molecular architecture and assembly of the SARS-CoV-2 in native context.

13 Jul 2020

Door to the cell for COVID-19 opened, leading way to therapies

A very recent study by Lan et al. published in Nature (https://doi.org/10.1038/s41586-020-2180-5) determined the crystal structure of the severe acute respiratory syndrome coronavirus (SARS-CoV)-2 receptor-binding domain (RBD) bound to angiotensin-converting enzyme 2 (ACE2). The structure reveals the mechanism of SARS-CoV-2 RBD recognition by its receptor ACE2, which is highly conserved in ACE2 recognition of SARS-CoV RBD. The study provides structural information on developing small molecules targeting SARS-CoV-2 RBD/ACE2 and implies the existence of other mechanisms than receptor binding for the markedly different infection activity of the two evolutionarily close viruses.

12 Sep 2019

Aminoacyl-tRNA synthetases as therapeutic targets

Aminoacyl-tRNA synthetases (ARSs) are essential enzymes for protein synthesis with evolutionarily conserved enzymatic mechanisms. Recent genomic, proteomic and functionomic advances have unveiled unexpected disease-associated mutations and altered expression, secretion and interactions in human ARSs, revealing hidden biological functions beyond their catalytic roles in protein synthesis. These studies have also brought to light their potential as a rich and unexplored source for new therapeutic targets and agents through multiple avenues, including direct targeting of the catalytic sites, controlling disease-associated protein–protein interactions and developing novel biologics from the secreted ARS proteins or their parts. Sunghoon Kim et. al. in Nature Reviews Drug Discovery address the emerging biology and therapeutic applications of human ARSs in diseases including autoimmune and rare diseases, and cancer.

4 Sep 2019

Atomic Force Microscopy Based Tip-Enhanced Raman Spectroscopy in Biology

Tip-enhanced Raman spectroscopy (TERS), one of the burgeoning probing techniques, can provide not only the topography characterization with high resolution, but also can deliver the chemical or molecular information of a sample beyond the optical diffraction limitation. In this review, Bo Liu et. al. mainly focus on the applications of AFM-TERS in three biological systems: nucleic acids, proteins and pathogens. From the TERS characterization to the data analysis, this review demonstrates that AFM-TERS has great potential applications to visually characterizing the biomolecular structure and crucially detecting more nano-chemical information of biological systems.

4 Sep 2019

Compressive Force Spectroscopy: From Living Cells to Single Proteins

One of the most successful applications of atomic force microscopy (AFM) in biology involves monitoring the effect of force on single biological molecules, often referred to as force spectroscopy. A less recognized variation of this method, the application of compressive force, allows studies from large samples (living cells) to smaller, multi-molecular complexes (viruses) down to single protein molecules. These studies have enabled the detailed characterization of individual cell states, subtle differences between seemingly identical viral structures, as well as the quantification of rate constants of functionally important, structural transitions in single proteins. Here, Daniel Mark Czajkowsky et. al. briefly review some of the recent achievements and highlight exciting areas of its future development.

23 Aug 2019

RNA Dynamics by NMR Spectroscopy

To reveal dynamic processes and higher energy structures, new NMR methods have been developed to elucidate dynamics in RNA with atomic resolution. In this review, Katja Petzold et. al. provide an introduction to dynamics novices and an overview of methods that access most dynamic timescales, from picoseconds to hours.

22 Aug 2019

How Good Can Single-Particle Cryo-EM Become? What Remains Before It Approaches Its Physical Limits?

Impressive though the achievements of single-particle cryo–electron microscopy are today, a substantial gap still remains between what is currently accomplished and what is theoretically possible. As is reviewed by Robert M. Glaeser, twofold or more improvements are possible as regards (a) the detective quantum efficiency of cameras at high resolution, (b) converting phase modulations to intensity modulations in the image, and (c) recovering the full amount of high-resolution signal in the presence of beam-induced motion of the specimen. In addition, potential for improvement is reviewed for other topics such as optimal choice of electron energy, use of aberration correctors, and quantum metrology. With the help of such improvements, it does not seem to be too much to imagine that determining the structural basis for every aspect of catalytic control, signaling, and regulation, in any type of cell of interest, could easily be accelerated fivefold or more.

15 Aug 2019

Potential of cryo-EM for high-resolution structural analysis of gap junction channels

The review by Oshima et. al. in Current Opinions in Structural Biology outlines structural biology of gap junction channels utilizing crystallography and single-particle cryo-EM to shed light on the functional mechanisms of cell-cell communication that are essential for multicellular organisms.

15 Aug 2019

Molecular dynamics simulations of macromolecular crystals

The paper of David Case and David Cerutti in WIREs Computational Molecular Science evaluates past fusions of simulations and crystallography and looks ahead to the ways that simulations of crystal structures will enhance structural biology in the future.

14 Aug 2019

Combining Mass Spectrometry (MS) and Nuclear Magnetic Resonance (NMR) Spectroscopy for Integrative Structural Biology of Protein–RNA Complexes

In this publication Fred Allain et.al. reviews recent advances in hybrid structural approaches with a focus on combining MS analysis of cross-linked protein–RNA complexes and NMR spectroscopy.