Allosteric Cross-Talk among Spike’s Receptor-Binding Domain Mutations of the SARS-CoV-2 South African Variant Triggers an Effective Hijacking of Human Cell Receptor

SummaryThe recently emerged SARS-CoV-2 South African (B. 1.351) and Brazil/Japan (P.1) variants of concern (VoCs) include a key mutation (N501Y) found in the UK variant that enhances affinity of the spike protein for its receptor, ACE2. Additional mutations are found in these variants at residues 417 and 484 that appear to promote antibody evasion. In contrast, the Californian VoCs (B.1.427/429) lack the N501Y mutation, yet exhibit antibody evasion. We engineered spike proteins to express these RBD VoC mutations either in isolation, or in different combinations, and analyzed the effects using biochemical assays and cryo-EM structural analyses. Overall, our findings suggest that the emergence of new SARS-CoV-2 variant spikes can be rationalized as the result of mutations that confer either increased ACE2 affinity, increased antibody evasion, or both, providing a framework to dissect the molecular factors that drive VoC evolution.

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SARS-CoV-2 Prion-Like Domains in Spike Proteins Enables Higher Affinity to ACE2

10.20944/preprints202003.0422.v1 ◽

2020 ◽

Author(s):

George Tetz ◽

Victor Tetz

Keyword(s):

Receptor Binding ◽

Cell Receptor ◽

Binding Domain ◽

Spike Protein ◽

Striking Difference ◽

Receptor Binding Domain ◽

Viral Receptor ◽

Functional Roles ◽

Binding Domains ◽

Spike Proteins

Currently, the world is struggling with the coronavirus disease 2019 (COVID-19) pandemic, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Prion-like domains are critical for virulence and the development of therapeutic targets; however, the prion-like domains in the SARS-CoV-2 proteome have not been analyzed. In this in silico study, using the PLAAC algorithm, we identified the presence of prion-like domains in SARS-CoV-2 spike protein. Compared with other viruses, a striking difference was observed in the distribution of prion-like domains in the spike, since SARS-CoV-2 was the only coronavirus with a prion-like domain found in the receptor-binding domain of the S1 region of the spike protein. The presence and unique distribution of prion-like domains in the SARS-CoV-2 receptor-binding domains of spike proteins is particularly interesting, since although SARS-CoV-2 and SARS-CoV S share the same host cell receptor, angiotensin-converting enzyme 2 (ACE2), SARS-CoV-2 demonstrates a 10- to 20-fold higher affinity for ACE2. Finally, we identified prion-like domains in the α1 helix of the ACE2 receptor that interacts with the viral receptor-binding domain of SARS-CoV-2. Taken together, the present findings indicate that the identified PrDs in the SARS-CoV-2 receptor-binding domain (RBD) and ACE2 region that interacts with RBD have important functional roles in viral adhesion and entry.

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Computational insights into differential interaction of mamalian ACE2 with the SARS-CoV-2 spike receptor binding domain

10.1101/2021.02.02.429327 ◽

2021 ◽

Author(s):

Cecylia S. Lupala ◽

Vikash Kumar ◽

Xiao-dong Su ◽

Chun Wu ◽

Haiguang Liu

Keyword(s):

Receptor Binding ◽

Cell Receptor ◽

Binding Domain ◽

Cell Entry ◽

Receptor Binding Domain ◽

Sequence Structure ◽

Angiotensin Converting Enzyme 2 ◽

Host Cell Receptor ◽

Comparison Results ◽

Horseshoe Bat

AbstractThe severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causing agent of the COVID-19 pandemic, has spread globally. Angiotensin-converting enzyme 2 (ACE2) has been identified as the host cell receptor that binds to receptor-binding domain (RBD) of the SARS-COV-2 spike protein and mediates cell entry. Because the ACE2 proteins are widely available in mammals, it is important to investigate the interactions between the RBD and the ACE2 of other mammals. Here we analyzed the sequences of ACE2 proteins from 16 mammals and predicted the structures of ACE2-RBD complexes. Analyses on sequence, structure, and dynamics synergistically provide valuable insights into the interactions between ACE2 and RBD. The comparison results suggest that the ACE2 of bovine, cat and panda form strong binding with RBD, while in the cases of rat, least horseshoe bat, horse, pig, mouse and civet, the ACE2 proteins interact weakly with RBD.

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XAV-19, a Swine Glyco-Humanized Polyclonal Antibody Against SARS-CoV-2 Spike Receptor-Binding Domain, Targets Multiple Epitopes and Broadly Neutralizes Variants

Frontiers in Immunology ◽

10.3389/fimmu.2021.761250 ◽

2021 ◽

Vol 12 ◽

Author(s):

Bernard Vanhove ◽

Stéphane Marot ◽

Ray T. So ◽

Benjamin Gaborit ◽

Gwénaëlle Evanno ◽

...

Keyword(s):

South African ◽

Receptor Binding ◽

Polyclonal Antibodies ◽

Neutralizing Antibody ◽

Binding Domain ◽

Spike Protein ◽

Receptor Binding Domain ◽

Live Virus ◽

Angiotensin Converting Enzyme 2 ◽

The United Kingdom

Amino acid substitutions and deletions in the Spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants can reduce the effectiveness of monoclonal antibodies (mAbs). In contrast, heterologous polyclonal antibodies raised against S protein, through the recognition of multiple target epitopes, have the potential to maintain neutralization capacities. XAV-19 is a swine glyco-humanized polyclonal neutralizing antibody raised against the receptor binding domain (RBD) of the Wuhan-Hu-1 Spike protein of SARS-CoV-2. XAV-19 target epitopes were found distributed all over the RBD and particularly cover the receptor binding motives (RBMs), in direct contact sites with the angiotensin converting enzyme-2 (ACE-2). Therefore, in Spike/ACE-2 interaction assays, XAV-19 showed potent neutralization capacities of the original Wuhan Spike and of the United Kingdom (Alpha/B.1.1.7) and South African (Beta/B.1.351) variants. These results were confirmed by cytopathogenic assays using Vero E6 and live virus variants including the Brazil (Gamma/P.1) and the Indian (Delta/B.1.617.2) variants. In a selective pressure study on Vero E6 cells conducted over 1 month, no mutation was associated with the addition of increasing doses of XAV-19. The potential to reduce viral load in lungs was confirmed in a human ACE-2 transduced mouse model. XAV-19 is currently evaluated in patients hospitalized for COVID-19-induced moderate pneumonia in phase 2a-2b (NCT04453384) where safety was already demonstrated and in an ongoing 2/3 trial (NCT04928430) to evaluate the efficacy and safety of XAV-19 in patients with moderate-to-severe COVID-19. Owing to its polyclonal nature and its glyco-humanization, XAV-19 may provide a novel safe and effective therapeutic tool to mitigate the severity of coronavirus disease 2019 (COVID-19) including the different variants of concern identified so far.

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The Molecular Mechanism of Shiga Toxin Stx2e Neutralization by a Single-domain Antibody Targeting the Cell Receptor-binding Domain

Journal of Biological Chemistry ◽

10.1074/jbc.m114.566257 ◽

2014 ◽

Vol 289 (36) ◽

pp. 25374-25381 ◽

Cited By ~ 19

Author(s):

Alvin W. H. Lo ◽

Kristof Moonens ◽

Maia De Kerpel ◽

Lea Brys ◽

Els Pardon ◽

...

Keyword(s):

Molecular Mechanism ◽

Receptor Binding ◽

Shiga Toxin ◽

Cell Receptor ◽

Binding Domain ◽

Single Domain ◽

Receptor Binding Domain ◽

Single Domain Antibody ◽

Domain Antibody ◽

Antibody Targeting

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Neutralization of European, South African, and United States SARS-CoV-2 mutants by a human antibody and antibody domains

10.1101/2021.03.22.436481 ◽

2021 ◽

Author(s):

Zehua Sun ◽

Andrew Kim ◽

Michele D Sobolewski ◽

Nathan Enick ◽

Chuan Chen ◽

...

Keyword(s):

United States ◽

Amino Acid ◽

Severe Acute Respiratory Syndrome ◽

South African ◽

Receptor Binding ◽

Binding Domain ◽

Human Antibody ◽

Elisa Assay ◽

Receptor Binding Domain ◽

Amino Acid Mutations

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) transmission with several emerging variants remain uncontrolled in many countries, indicating the pandemic remains severe. Recent studies showed reduction of neutralization against these emerging SARS-CoV-2 variants by vaccine-elicited antibodies. Among those emerging SARS-CoV-2 variants, a panel of amino acid mutations was characterized including those in the receptor-binding domain (RBD) of the SARS-CoV-2 spike (S) glycoprotein. In the present study, we evaluated our previously identified antibody and antibody domains for binding to these RBD variants with the emerging mutations, and neutralization of pseudo typed viruses carrying spike proteins with such mutations. Our results showed that one previously identified antibody domain, ab6, can bind 32 out of 35 RBD mutants tested in an ELISA assay. All three antibodies and antibody domains can neutralize pseudo typed B.1.1.7 (UK variant), but only the antibody domain ab6 can neutralize the pseudo typed virus with the triple mutation (K417N, E484K, N501Y). This domain and its improvements have potential for therapy of infections caused by SARS-CoV-2 mutants.

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Allosteric Cross-Talk Among SARS-CoV-2 Spike’s Receptor-Binding Domain Mutations Triggers an Effective Hijacking of Human Cell Receptor

10.1101/2021.04.30.441093 ◽

2021 ◽

Author(s):

Angelo Spinello ◽

Andrea Saltalamacchia ◽

Jure Borišek ◽

Alessandra Magistrato

Keyword(s):

Receptor Binding ◽

Viral Entry ◽

Cell Receptor ◽

Binding Domain ◽

Host Cells ◽

Evolutionary Strategies ◽

Receptor Binding Domain ◽

Angiotensin Converting Enzyme 2 ◽

Societal Challenge ◽

Dynamics Simulations

ABSTRACTThe rapid and relentless emergence of novel highly transmissible SARS-CoV-2 variants, possibly decreasing vaccine efficacy, currently represents a formidable medical and societal challenge. These variants frequently hold mutations on the Spike protein’s Receptor-Binding Domain (RBD), which, binding to the Angiotensin-Converting Enzyme 2 (ACE2) receptor, mediates viral entry into the host cells.Here, all-atom Molecular Dynamics simulations and Dynamical Network Theory of the wild-type and mutant RBD/ACE2 adducts disclose that while the N501Y mutation (UK variant) enhances the Spike’s binding affinity towards ACE2, the N501Y, E484K and K417N mutations (South African variant) aptly adapt to increase SARS-CoV-2 propagation via a two-pronged strategy: (i) effectively grasping ACE2 through an allosteric signaling between pivotal RBD structural elements; and (ii) impairing the binding of antibodies elicited by infected/vaccinated patients. This information, unlocking the molecular terms and evolutionary strategies underlying the increased virulence of emerging SARS-CoV-2 variants, set the basis for developing the next-generation anti-COVID-19 therapeutics.TOC GRAPHICS

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Peptide inhibitors of the interaction of the SARS-CoV-2 receptor-binding domain with the ACE2 cell receptor

Biomeditsinskaya Khimiya ◽

10.18097/pbmc20216703244 ◽

2021 ◽

Vol 67 (3) ◽

pp. 244-250

Author(s):

R.Sh. Bibilashvili ◽

M.V. Sidorova ◽

U.S. Dudkina ◽

M.E. Palkeeva ◽

A.S. Molokoedov ◽

...

Keyword(s):

Amino Acid ◽

Receptor Binding ◽

Disulfide Bonds ◽

Protein S ◽

Cell Receptor ◽

Binding Domain ◽

Amino Acid Sequences ◽

Receptor Binding Domain ◽

Amino Acid Residues ◽

Chimeric Molecules

Computer simulation has been used to identify peptides that mimic the natural target of the SARS-CoV-2 coronavirus spike (S) protein, the angiotensin converting enzyme type 2 (ACE2) cell receptor. Based on the structure of the complex of the protein S receptor-binding domain (RBD) and ACE2, the design of chimeric molecules consisting of two 22-23-mer peptides linked to each other by disulfide bonds was carried out. The chimeric molecule X1 was a disulfide dimer, in which edge cysteine residues in the precursor molecules h1 and h2 were connected by the S-S bond. In the chimeric molecule X2, the disulfide bond was located in the middle of the molecule of each of the precursor peptides. The precursors h1 and h2 modelled amino acid sequences of α1- and α2-helices of the extracellular peptidase domain of ACE2, respectively, keeping intact most of the amino acid residues involved in the interaction with RBD. The aim of the work was to evaluate the binding efficiency of chimeric molecules and their RBD-peptides (particularly in dependence of the middle and edge methods of fixing the initial peptides h1 and h2). The proposed polypeptides and chimeric molecules were synthesized by chemical methods, purified (to 95-97% purity), and characterized by HPLC and MALDI-TOF mass spectrometry. The binding of the peptides to the SARS-CoV-2 RBD was evaluated by microthermophoresis with recombinant domains corresponding in sequence to the original Chinese (GenBank ID NC_045512.2) and the British (B. 1.1.7, GISAID EPI_ISL_683466) variants. Binding to the original RBD of the Chinese variant was detected in three synthesized peptides: linear h2 and both chimeric variants. Chimeric peptides were also bound to the RBD of the British variant with micromolar constants. The antiviral activity of the proposed peptides in Vero cell culture was also evaluated.

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1HN, 13C, and 15N resonance assignments of the Clostridioides difficile receptor binding domain 2 (CDTb, residues 757–876)

Biomolecular NMR Assignments ◽

10.1007/s12104-020-09979-y ◽

2020 ◽

Author(s):

Mary E. Cook ◽

Kristen M. Varney ◽

Raquel Godoy-Ruiz ◽

David J. Weber

Keyword(s):

Amino Acid ◽

Receptor Binding ◽

Bacterial Pathogen ◽

Resonance Assignments ◽

Cell Receptor ◽

Binding Domain ◽

Binary Toxin ◽

Receptor Binding Domain ◽

Clostridioides Difficile ◽

Binary Toxins

Abstract Clostridioides difficile is a bacterial pathogen responsible for the majority of nosocomial infections in the developed world. C. difficile infection (CDI) is difficult to treat in many cases because hypervirulent strains have evolved that contain a third toxin, termed the C. difficile toxin (CDT), in addition to the two enterotoxins TcdA and TcdB. CDT is a binary toxin comprised of an enzymatic, ADP-ribosyltransferase (ART) toxin component, CDTa, and a pore-forming or delivery subunit, CDTb. In the absence of CDTa, CDTb assembles into two distinct di-heptameric states, a symmetric and an asymmetric form with both states having two surface-accessible host cell receptor-binding domains, termed RBD1 and RBD2. RBD1 has a unique amino acid sequence, when aligned to other well-studied binary toxins (i.e., anthrax), and it contains a novel Ca2+-binding site important for CDTb stability. The other receptor binding domain, RBD2, is critically important for CDT toxicity, and a domain such as this is missing altogether in other binary toxins and shows further that CDT is unique when compared to other binary toxins. In this study, the 1H, 13C, and 15N backbone and sidechain resonances of the 120 amino acid RBD2 domain of CDTb (residues 757–876) were assigned sequence-specifically and provide a framework for future NMR-based drug discovery studies directed towards targeting the most virulent strains of CDI.

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