A hexapeptide of the receptor-binding domain of SARS corona virus spike protein blocks viral entry into host cells via the human receptor ACE2

Severe acute respiratory syndrome coronavirus has a great role in causing respiratory illness in humans and has the most important relationship of its spike proteins with host ACE-2 receptors. After entry into the human body, the viral S protein receptor-binding domain binds to human ACE-2 receptor. Two modes explained in this paper of an ACE-2 shedding. The shedding induces the process of viral entry to host cells by binding SARS-CoV-2 proteins. The residues of arginine and lysine in the ACE-2 receptor from 652 to 659 amino acid cleavage by ADAM17 but in TMPRSS2 the residues can be seen on amino acid from 697 to 716. Corona virus genome shows some structural proteins that are responsible for the cellular entry and facilitate the attachment of a virus to the host cell. Virus recognizes the attachment site and binds with it and enter into the cell. Spike protein is split from the cleavage site along its two subunits S1 and S2 then during this process. S2 subunit release RBD (Receptor- Binding Domain) of S1 mediated to the ACE-2. The RBD of S1 consists of 200 amino acid domains. The unknown protein B6ATI which is a neutral amino acid transporter located in ileum is the basic cause for formation of ACE-2 homodimer. In this way S1 domain provides site for another S2 domain. This leads to concealing of the ACE-2 ectodomain cleavage-sites, shedding. It prevents endocytosis of the receptor blocking a major pathway in the viral entry.

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A Fungal Defensin Targets the SARS−CoV−2 Spike Receptor−Binding Domain

Journal of Fungi ◽

10.3390/jof7070553 ◽

2021 ◽

Vol 7 (7) ◽

pp. 553

Author(s):

Bin Gao ◽

Shunyi Zhu

Keyword(s):

Receptor Binding ◽

Viral Entry ◽

Single Point ◽

Binding Domain ◽

Host Cells ◽

Single Point Mutation ◽

Binding Motif ◽

Microsporum Canis ◽

Receptor Binding Domain ◽

Fungal Defensin

Coronavirus Disease 2019 (COVID−19) elicited by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS−CoV−2) is calling for novel targeted drugs. Since the viral entry into host cells depends on specific interactions between the receptor−binding domain (RBD) of the viral Spike protein and the membrane−bound monocarboxypeptidase angiotensin converting enzyme 2 (ACE2), the development of high affinity RBD binders to compete with human ACE2 represents a promising strategy for the design of therapeutics to prevent viral entry. Here, we report the discovery of such a binder and its improvement via a combination of computational and experimental approaches. The binder micasin, a known fungal defensin from the dermatophytic fungus Microsporum canis with antibacterial activity, can dock to the crevice formed by the receptor−binding motif (RBM) of RBD via an extensive shape complementarity interface (855.9 Å2 in area) with numerous hydrophobic and hydrogen−bonding interactions. Using microscale thermophoresis (MST) technique, we confirmed that micasin and its C−terminal γ−core derivative with multiple predicted interacting residues exhibited a low micromolar affinity to RBD. Expanding the interface area of micasin through a single point mutation to 970.5 Å2 accompanying an enhanced hydrogen bond network significantly improved its binding affinity by six−fold. Our work highlights the naturally occurring fungal defensins as an emerging resource that may be suitable for the development into antiviral agents for COVID−19.

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Stereotypic neutralizing VH antibodies against SARS-CoV-2 spike protein receptor binding domain in COVID-19 patients and healthy individuals

Science Translational Medicine ◽

10.1126/scitranslmed.abd6990 ◽

2021 ◽

pp. eabd6990

Author(s):

Sang Il Kim ◽

Jinsung Noh ◽

Sujeong Kim ◽

Younggeun Choi ◽

Duck Kyun Yoo ◽

...

Keyword(s):

B Cells ◽

Receptor Binding ◽

Neutralizing Antibodies ◽

De Novo ◽

Binding Domain ◽

Host Cells ◽

Spike Protein ◽

Receptor Binding Domain ◽

Healthy Individuals

Stereotypic antibody clonotypes exist in healthy individuals and may provide protective immunity against viral infections by neutralization. We observed that 13 out of 17 patients with COVID-19 had stereotypic variable heavy chain (VH) antibody clonotypes directed against the receptor-binding domain (RBD) of SARS-CoV-2 spike protein. These antibody clonotypes were comprised of immunoglobulin heavy variable (IGHV)3-53 or IGHV3-66 and immunoglobulin heavy joining (IGHJ)6 genes. These clonotypes included IgM, IgG3, IgG1, IgA1, IgG2, and IgA2 subtypes and had minimal somatic mutations, which suggested swift class switching after SARS-CoV-2 infection. The different immunoglobulin heavy variable chains were paired with diverse light chains resulting in binding to the RBD of SARS-CoV-2 spike protein. Human antibodies specific for the RBD can neutralize SARS-CoV-2 by inhibiting entry into host cells. We observed that one of these stereotypic neutralizing antibodies could inhibit viral replication in vitro using a clinical isolate of SARS-CoV-2. We also found that these VH clonotypes existed in six out of 10 healthy individuals, with IgM isotypes predominating. These findings suggest that stereotypic clonotypes can develop de novo from naïve B cells and not from memory B cells established from prior exposure to similar viruses. The expeditious and stereotypic expansion of these clonotypes may have occurred in patients infected with SARS-CoV-2 because they were already present.

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SARS-CoV-2, an evolutionary perspective of interaction with human ACE2 reveals undiscovered amino acids necessary for complex stability

10.1101/2020.03.21.001933 ◽

2020 ◽

Cited By ~ 4

Author(s):

Vinicio Armijos-Jaramillo ◽

Justin Yeager ◽

Claire Muslin ◽

Yunierkis Perez-Castillo

Keyword(s):

Receptor Binding ◽

Evolutionary Dynamics ◽

Hot Spot ◽

Critical Role ◽

Human Cells ◽

Binding Domain ◽

Host Cells ◽

Spike Protein ◽

Receptor Binding Domain ◽

Salt Bridges

AbstractThe emergence of SARS-CoV-2 has resulted in more than 200,000 infections and nearly 9,000 deaths globally so far. This novel virus is thought to have originated from an animal reservoir, and acquired the ability to infect human cells using the SARS-CoV cell receptor hACE2. In the wake of a global pandemic it is essential to improve our understanding of the evolutionary dynamics surrounding the origin and spread of a novel infectious disease. One way theory predicts selection pressures should shape viral evolution is to enhance binding with host cells. We first assessed evolutionary dynamics in select betacoronavirus spike protein genes to predict where these genomic regions are under directional or purifying selection between divergent viral lineages at various scales of relatedness. With this analysis, we determine a region inside the receptor-binding domain with putative sites under positive selection interspersed among highly conserved sites, which are implicated in structural stability of the viral spike protein and its union with human receptor hACE2. Next, to gain further insights into factors associated with coronaviruses recognition of the human host receptor, we performed modeling studies of five different coronaviruses and their potential binding to hACE2. Modeling results indicate that interfering with the salt bridges at hot spot 353 could be an effective strategy for inhibiting binding, and hence for the prevention of coronavirus infections. We also propose that a glycine residue at the receptor binding domain of the spike glycoprotein can have a critical role in permitting bat variants of the coronaviruses to infect human cells.

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A modular molecular framework for quickly estimating the binding affinity of the spike protein of SARS-CoV-2 variants for ACE2, in presence of mutations at the spike receptor binding domain.

10.1101/2021.05.26.445422 ◽

2021 ◽

Author(s):

Vincenzo Tragni ◽

Francesca Preziusi ◽

Luna Laera ◽

Angelo Onofrio ◽

Simona Todisco ◽

...

Keyword(s):

Host Cell ◽

Receptor Binding ◽

Conformational Changes ◽

Binding Domain ◽

Host Cells ◽

Spike Protein ◽

Binding Affinities ◽

Receptor Binding Domain ◽

Rapid Spread ◽

Related Proteins

The rapid spread of new SARS-CoV-2 variants needs the development of rapid tools for predicting the affinity of the mutated proteins responsible for the infection, i.e., the SARS-CoV-2 spike protein, for the human ACE2 receptor, aiming to understand if a variant can be more efficient in invading host cells. Here we show how our computational pipeline, previously used for studying SARS-CoV-2 spike receptor-binding domain (RBD)/ACE2 interactions and pre-/post-fusion conformational changes, can be used for predicting binding affinities of the human ACE2 receptor for the spike protein RBD of the characterized infectious variants of concern/interest B.1.1.7-UK (carrying the mutations N501Y, S494P, E484K at the RBD), P.1-Japan/Brazil (RBD mutations: K417N/T, E484K, N501Y), B.1.351-South Africa (RBD mutations: K417N, E484K, N501Y), B.1.427/B.1.429-California (RBD mutations: L452R), the B.1.141 variant (RBD mutations: N439K), and the recent B.1.617.1-India (RBD mutations: L452R; E484Q) and the B.1.620 (RBD mutations: S477N; E484K). Furthermore, we searched for ACE2 structurally related proteins that might be involved in interactions with the SARS-CoV-2 spike protein, in those tissues showing low ACE2 expression, revealing two new proteins, THOP1 and NLN, deserving to be investigated for their possible inclusion in the group of host-cell entry factors responsible for host-cell SARS-CoV-2 invasion and immunity response.

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Competitive SARS-CoV-2 Serology Reveals Most Antibodies Targeting the Spike Receptor-Binding Domain Compete for ACE2 Binding

mSphere ◽

10.1128/msphere.00802-20 ◽

2020 ◽

Vol 5 (5) ◽

Cited By ~ 3

Author(s):

James R. Byrnes ◽

Xin X. Zhou ◽

Irene Lui ◽

Susanna K. Elledge ◽

Jeff E. Glasgow ◽

...

Keyword(s):

Angiotensin Converting Enzyme ◽

Receptor Binding ◽

Viral Entry ◽

Binding Domain ◽

Spike Protein ◽

Receptor Binding Domain ◽

Converting Enzyme ◽

Block Interaction ◽

Angiotensin Converting Enzyme 2 ◽

Protein Receptor

ABSTRACT As severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to spread around the world, there is an urgent need for new assay formats to characterize the humoral response to infection. Here, we present an efficient, competitive serological assay that can simultaneously determine an individual’s seroreactivity against the SARS-CoV-2 Spike protein and determine the proportion of anti-Spike antibodies that block interaction with the human angiotensin-converting enzyme 2 (ACE2) required for viral entry. In this approach based on the use of enzyme-linked immunosorbent assays (ELISA), we present natively folded viral Spike protein receptor-binding domain (RBD)-containing antigens via avidin-biotin interactions. Sera are then competed with soluble ACE2-Fc, or with a higher-affinity variant thereof, to determine the proportion of ACE2 blocking anti-RBD antibodies. Assessment of sera from 144 SARS-CoV-2 patients ultimately revealed that a remarkably consistent and high proportion of antibodies in the anti-RBD pool targeted the epitope responsible for ACE2 engagement (83% ± 11%; 50% to 107% signal inhibition in our largest cohort), further underscoring the importance of tailoring vaccines to promote the development of such antibodies. IMPORTANCE With the emergence and continued spread of the SARS-CoV-2 virus, and of the associated disease, coronavirus disease 2019 (COVID-19), there is an urgent need for improved understanding of how the body mounts an immune response to the virus. Here, we developed a competitive SARS-CoV-2 serological assay that can simultaneously determine whether an individual has developed antibodies against the SARS-CoV-2 Spike protein receptor-binding domain (RBD) and measure the proportion of these antibodies that block interaction with the human angiotensin-converting enzyme 2 (ACE2) required for viral entry. Using this assay and 144 SARS-CoV-2 patient serum samples, we found that a majority of anti-RBD antibodies compete for ACE2 binding. These results not only highlight the need to design vaccines to generate such blocking antibodies but also demonstrate the utility of this assay to rapidly screen patient sera for potentially neutralizing antibodies.

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Molecular Docking and Dynamics Simulation of a Screening Library from Life Chemicals Database for Potential Protein-Protein Interactions (PPIs) Inhibitors against SARS-CoV-2 Spike Protein

Journal of Pharmaceutical Research International ◽

10.9734/jpri/2021/v33i20a31350 ◽

2021 ◽

pp. 74-84

Author(s):

Hasanain Abdulhameed Odhar ◽

Salam Waheed Ahjel ◽

Ahmed Fadhil Hashim ◽

Ali Mahmood Rayshan

Keyword(s):

Molecular Docking ◽

Receptor Binding ◽

Protein Interactions ◽

Binding Domain ◽

Host Cells ◽

Dynamics Simulation ◽

Spike Protein ◽

Receptor Binding Domain ◽

Protein Protein Interactions ◽

Molecular Docking And Dynamics

The ongoing pandemic of coronavirus 2 represents a major challenge for global public health authorities. Coronavirus disease 2019 can be fatal especially in elderly people and those with comorbidities. Currently, several vaccines against coronavirus 2 are under application in multiple countries with emergency use authorization. In the same time, many vaccine candidates are under development and assessment. It is worth noting that the design of some of these vaccines depends on the expression of receptor binding domain for viral spike protein to induce host immunity. As such, blocking the spike protein interface with antibodies, peptides or small molecular compounds can impede the ability of coronavirus 2 to invade host cells by intervention with interactions between viral spike protein and cellular angiotensin converting enzyme 2. In this virtual screening study, we have used predictive webservers, molecular docking and dynamics simulation to evaluate the ability of 3000 compounds to interact with interface residues of spike protein receptor binding domain. This library of chemicals was focused by Life Chemicals as potential protein-protein interactions inhibitor. Here, we report that hit compound 7, with IUPAC name of 3‐cyclohexyl‐N‐(4‐{[(1R,9R) ‐6‐oxo‐7,11‐ diazatricyclo [7.3.1.02,7] trideca‐2,4‐dien‐11‐yl] sulfonyl} phenyl) propenamide, may have the capacity to interact with interface of receptor binding domain for viral spike protein and thereby reduce cellular entry of the virus. However, in vitro and in vivo assessments may be required to validate these virtual findings.

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SARS-CoV-2 Spike receptor-binding domain with a G485R mutation in complex with human ACE2

10.1101/2021.03.16.434488 ◽

2021 ◽

Author(s):

Claire M. Weekley ◽

Damian F. J. Purcell ◽

Michael W. Parker

Keyword(s):

Receptor Binding ◽

Point Mutations ◽

Direct Interaction ◽

Binding Domain ◽

Spike Protein ◽

Genomic Sequencing ◽

Binding Motif ◽

Receptor Binding Domain ◽

Structural Characterisation ◽

Human Receptor

AbstractSince SARS-CoV-2 emerged in 2019, genomic sequencing has identified mutations in the viral RNA including in the receptor-binding domain of the Spike protein. Structural characterisation of the Spike carrying point mutations aids in our understanding of how these mutations impact binding of the protein to its human receptor, ACE2, and to therapeutic antibodies. The Spike G485R mutation has been observed in multiple isolates of the virus and mutation of the adjacent residue E484 to lysine is known to contribute to antigenic escape. Here, we have crystallised the SARS-CoV-2 Spike receptor-binding domain with a G485R mutation in complex with human ACE2. The crystal structure shows that while the G485 residue does not have a direct interaction with ACE2, its mutation to arginine affects the structure of the loop made by residues 480-488 in the receptor-binding motif, disrupting the interactions of neighbouring residues with ACE2 and with potential implications for antigenic escape from vaccines, antibodies and other biologics directed against SARS-CoV-2 Spike.

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New insights into nCOVID-19 binding domain and its cellular receptors

10.1101/2020.09.06.285023 ◽

2020 ◽

Author(s):

Ankush Garg ◽

Gaurav Kumar ◽

Sharmistha Sinha

Keyword(s):

Receptor Binding ◽

Viral Entry ◽

Binding Domain ◽

Spike Protein ◽

Binding Motif ◽

Receptor Binding Domain ◽

Angiotensin Converting Enzyme 2 ◽

Cell Receptors ◽

High Propensity ◽

Multiple Receptor

AbstractnCOVID-19 virus makes cellular entry using its spike protein protruding out on its surface. Angiotensin converting enzyme 2 receptor has been identified as a receptor that mediates the viral entry by binding with the receptor binding motif of spike protein. In the present study, we elucidate the significance of N-terminal domain of spike protein in spike-receptor interactions. Recent clinical reports indicate a link between nCOVID-19 infections with patient comorbidities. The underlying reason behind this relationship is not clear. Using molecular docking, we study the affinity of the nCOVID-19 spike protein with cell receptors overexpressed under disease conditions. Our results suggest that certain cell receptors such as DC/L-SIGN, DPP4, IL22R and ephrin receptors could act as potential receptors for the spike protein. The receptor binding domain of nCOVID-19 is more flexible than that of SARS-COV and has a high propensity to undergo phase separation. Higher flexibility of nCOVID-19 receptor binding domain might enable it to bind multiple receptor partners. Further experimental work on the association of these receptors with spike protein may help us to explain the severity of nCOVID-19 infection in patients with comorbidities.

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SARS-CoV-2 receptor binding domain fusion protein efficiently neutralizes virus infection

PLoS Pathogens ◽

10.1371/journal.ppat.1010175 ◽

2021 ◽

Vol 17 (12) ◽

pp. e1010175

Author(s):

Abigael Eva Chaouat ◽

Hagit Achdout ◽

Inbal Kol ◽

Orit Berhani ◽

Gil Roi ◽

...

Keyword(s):

Virus Infection ◽

Enzymatic Activity ◽

Receptor Binding ◽

Viral Entry ◽

Surface Expression ◽

Binding Domain ◽

Host Cells ◽

Receptor Binding Domain ◽

High Virus

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the COVID-19 pandemic. Currently, as dangerous mutations emerge, there is an increased demand for specific treatments for SARS-CoV-2 infected patients. The spike glycoprotein on the virus membrane binds to the angiotensin converting enzyme 2) ACE2 (receptor on host cells through its receptor binding domain (RBD) to mediate virus entry. Thus, blocking this interaction may inhibit viral entry and consequently stop infection. Here, we generated fusion proteins composed of the extracellular portions of ACE2 and RBD fused to the Fc portion of human IgG1 (ACE2-Ig and RBD-Ig, respectively). We demonstrate that ACE2-Ig is enzymatically active and that it can be recognized by the SARS-CoV-2 RBD, independently of its enzymatic activity. We further show that RBD-Ig efficiently inhibits in-vivo SARS-CoV-2 infection better than ACE2-Ig. Mechanistically, we show that anti-spike antibody generation, ACE2 enzymatic activity, and ACE2 surface expression were not affected by RBD-Ig. Finally, we show that RBD-Ig is more efficient than ACE2-Ig at neutralizing high virus titers. We thus propose that RBD-Ig physically blocks virus infection by binding to ACE2 and that RBD-Ig should be used for the treatment of SARS-CoV-2-infected patients.

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