Stapled Peptides Targeting SARS-CoV-2 Spike Protein HR1 Inhibit the Fusion of Virus to Its Cell Receptor

Coronavirus belongs to the family of Coronaviridae, comprising single-stranded, positive-sense RNA genome (+ ssRNA) of around 26 to 32 kilobases, and has been known to cause infection to a myriad of mammalian hosts, such as humans, cats, bats, civets, dogs, and camels with varied consequences in terms of death and debilitation. Strikingly, novel coronavirus (2019-nCoV), later renamed as severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), and found to be the causative agent of coronavirus disease-19 (COVID-19), shows 88% of sequence identity with bat-SL-CoVZC45 and bat-SL-CoVZXC21, 79% with SARS-CoV and 50% with MERS-CoV, respectively. Despite key amino acid residual variability, there is an incredible structural similarity between the receptor binding domain (RBD) of spike protein (S) of SARS-CoV-2 and SARS-CoV. During infection, spike protein of SARS-CoV-2 compared to SARS-CoV displays 10–20 times greater affinity for its cognate host cell receptor, angiotensin-converting enzyme 2 (ACE2), leading proteolytic cleavage of S protein by transmembrane protease serine 2 (TMPRSS2). Following cellular entry, the ORF-1a and ORF-1ab, located downstream to 5′ end of + ssRNA genome, undergo translation, thereby forming two large polyproteins, pp1a and pp1ab. These polyproteins, following protease-induced cleavage and molecular assembly, form functional viral RNA polymerase, also referred to as replicase. Thereafter, uninterrupted orchestrated replication-transcription molecular events lead to the synthesis of multiple nested sets of subgenomic mRNAs (sgRNAs), which are finally translated to several structural and accessory proteins participating in structure formation and various molecular functions of virus, respectively. These multiple structural proteins assemble and encapsulate genomic RNA (gRNA), resulting in numerous viral progenies, which eventually exit the host cell, and spread infection to rest of the body. In this review, we primarily focus on genomic organization, structural and non-structural protein components, and potential prospective molecular targets for development of therapeutic drugs, convalescent plasm therapy, and a myriad of potential vaccines to tackle SARS-CoV-2 infection.

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Conformational Flexibility and Local Frustration in the Functional States of the SARS-CoV-2 Spike B.1.1.7 and B.1.351 Variants: Mutation-Induced Allosteric Modulation Mechanism of Functional Dynamics and Protein Stability

10.1101/2021.12.22.473892 ◽

2021 ◽

Author(s):

Gennady Verkhivker

Keyword(s):

Protein Stability ◽

Cell Receptor ◽

Spike Protein ◽

Coarse Grained ◽

Virus Infectivity ◽

Functional Changes ◽

Conformational Substates ◽

Functional Relationships ◽

Functional Dynamics ◽

Open States

The experimental and computational studies of the SARS-CoV-2 spike protein variants revealed an important role of the D614G mutation that is shared across variants of concern(VOCs), linking the effect of this mutation with the enhanced virus infectivity and transmissibility. The recent structural and biophysical studies characterized the closed and open states of the B.1.1.7 (B.1.1.7) and B.1.351 (Beta) spike variants allowing for a more detailed atomistic characterization of the conformational landscapes and functional changes. In this study, we employed coarse-grained simulations of the SARS-CoV-2 spike variant trimers together with the ensemble-based mutational frustration analysis to characterize the dynamics signatures of the conformational landscapes. By combining the local frustration analysis of the conformational ensembles with collective dynamics and residue-based mutational scanning of protein stability, we determine protein stability hotspots and identify potential energetic drivers favoring the receptor-accessible open spike states for the B.1.1.7 and B.1.351 spike variants. Through mutational scanning of protein stability changes we quantify mutational adaptability of the S-G614, S-B.1.1.7 and S-B.1.351 variants in different functional forms. Using this analysis, we found a significant conformational and mutational plasticity of the open states for all studied variants. The results of this study suggest that modulation of the energetic frustration at the inter-protomer interfaces can serve as a mechanism for allosteric couplings between mutational sites, the inter-protomer hinges of functional motions and motions of the receptor-binding domain required for binding of the host cell receptor. The proposed mechanism of mutation-induced energetic frustration may result in the greater adaptability and the emergence of multiple conformational substates in the open form. This study also suggested functional relationships between mutation-induced modulation of protein dynamics, local frustration and allosteric regulation of the SARS-CoV-2 spike protein.

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CD147-spike protein is a novel route for SARS-CoV-2 infection to host cells

Signal Transduction and Targeted Therapy ◽

10.1038/s41392-020-00426-x ◽

2020 ◽

Vol 5 (1) ◽

Cited By ~ 3

Author(s):

Ke Wang ◽

Wei Chen ◽

Zheng Zhang ◽

Yongqiang Deng ◽

Jian-Qi Lian ◽

...

Keyword(s):

Virus Entry ◽

Rapid Evolution ◽

Cell Receptor ◽

Host Cells ◽

Spike Protein ◽

Dependent Manner ◽

Viral Loads ◽

Host Cell Receptor ◽

Human T Cells ◽

Effective Drugs

AbstractIn face of the everlasting battle toward COVID-19 and the rapid evolution of SARS-CoV-2, no specific and effective drugs for treating this disease have been reported until today. Angiotensin-converting enzyme 2 (ACE2), a receptor of SARS-CoV-2, mediates the virus infection by binding to spike protein. Although ACE2 is expressed in the lung, kidney, and intestine, its expressing levels are rather low, especially in the lung. Considering the great infectivity of COVID-19, we speculate that SARS-CoV-2 may depend on other routes to facilitate its infection. Here, we first discover an interaction between host cell receptor CD147 and SARS-CoV-2 spike protein. The loss of CD147 or blocking CD147 in Vero E6 and BEAS-2B cell lines by anti-CD147 antibody, Meplazumab, inhibits SARS-CoV-2 amplification. Expression of human CD147 allows virus entry into non-susceptible BHK-21 cells, which can be neutralized by CD147 extracellular fragment. Viral loads are detectable in the lungs of human CD147 (hCD147) mice infected with SARS-CoV-2, but not in those of virus-infected wild type mice. Interestingly, virions are observed in lymphocytes of lung tissue from a COVID-19 patient. Human T cells with a property of ACE2 natural deficiency can be infected with SARS-CoV-2 pseudovirus in a dose-dependent manner, which is specifically inhibited by Meplazumab. Furthermore, CD147 mediates virus entering host cells by endocytosis. Together, our study reveals a novel virus entry route, CD147-spike protein, which provides an important target for developing specific and effective drug against COVID-19.

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An ultrapotent synthetic nanobody neutralizes SARS-CoV-2 by stabilizing inactive Spike

Science ◽

10.1126/science.abe3255 ◽

2020 ◽

pp. eabe3255 ◽

Cited By ~ 7

Author(s):

Michael Schoof ◽

Bryan Faust ◽

Reuben A. Saunders ◽

Smriti Sangwan ◽

Veronica Rezelj ◽

...

Keyword(s):

Cell Receptor ◽

Affinity Maturation ◽

Host Cells ◽

Spike Protein ◽

Airway Epithelia ◽

Angiotensin Converting Enzyme 2 ◽

Inactive Conformation ◽

Binding Domains ◽

Host Cell Receptor ◽

Yeast Surface

The SARS-CoV-2 virus enters host cells via an interaction between its Spike protein and the host cell receptor angiotensin converting enzyme 2 (ACE2). By screening a yeast surface-displayed library of synthetic nanobody sequences, we developed nanobodies that disrupt the interaction between Spike and ACE2. Cryogenic electron microscopy (cryo-EM) revealed that one nanobody, Nb6, binds Spike in a fully inactive conformation with its receptor binding domains (RBDs) locked into their inaccessible down-state, incapable of binding ACE2. Affinity maturation and structure-guided design of multivalency yielded a trivalent nanobody, mNb6-tri, with femtomolar affinity for Spike and picomolar neutralization of SARS-CoV-2 infection. mNb6-tri retains function after aerosolization, lyophilization, and heat treatment, which enables aerosol-mediated delivery of this potent neutralizer directly to the airway epithelia.

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Profiling B cell immune responses to identify neutralizing antibodies from convalescent COVID-19 patients

10.21203/rs.3.rs-38173/v2 ◽

2020 ◽

Author(s):

Lisu Huang ◽

Bingqing Shen ◽

Yu Guo ◽

Shu Shen ◽

Heyu Huang ◽

...

Keyword(s):

B Cell ◽

Neutralizing Antibodies ◽

Therapeutic Option ◽

Antiviral Drug ◽

Cell Receptor ◽

High Serum ◽

Spike Protein ◽

Live Virus ◽

Angiotensin Converting Enzyme 2 ◽

Humoral Responses

Abstract The pandemic Coronavirus Disease 2019 (COVID-19) causes noticeable morbidity and mortality worldwide. In addition to vaccine and antiviral drug therapy, the use of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) neutralizing antibodies for treatment purposes is a viable alternative. In this study, we aimed to profile the humoral responses and identify neutralizing antibodies against SARS-CoV-2 using high-throughput single-cell sequencing that tailored to B cell receptor sequencing. From two convalescent patients with high serum titer against SARS-COV-2, we identified seven antibodies specifically binding to SARS-CoV-2. Among these, the most potent antibody, P4A1 was demonstrated to block the binding of spike protein to its receptor angiotensin-converting enzyme 2 (ACE2), and prevent the viral infection in neutralization assays with pseudovirus as well as live virus at nM to sub-nM range. Moreover, antibody P4A1 can also bind strongly to spike protein with N354D/D364Y, R408I, W436R, V367F or D614G mutations respectively, suggesting that the antibody alone or in combination with other antibodies that recognize different variations of SARS-CoV-2, may provide a broad spectrum therapeutic option for COVID-19 patients. Authors Lisu Huang, Bingqing Shen, Yu Guo, and Shu Shen contributed equally to this work.

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Can ChAdOx1, a replication-deficient adenoviral (Ad) vector used to deliver SARS-Cov2 spike protein as a vaccine, recombine with the homologous human Ad to generate a novel Ad+Spike virus?

10.31219/osf.io/2hj9c ◽

2020 ◽

Author(s):

Sandeep Chakraborty

Keyword(s):

Viral Vector ◽

Rna Virus ◽

Human Adenovirus ◽

Cell Receptor ◽

Spike Protein ◽

Transcriptional Unit ◽

Human Virus ◽

Dna And Rna ◽

Host Cell Receptor ◽

Weakened Version

The development of a vaccine for Covid19 is being expedited [1]. The underlying technology for the vaccines are varied: ‘nucleic acid (DNA and RNA), virus-like particle, peptide, viral vector (replicating and non- replicating), recombinant protein, live attenuated virus and inactivated virus’ [2]. Among these, ChAdOx1, a genetically modified, weakened version of a common cold virus (adenovirus) is now in human clinical trials [3]. The ChAd vector (Chimpanzee adenovirus) was introduced in 2012 Chimpanzee adenovirus Y25 [4]. A large proportion of human adults possess significant titres of neutralising antibodies to human Adv, hence the requirement for a different adenovirus. The deletion of a single transcriptional unit, E1, ensures these viruses cant replicate. Other genes like the E3 region may also be deleted. Now, in the Covid19 vaccine ChAdOx1, the spike protein gene from MERS-CoV strain Camel/Qatar/2/2014 ‘was inserted into the E1 locus of a genomic clone of ChAdOx1 using site-specific recombination’ [5].One of the theories about the genesis of SARS-Cov2 is recombination with coronaviruses from pan- golins [6]. Whether or not it happened in SARS-Cov2, there is no denying that such recombinations do happen.How do we know that the spike protein wont be inserted into a human adenovirus using recombination?Human adenovirus shares 95% homology to ChAd. The spike protein may be inserted after the E1 protein in a viable human virus. What will happen after that to the virus is anyone’s guess. Note, that there is precedence for such recombinant adenoviruses - using ‘ping-pong” zoonosis and anthroponosis’, where the genome of a promiscuous pathogen is ‘embedded with evidence of unprecedented multiple, multidirectional, stable, and reciprocal cross-species infections of hosts from three species (human, chimpanzee, and bonobo)’ [7].Another critique - co-stimulation in host cellsA spike protein from SARS-Cov2, which is supposed to bind to ACE2 and CD147 [8], has been inserted in an adenovirus. The adenovirus has its own host-cell receptor preferences [9] - what will be the consequences of co-stimulation in those cells in which both these receptors are expressed?

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Host-cell recognition through GRP78 is enhanced in the new UK variant of SARS-CoV-2, in silico

10.21203/rs.3.rs-147971/v1 ◽

2021 ◽

Author(s):

Abdo A Elfiky ◽

Ibrahim M Ibrahim

Keyword(s):

Host Cell ◽

Cell Receptor ◽

Spike Protein ◽

Cell Recognition ◽

Receptor Binding Domain ◽

Host Cell Receptor ◽

Mutant Variant ◽

New Variant ◽

The Uk ◽

Host Cell Recognition

Abstract New SARS-CoV-2 variant VUI 202012/01 started in the UK and currently spreading in Europe and Australia during the last few days. The new variant bears about nine mutations in the spike protein (Δ69-70, Δ145, N501Y, A570D, D614G, P681H, T716I, S982A, and D1118H). The N501Y lies in the receptor-binding domain (RBD) of the spike and interacts with the host-cell receptor ACE2 responsible for viral recognition and entry. We tried to simulate the system of ACE2-SARS-CoV-2 spike RBD in the wildtype and mutated isoform of the RBD (N501Y). Additionally, the GRP78 association with the ACE2-SARS-CoV-2 spike RBD is modeled at the presence of this mutant variant of the viral spike.

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Elucidating Interactions Between SARS-CoV-2 Trimeric Spike Protein and ACE2 Using Homology Modeling and Molecular Dynamics Simulations

Frontiers in Chemistry ◽

10.3389/fchem.2020.622632 ◽

2021 ◽

Vol 8 ◽

Author(s):

Sugunadevi Sakkiah ◽

Wenjing Guo ◽

Bohu Pan ◽

Zuowei Ji ◽

Gokhan Yavas ◽

...

Keyword(s):

Molecular Dynamics ◽

Homology Modeling ◽

Molecular Dynamics Simulations ◽

Host Cell ◽

Tertiary Structure ◽

Cell Receptor ◽

Host Cells ◽

Spike Protein ◽

Receptor Protein ◽

Dynamics Simulations

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19). As of October 21, 2020, more than 41.4 million confirmed cases and 1.1 million deaths have been reported. Thus, it is immensely important to develop drugs and vaccines to combat COVID-19. The spike protein present on the outer surface of the virion plays a major role in viral infection by binding to receptor proteins present on the outer membrane of host cells, triggering membrane fusion and internalization, which enables release of viral ssRNA into the host cell. Understanding the interactions between the SARS-CoV-2 trimeric spike protein and its host cell receptor protein, angiotensin converting enzyme 2 (ACE2), is important for developing drugs and vaccines to prevent and treat COVID-19. Several crystal structures of partial and mutant SARS-CoV-2 spike proteins have been reported; however, an atomistic structure of the wild-type SARS-CoV-2 trimeric spike protein complexed with ACE2 is not yet available. Therefore, in our study, homology modeling was used to build the trimeric form of the spike protein complexed with human ACE2, followed by all-atom molecular dynamics simulations to elucidate interactions at the interface between the spike protein and ACE2. Molecular Mechanics Poisson-Boltzmann Surface Area (MMPBSA) and in silico alanine scanning were employed to characterize the interacting residues at the interface. Twenty interacting residues in the spike protein were identified that are likely to be responsible for tightly binding to ACE2, of which five residues (Val445, Thr478, Gly485, Phe490, and Ser494) were not reported in the crystal structure of the truncated spike protein receptor binding domain (RBD) complexed with ACE2. These data indicate that the interactions between ACE2 and the tertiary structure of the full-length spike protein trimer are different from those between ACE2 and the truncated monomer of the spike protein RBD. These findings could facilitate the development of drugs and vaccines to prevent SARS-CoV-2 infection and combat COVID-19.

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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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Interaction of the spike protein RBD from SARS-CoV-2 with ACE2: similarity with SARS-CoV, hot-spot analysis and effect of the receptor polymorphism

10.1101/2020.03.04.976027 ◽

2020 ◽

Cited By ~ 1

Author(s):

Houcemeddine Othman ◽

Zied Bouslama ◽

Jean-Tristan Brandenburg ◽

Jorge da Rocha ◽

Yosr Hamdi ◽

...

Keyword(s):

Hot Spot ◽

Evolutionary Process ◽

Cell Receptor ◽

Host Protein ◽

Spike Protein ◽

Amino Acid Residues ◽

Binding Assays ◽

Angiotensin Converting Enzyme 2 ◽

The Stability ◽

The Impact

AbstractThe spread of COVID-19 caused by the SARS-CoV-2 outbreak has been growing since its first identification in December 2019. The publishing of the first SARS-CoV-2 genome made a valuable source of data to study the details about its phylogeny, evolution, and interaction with the host. Protein-protein binding assays have confirmed that Angiotensin-converting enzyme 2 (ACE2) is more likely to be the cell receptor through which the virus invades the host cell. In the present work, we provide an insight into the interaction of the viral spike Receptor Binding Domain (RBD) from different coronavirus isolates with host ACE2 protein. By calculating the binding energy score between RBD and ACE2, we highlighted the putative jump in the affinity from a progenitor form of SARS-CoV-2 to the current virus responsible for COVID-19 outbreak. Our result was consistent with previously reported phylogenetic analysis and corroborates the opinion that the interface segment of the spike protein RBD might be acquired by SARS-CoV-2 via a complex evolutionary process rather than a progressive accumulation of mutations. We also highlighted the relevance of Q493 and P499 amino acid residues of SARS-CoV-2 RBD for binding to human ACE2 and maintaining the stability of the interface. Moreover, we show from the structural analysis that it is unlikely for the interface residues to be the result of genetic engineering. Finally, we studied the impact of eight different variants located at the interaction surface of ACE2, on the complex formation with SARS-CoV-2 RBD. We found that none of them is likely to disrupt the interaction with the viral RBD of SARS-CoV-2.

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