Protein structure analysis of the interactions between SARS-CoV-2 spike protein and the human ACE2 receptor: from conformational changes to novel neutralizing antibodies

ABSTRACTThe recent severe acute respiratory syndrome, known as Corona Virus Disease 2019 (COVID-19) has spread so much rapidly and severely to induce World Health Organization (WHO) to declare state of emergency over the new coronavirus SARS-CoV-2 pandemic. While several countries have chosen the almost complete lock-down for slowing down SARS-CoV-2 spread, scientific community is called to respond to the devastating outbreak by identifying new tools for diagnosis and treatment of the dangerous COVID-19. With this aim we performed an in silico comparative modeling analysis, which allows to gain new insights about the main conformational changes occurring in the SARS-CoV-2 spike protein, at the level of the receptor binding domain (RBD), along interactions with human cells angiotensin converting enzyme 2 (ACE2) receptor, that favour human cell invasion. Furthermore, our analysis provides i) an ideal pipeline to identify already characterized antibodies that might target SARS-CoV-2 spike RBD, for preventing interactions with the human ACE2, and ii) instructions for building new possible neutralizing antibodies, according to chemical/physical space restraints and complementary determining regions (CDR) mutagenesis of the identified existing antibodies. The proposed antibodies show in silico a high affinity for SARS-CoV-2 spike RBD and can be used as reference antibodies also for building new high affinity antibodies against present and future coronavirus able to invade human cells through interactions of their spike proteins with the human ACE2. More in general, our analysis provides indications for the set-up of the right biological molecular context for investigating spike RBD-ACE2 interactions for the development of new vaccines, diagnosis kits and other treatments based on the usage or the targeting of SARS-CoV-2 spike protein.

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Protein structure analysis of the interactions between SARS-CoV-2 spike protein and the human ACE2 receptor: from conformational changes to novel neutralizing antibodies

Cellular and Molecular Life Sciences ◽

10.1007/s00018-020-03580-1 ◽

2020 ◽

Cited By ~ 7

Author(s):

Ivan Mercurio ◽

Vincenzo Tragni ◽

Francesco Busto ◽

Anna De Grassi ◽

Ciro Leonardo Pierri

Keyword(s):

Protein Structure ◽

Structure Analysis ◽

Conformational Changes ◽

Neutralizing Antibodies ◽

Spike Protein ◽

Protein Structure Analysis

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In Silico Investigation of the New UK (B.1.1.7) and South African (501Y.V2) SARS-CoV-2 Variants with a Focus at the ACE2–Spike RBD Interface

International Journal of Molecular Sciences ◽

10.3390/ijms22041695 ◽

2021 ◽

Vol 22 (4) ◽

pp. 1695

Author(s):

Bruno O. Villoutreix ◽

Vincent Calvez ◽

Anne-Geneviève Marcelin ◽

Abdel-Majid Khatib

Keyword(s):

Amino Acid ◽

South African ◽

In Silico ◽

Neutralizing Antibodies ◽

Viral Escape ◽

Spike Protein ◽

Amino Acid Substitutions ◽

The South ◽

African Strain ◽

The Uk

SARS-CoV-2 exploits angiotensin-converting enzyme 2 (ACE2) as a receptor to invade cells. It has been reported that the UK and South African strains may have higher transmission capabilities, eventually in part due to amino acid substitutions on the SARS-CoV-2 Spike protein. The pathogenicity seems modified but is still under investigation. Here we used the experimental structure of the Spike RBD domain co-crystallized with part of the ACE2 receptor, several in silico methods and numerous experimental data reported recently to analyze the possible impacts of three amino acid replacements (Spike K417N, E484K, N501Y) with regard to ACE2 binding. We found that the N501Y replacement in this region of the interface (present in both the UK and South African strains) should be favorable for the interaction with ACE2, while the K417N and E484K substitutions (South African strain) would seem neutral or even unfavorable. It is unclear if the N501Y substitution in the South African strain could counterbalance the K417N and E484K Spike replacements with regard to ACE2 binding. Our finding suggests that the UK strain should have higher affinity toward ACE2 and therefore likely increased transmissibility and possibly pathogenicity. If indeed the South African strain has a high transmission level, this could be due to the N501Y replacement and/or to substitutions in regions located outside the direct Spike–ACE2 interface but not so much to the K417N and E484K replacements. Yet, it should be noted that amino acid changes at Spike position 484 can lead to viral escape from neutralizing antibodies. Further, these amino acid substitutions do not seem to induce major structural changes in this region of the Spike protein. This structure–function study allows us to rationalize some observations made for the UK strain but raises questions for the South African strain.

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Conformational Perturbation of SARS-CoV-2 Spike Protein Using N-Acetyl Cysteine, a Molecular Scissor: A Probable Strategy to Combat COVID-19

10.26434/chemrxiv.12687923.v1 ◽

2020 ◽

Author(s):

Utsab Debnath ◽

Varun Dewaker ◽

Yenamandra S. Prabhakar ◽

Parthasarathi Bhattacharyya ◽

Amit Mandal

Keyword(s):

Disulfide Bond ◽

In Silico ◽

Ex Vivo ◽

Virus Disease ◽

Spike Protein ◽

Early 21St Century ◽

Acetyl Cysteine ◽

Structural Architecture

The infection caused by Severe Acute Respiratory Syndrome–CoronaVirus-2 (SARS-CoV-2) resulted in a pandemic across the globe with a huge death toll. The symptoms from SARS-CoV2 appear somewhat similar to the SARS-CoV-1 infection that appeared in early 21st century but the infectivity is far higher for the SARS-CoV-2. The virus attaches itself to exposed human epithelial cells through the spike protein. Recently discovered crystal structure of the complex of spike protein of SARS-CoV-2 with human angiotensin-converting enzyme 2 (ACE2) receptor indicated that the virus binds with the host cell very strongly. We hypothesized that the perturbation of the functionally active conformation of spike protein through the reduction of a solvent accessible disulfide bond (Cys391-Cys525) that provides its structural architecture, may 2 be a feasible strategy to disintegrate the spike protein from ACE2 receptor and thereby prevent the infection. Using in silico platform we showed that N-acetyl cysteine (NAC), a drug used as antioxidant and mucolytic agent, binds in the close proximity of above disulfide bond. The reduction of the disulfide bond via thiol/disulfide exchange, followed by covalent conjugation of NAC perturbed the stereo specific orientations of interacting key residues of spike protein. This resulted in threefold weakening in the binding affinity of spike protein with ACE2 receptor. This opens avenues for exploring the effect of NAC in vitro, ex vivo and in vivo and on successful observation of the similar effect as in silico, the intervention of NAC may be translated in the pharmacoprevention and treatment of Corona virus disease 2019.

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An Overview on Lambda, Epsilon, Kappa, Iota and Zeta Variants of Covid-19 and its Probability to Merge with Delta & Delta Plus, why it is a Concern

Biointerface Research in Applied Chemistry ◽

10.33263/briac125.68956914 ◽

2021 ◽

Vol 12 (5) ◽

pp. 6895-6914

Keyword(s):

World Health Organization ◽

Neutralizing Antibodies ◽

Spike Protein ◽

World Health ◽

South American ◽

Infected Person ◽

High Prevalence ◽

The World ◽

Health Organization

COVID-19 is caused by the virus SARS-CoV-2 that belongs to the Corona groups. The subgroups of the coronavirus families are α, β, γ, and δ coronavirus. On June 15, 2021, the string λ of SARS-CoV-2 was evaluated as a variant of interest via the World Health Organization. This string has a high prevalence in some parts of South American countries, but it occurred only occasionally in Brazil. This study confirms that mutations in the λ -spike protein can be destroyed the neutralizing antibodies and increase infectivity. Coronaviruses such as SARS-CoV-2 have an evolutionary superpower called “recombination” which permits the mixing of their genomes into novel combinations. Unlike regular mutation, which precedes slowly one change at a time, recombination can produce whole changes in a coronavirus genome. Although right now, δ-variant is a concern, a mixing of λ with other variants such as δ-variant is much more of a concern compared to alone variants. There is another item: the recombination can arise within the sample after it was taken from the infected person, not while it was inside their body.

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Peptide Scanning of SARS-CoV and SARS-CoV-2 Spike Protein Subunit 1 Reveals Potential Additional Receptor Binding Sites

10.1101/2021.08.16.456470 ◽

2021 ◽

Author(s):

Weilin Lin ◽

Jannatul Rafeya ◽

Vanessa Roschewitz ◽

David Smith ◽

Adrian Keller ◽

...

Keyword(s):

Receptor Binding ◽

Binding Sites ◽

Neutralizing Antibodies ◽

Solid Phase ◽

Solid Phase Peptide Synthesis ◽

Spike Protein ◽

Loss Of Function ◽

Protein Subunit ◽

Viral Attachment ◽

High Affinity

The binding of SARS-CoV and SARS-CoV-2 to the ACE2 receptor on human cells is mediated by the spike protein subunit 1 (S1) on the virus surfaces, while the receptor binding domains (RBDs) of S1 are the major determinants for the interaction with ACE2 and dominant targets of neutralizing antibodies. However, at the virus-host interface, additional biomolecular interactions, although being relatively weak in affinity and low in specificity, could also contribute to viral attachment and play important roles in gain- or loss-of-function mutations. In this work, we performed a peptide scanning of the S1 domains of SARS-CoV and SARS-CoV-2 by synthesizing 972 16-mer native and mutated peptide fragments using a high throughput in situ array synthesis technology. By probing the array using fluorescently labelled ACE2, a number of previously unknown potential receptor binding sites of S1 have been revealed. 20 peptides were synthesized using solid phase peptide synthesis, in order to validate and quantify their binding to ACE2. Four ACE2-binding peptides were selected, to investigate whether they can be assembled through a biotinylated peptide/neutravidin system to achieve high affinity to ACE2. A number of constructs exhibited high affinity to ACE2 with Kd values of pM to low nM.

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Low neutralizing antibody titers against the Mu variant of SARS-CoV-2 in BNT162b2 vaccinated individuals

10.1101/2021.11.19.21266552 ◽

2021 ◽

Author(s):

Diego Alejandro Alvarez-Diaz ◽

Ana L Munoz ◽

Pilar Tavera-Rodriguez ◽

Maria T Herrera-Sepulveda ◽

Hector A Ruiz-Moreno ◽

...

Keyword(s):

Neutralizing Antibodies ◽

Neutralizing Antibody ◽

Spike Protein ◽

World Health ◽

The World ◽

Need To Evaluate ◽

Surveillance Programs ◽

Antibody Titers ◽

Epidemiological Impact ◽

Health Organization

Background Global surveillance programs for the virus that causes COVID-19 are showing the emergence of variants with mutations in the Spike protein, including the Mu variant, recently declared as a Variant of Interest (VOI) by the World Health Organization. Because these types of variants can be more infectious or less susceptible to antiviral treatments and vaccine-induced antibodies. Objectives To evaluate the sensitivity of the Mu variant (B.1.621) to neutralizing antibodies induced by the BNT162b2 vaccine. Study design Three of the most predominant SARS-CoV-2 variants in Colombia during the epidemiological peaks of 2021 were isolated. Microneutralization assays were performed by incubating 120 TCDI50 of each SARS-CoV-2 isolate with five 2-fold serial dilutions of sera from 14 BNT162b2 vaccinated volunteers. The MN50 titer was calculated by the Reed-Muench formula Results The three isolated variants were Mu, a Variant of Interest (VOI), Gamma, a variant of concern (VOC), and B.1.111 that lacks genetic markers associated with greater virulence. At the end of August, the Mu and Gamma variants were widely distributed in Colombia. Mu was predominant (49%), followed by Gamma (25%). In contrast, B.1.111 became almost undetectable. The evaluation of neutralizing antibodies suggests that patients vaccinated with BNT162-2 generate neutralizing antibody titers against the Mu variant at significantly lower concentrations relative to B.1.111 and Gamma. Conclusions This study shows the importance of continuing with surveillance programs of emerging variants as well as the need to evaluate the neutralizing antibody response induced by other vaccines circulating in the country against Mu and other variants with high epidemiological impact.

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Recombinant subunits of SARS-CoV-2 spike protein as vaccine candidates to elicit neutralizing antibodies

10.21203/rs.3.rs-414366/v1 ◽

2021 ◽

Author(s):

Faezeh Noorabad ghahroodi ◽

Saeed Khalili ◽

Mohammad Javad Rasaee

Keyword(s):

Recombinant Proteins ◽

In Silico ◽

Neutralizing Antibodies ◽

Spike Protein ◽

Serum Samples ◽

Strong Immune Response ◽

Live Virus ◽

Vaccine Candidates ◽

Antibody Titers

Abstract The spike protein has been reported as one of the most critical targets for vaccine design strategies against the SARS-COV-2 infection. Hence, we have designed, produced, and evaluated the potential use of recombinant proteins derived from spike protein as vaccine candidates capable of neutralizing SARS-COV-2 virus. In silico tools were used to design spike-based subunit recombinant proteins (P1, P2, and P3). These proteins were checked for their ability to be identified by the anti-SARS-COV-2 antibodies by exposing them to Covid-19 serum samples. The proteins were then injected into mice and rabbits and the antibody titers were measured for 170 days. The virus neutralization test (VNT) was performed to analyze the obtained antibodies for their neutralization efficiency. The antibodies that existed in the serum of COVID-19 patients have identified the designed proteins. The anti-spike antibody titer was increased in the animals injected with recombinant proteins. The VNT results revealed that the produced antibodies could neutralize the cultured live virus. The long-lasting antibody titers (130 and 170 days for rabbit and mouse groups) indicated the elicitation of a strong immune response by the recombinant proteins. Subunit vaccines could also be considered as robust tools for effective vaccination against COVID-19. Using a combination of in silico, in vitro, and in vivo experiments, it was shown that the injection of spike-based recombinant proteins could stimulate long-lasting and neutralizing antibody responses. Further evaluation of the recombinant proteins examined in our studies in higher primates and human would elucidate their true potentials.

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SARS CoV-2 Spike Protein in silico Interaction With ACE2 Receptors From Wild and Domestic Species

Frontiers in Genetics ◽

10.3389/fgene.2021.571707 ◽

2021 ◽

Vol 12 ◽

Author(s):

Santiago Rendon-Marin ◽

Marlen Martinez-Gutierrez ◽

Gary R. Whittaker ◽

Javier A. Jaimes ◽

Julian Ruiz-Saenz

Keyword(s):

In Silico ◽

Spike Protein ◽

World Health ◽

Health Concern ◽

Receptor Interaction ◽

Domestic Cats ◽

Intermediate Hosts ◽

Angiotensin Converting Enzyme 2 ◽

Domestic Species ◽

Health Organization

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been declared a pandemic by the World Health Organization (WHO), and since its first report, it has become a major public health concern. SARS-CoV-2 is closely related to SARS-CoV and SARS-related bat coronaviruses, and it has been described to use angiotensin-converting enzyme 2 (ACE2) as a receptor. Natural SARS-CoV-2 infection in domestic and wildlife animals, measured by RT-qPCR, has been confirmed in different countries, especially from the Felidae family.In silicoanalysis of the interaction between the SARS-CoV-2 spike protein and the cellular receptor ACE2 in various animal species has suggested that wild felids and domestic cats could be susceptible to SARS-CoV-2 based on this interaction. Here, we performed a protein-protein molecular docking analysis of SARS-CoV-2 spike protein with the ACE2 receptor from different animals to elucidate the potential of those species as intermediate hosts or susceptible animals for SARS-CoV-2 infection. Compared to human ACE2, we found that ACE2 receptors from domestic cats and tigers could efficiently interact with RBD of SARS CoV-2 Spike protein. However, dog, ferret, and hamster ACE2 receptor interaction with SARS-CoV-2 S protein RBD was not predicted as favorable, demonstrating a potential differentiated susceptibility in the evaluated species.

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Flavonoids and Nucleotide Analogs Show High Affinity for Viral Proteins of SARS-CoV-2 by in silico Analysis: New Candidates for the Treatment of COVID-19.

10.21203/rs.3.rs-67272/v1 ◽

2020 ◽

Author(s):

Luis Adrián De Jesús-González ◽

Juan Fidel Osuna-Ramos ◽

José Manuel Reyes-Ruiz ◽

Carlos Noe Farfan-Morales ◽

Selvin Noé Palacios-Rápalo ◽

...

Keyword(s):

Rna Polymerase ◽

Inhibitory Activity ◽

In Silico ◽

Antiviral Drug ◽

World Health ◽

Nucleotide Analogs ◽

High Affinity ◽

Uridine Nucleotide

Abstract The recent epidemic of COVID-19 caused by SARS-CoV-2 was declared by the World Health Organization as a public health emergency of international concern. The absence of an approved vaccine or a specific antiviral drug has made bioinformatic tools crucial for the identification of potential therapeutic targets and drugs for its control. As in other RNA viruses, the protease 3C-like and the RNA-polymerase are two of the SARS-CoV-2 targets to test drugs that can be analyzed in silico. In the present study, compounds derived from plants, fungi, and nucleoside 5'-triphosphate or uridine nucleotide analogs, with anti-DENV activity in vitro or in vivo, were analyzed by molecular docking as potential anti-SARS-CoV-2 drugs. Anthraquinone, with a DENV NS3 protease inhibitory activity; Balapiravir, Fisetin, Hyperoside, and Sofosbuvir, with a DENV NS5 RNA-polymerase inhibitory activity; and Quercetin, with both anti-NS3-NS5 activities, were tested against 3C-like protease and RNA-polymerase of SARS-CoV-2. All these drugs demonstrated a high affinity for the corresponding SARS-CoV-2 proteins, representing excellent candidates for the treatment of COVID-19. Therefore, in vitro or in vivo studies should be carried out using these compounds on models for SARS-CoV-2 infection.

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Targeting Virus-Host Interaction: An in Silico Approach to Develop Promising Inhibitors Against COVID-19

10.26434/chemrxiv.12198369 ◽

2020 ◽

Author(s):

Jitendra Subhash Rane ◽

Aroni Chatterjee ◽

Rajni Khan ◽

Abhijeet Kumar ◽

Shashikant Ray

Keyword(s):

Angiotensin Converting Enzyme ◽

In Silico ◽

Virus Disease ◽

Spike Protein ◽

Converting Enzyme ◽

Host Cell Membrane ◽

Angiotensin Converting Enzyme 2 ◽

Host Interaction

The entire human population all over the globe is currently facing appalling conditions due to the spread of infection from COVID-19 (corona virus disease-2019). In the last few months enormous amount of studies have been continuously trying to target several potential drug sites to identify a novel therapeutic target. Spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is also being targeted by several scientific groups as a novel drug target. The spike glycoprotein protein is present on the surface of the virion and binds to the human angiotensin-converting enzyme-2 (hACE2) membrane receptor thereby promoting its fusion to the host cell membrane. The binding and internalization of the virus is a crucial step in the process of infection and hence any molecule that can inhibit this, certainly holds a significant therapeutic value. We have identified AP-NP (2-(2-amino-5-(naphthalen-2- yl)pyrimidin-4-yl)phenol) and AP-4-Me-Ph (2-(2-amino-5-(p-tolyl)pyrimidin-4-yl)phenol) from a group of diaryl pyrimidine derivatives which appear to bind at the interface of hACE2-SARS-CoV-2S complex (human angiotensin converting enzyme 2 and spike glycoprotein complex) with a low binding energy (<-8 Kcal/mol). In this in-silico study we also found that AP-NP interacts with S1 domain of C-terminal part of SARS-CoV-2S however AP-4-Me-Ph was found to interact with S2 domain of SARS-CoV-2S. The result suggested that AP-NP and AP-4-Me-Ph have potential to inhibit the interaction between spike protein and hACE2 receptor also AP-4-Me-Ph might be prevent internalization of the virion within the host. Further in vitro and in vivo study will strengthen these drug candidates against the COVID-19.

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