Human Receptor Angiotensin-converting Enzyme Research Articles

Elucidation of N-/O-glycosylation and site-specific mapping of sialic acid linkage isomers of SARS-CoV-2 human receptor angiotensin-converting enzyme 2.

Human angiotensin-converting enzyme 2 (hACE2) is the primary receptor for cellular entry of SARS-CoV-2 into human host cells. hACE2 is heavily glycosylated and glycans on the receptor may play a role in viral binding. Thus, comprehensive characterization of hACE2 glycosylation could aid our understanding of interactions between the receptor and SARS-CoV-2 spike (S) protein, as well as provide a basis for the development of therapeutic drugs targeting this crucial interaction. Herein, 138 N-glycan compositions were identified, most of which are complex-type N-glycans, from seven N-glycosites of hACE2. Among them, 67% contain at least one sialic acid residue. At the level of glycopeptides, the overall quantification of sialylated glycan isomers observed on the sites N322 and N546 have a higher degree of NeuAc (α2-3)Gal (over 80.3%) than that of other N-glycosites (35.6-71.0%). In terms of O-glycans, 69 glycan compositions from 12 O-glycosites were identified, and especially, the C-terminus of hACE2 is heavily O-glycosylated. The terminal sialic acid linkage type of H1N1S1 and H1N1S2 are covered highly with α2,3-sialic acid. These findings could aid the investigation of the interaction between SARS-CoV-2 and human host cells.

Unveiling Acacia farnesiana phytochemicals as Angiotensin Converting Enzyme (ACE) inhibitors via in-silico drug design, molecular docking, and bioavailability predictions: An illustration against SARS-CoV-2 spike protein

Angiotensin-converting enzyme 2 (ACE2) is a transmembrane protein that functions as a receptor for coronavirus spike protein. When spike protein fragments as the ligand binds with ACE2 protein, this ACE2 protein functions as a virus receptor, participating in the biological process known as the viral particle entry in the host cell. Hence, an in-silico study was carried out since it is faster and less expensive than trial and error methods based on experimental investigations. To study the effect of Acacia farnesiana phytochemicals on spike protein, molecular docking analyses were carried out. In this study, twelve phytochemicals from Acacia farnesiana have been selected as small molecules based on their ACEI and anti- inflammatory nature to evaluate molecular interaction between spike protein of SARS-CoV2 with ACE2 of the human complex molecule. Gallic acid, methyl gallate, kaempferol, Rhamnocitrin, naringenin, apigenin, ellagic acid, ferulic acid, myricetin, Diosmetin, Caffeic acid, and Quercetin were chosen as competent natural compounds from Acacia farnesiana as potent small molecules against COVID-19 and further ADME analysis were carried out. The result indicated that due to the presence of ACEIs and anti-inflammatory phytochemicals in Acacia farnesiana, the bound structure of ACE2 and spike protein becomes unstable. Therefore, these natural compounds can show antiviral activity by destabilizing spike protein binding with the human host ACE2 receptor.

In silico study of SARS-CoV-2 spike protein RBD and human ACE-2 affinity dynamics across variants and Omicron subvariants.

The coronavirus disease 2019virus outbreak continues worldwide, with many variants emerging, some of which are considered variants of concern (VOCs). The WHO designated Omicron as a VOC and assigned it under variant B.1.1.529. Here, we used computational studies to examine the VOCs, including Omicron subvariants,and one variant of interest. Here we found that the binding affinity of human receptor angiotensin-converting enzyme 2 (hACE2) and receptor-binding domain(RBDs) increased in the order of wild type (Wuhan-strain) < Beta < Alpha < OmicronBA.5 < Gamma < Delta < Omicron BA.2.75 < BA.1 < BA.3 < BA.2. Interactions between docked complexes revealed that the RBD residue positions like 452, 478, 493, 498, 501, and 505 are crucial in creating strong interactions with hACE2. Omicron BA.2 shows the highest binding capacity to the hACE2 receptor among all the mutant complexes. The BA.5's L452R, F486V, and T478K mutation significantly impact the interaction network in the BA.5 RBD-hACE2 interface. Here for the first time, we report the His505, an active residue on the RBD forming a salt bridge in the BA.2, leading to increased mutation stability. When the active RBD residues are mutated, binding affinity and intermolecular interactions increase across all mutant complexes. By examining the differences in different variants, this study may provide a solid foundation for structure-based drug design for newly emerging variants.

Dendrimer-Peptide Conjugates for Effective Blockade of the Interactions between SARS-CoV-2 Spike Protein and Human ACE2 Receptor.

The coronavirus disease 2019 (COVID-19) pandemic has threatened the stability of global healthcare, which is becoming an endemic issue. Despite the development of various treatment strategies to fight COVID-19, the currently available treatment options have shown varied efficacy. Herein, we have developed an avidity-based SARS-CoV-2 antagonist using dendrimer-peptide conjugates (DPCs) for effective COVID-19 treatment. Two different peptide fragments obtained from angiotensin-converting enzyme 2 (ACE2) were integrated into a single sequence, followed by the conjugation to poly(amidoamine) (PAMAM) dendrimers. We hypothesized that the strong multivalent binding avidity endowed by dendrimers would help peptides effectively block the interaction between SARS-CoV-2 and ACE2, and this antagonist effect would be dependent upon the generation (size) of the dendrimers. To assess this, binding kinetics of the DPCs prepared from generation 4 (G4) and G7 PAMAM dendrimers to spike protein of SARS-CoV-2 were quantitatively measured using surface plasmon resonance. The larger dendrimer-based DPCs exhibited significantly enhanced binding strength by 3 orders of magnitude compared to the free peptides, whereas the smaller one showed a 12.8-fold increase only. An in vitro assay using SARS-CoV-2-mimicking microbeads also showed the improved SARS-CoV-2 blockade efficiency of the G7-peptide conjugates compared to G4. In addition, the interaction between the DPCs and SARS-CoV-2 was analyzed using molecular dynamics (MD) simulation, providing an insight into how the dendrimer-mediated multivalent binding effect can enhance the SARS-CoV-2 blockade. Our findings demonstrate that the DPCs having strong binding to SARS-CoV-2 effectively block the interaction between ACE2 and SARS-CoV-2, providing a potential as a high-affinity drug delivery system to direct anti-COVID payloads to the virus.

Potential antiviral peptides targeting the SARS-CoV-2 spike protein

BackgroundThe coronavirus disease caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection became an international pandemic and created a public health crisis. The binding of the viral Spike glycoprotein to the human cell receptor angiotensin-converting enzyme 2 (ACE2) initiates viral infection. The development of efficient treatments to combat coronavirus disease is considered essential.MethodsAn in silico approach was employed to design amino acid peptide inhibitor against the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein. The designed inhibitor (SARS-CoV-2 PEP 49) consists of amino acids with the α1 helix and the β4 - β5 sheets of ACE2. The PEP-FOLD3 web tool was used to create the 3D structures of the peptide amino acids. Analyzing the interaction between ACE2 and the RBD of the Spike protein for three protein data bank entries (6M0J, 7C8D, and 7A95) indicated that the interacting amino acids were contained inside two regions of ACE2: the α1 helical protease domain (PD) and the β4 - β5 sheets.ResultsMolecular docking analysis of the designed inhibitor demonstrated that SARS-CoV-2 PEP 49 attaches directly to the ACE2 binding site of the Spike protein with a binding affinity greater than the ACE2, implying that the SARS-CoV-2 PEP 49 model may be useful as a potential RBD binding blocker.

Potential use of the S-protein-Angiotensin converting enzyme 2 binding pathway in the treatment of coronavirus disease 2019.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the pathogen that causes coronavirus disease 2019 (COVID-19), infects humans through a strong interaction between the viral spike protein (S-protein) and angiotensin converting enzyme 2 (ACE2) receptors on the cell surface. The infection of host lung cells by SARS-CoV-2 leads to clinical symptoms in patients. However, ACE2 expression is not restricted to the lungs; altered receptors have been found in the nasal and oral mucosa, vessel, brain, pancreas, gastrointestinal tract, kidney, and heart. The future of COVID-19 is uncertain, however, new viral variants are likely to emerge. The SARS-CoV-2 Omicron variant has a total of 50 gene mutations compared with the original virus; 15 of which occur in the receptor binding domain (RBD). The RBD of the viral S-protein binds to the human ACE2 receptor for viral entry. Mutations of the ACE2-RBD interface enhance tight binding by increasing hydrogen bond interactions and expanding the accessible surface area. Extracorporeal membrane oxygenation, hyperbaric oxygen, and aggressive dialysis for the treatment of COVID-19 have shown various degrees of clinical success. The use of decoy receptors based on the ACE2 receptor as a broadly potent neutralizer of SARS-CoV-2 variants has potential as a therapeutic mechanism. Drugs such as 3E8 could block binding of the S1-subunit to ACE2 and restrict the infection of ACE2-expressing cells by a variety of coronaviruses. Here, we discuss the development of ACE2-targeted strategies for the treatment and prevention of COVID-19.

Drug Repurposing Against Angiotensin-Converting Enzyme-Related Carboxypeptidase (ACE2) Through Computational Approach.

Ongoing novel coronavirus (COVID-19) with high mortality is an infectious disease in the world which epidemic in 2019 with human-human transmission. According to the literature, S-protein is one of the main proteins of COVID-19 that bind to the human cell receptor angiotensin-converting enzyme 2 (ACE2). In this study, it was attempted to identify the main effective drugs approved that may be repurposed to the binding site of ACE2. High throughput virtual screening based on the docking study was performed to know which one of the small-molecules had a potential interaction with ACE2 structure. Forasmuch as investigating and identifying the best ACE2 inhibitors among more than 3,500 small-molecules is time-consuming, supercomputer was utilized to apply docking-based virtual screening. Outputs of the proposed computational model revealed that vincristine, vinbelastin and bisoctrizole can significantly bind to ACE2 and may interface with its normal activity.

Improvement of native structure-based peptides as efficient inhibitors of protein-protein interactions of SARS-CoV-2 spike protein and human ACE2.

New pathogens responsible for novel human disease outbreaks in the last two decades are mainly the respiratory system viruses. Not different was the last pandemic episode, caused by infection of a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). One of the extensively explored targets, in the recent scientific literature, as a possible way for rapid development of COVID-19 specific drug(s) is the interaction between the receptor-binding domain of the virus’ spike (S) glycoprotein and human receptor angiotensin-converting enzyme 2 (hACE2). This protein-protein recognition process is involved in the early stages of the SARS-CoV-2 life cycle leading to the host cell membrane penetration. Thus, disrupting this interaction may block or significantly reduce the infection caused by the novel pathogen. Previously we have designed (by in silico structure-based analysis) three very short peptides having sequences inspirited by hACE2 native fragments, which effectively bind to the SARS-CoV-2 S protein and block its interaction with the human receptor. In continuation of the above mentioned studies, here we presented an application of molecular modeling approach resulting in improved binding affinity of the previously proposed ligand and its enhanced ability to inhibit meaningful host-virus protein-protein interaction. The new optimized hexapeptide binds to the virus protein with affinity one magnitude higher than the initial ligand and, as a very short peptide, has also great potential for further drug development. The peptide-based strategy is rapid and cost-effective for developing and optimizing efficient protein-protein interactions disruptors and may be successfully applied to discover antiviral candidates against other future emerging human viral infections.

Integrating in silico and in vivo approach for investigating the role of polyherbal oil in prevention and treatment of COVID-19 infection

Currently, there are no FDA approved antiviral drugs available to treat COVID-19 patients. Also, due to emergence of new SARS-CoV-2 variants, the protective efficacy of vaccines could be reduced, hence it is urgent to have alternative treatments for combating the SARS-CoV-2 infection. Since, there is a long-standing history of herbal medicine in the treatment of respiratory diseases. In the present study, we investigated two polyherbal oil blend viz. Sudarshan AV and Elixir AV (SAV and EAV) in inhibiting SARS-COV-2. From GC-MS analysis of polyherbal oils (SAV and EAV) a total of 11 active compounds were selected, on the basis of their abundance and activity. Further, from the molecular docking studies, we found an inhibitory effect of these compounds on viral envelope and membrane, spike proteins whilst an agonistic effect with human host receptor angiotensin-converting enzyme 2 (ACE2) implicating the crucial role of the individual compound in resistance of SARS-CoV-2.Since, the in-silico results suggest that polyherbal oil (SAV and EAV) contributes in preventing the entry of SARS-CoV-2 into the human body, we further investigated the efficacy of polyherbal formulated essential oil (FEO; SAV & EAV) in prevention and treatment of COVID-19 in hamster model. The male golden Syrian hamsters (n = 23) were divided into 5 groups i.e., Group 1: Control (n = 3); Group 2: Infected (n = 5); Group 3: Infected + Remdesivir (n = 5); Group 4: Infected + FEO (n = 5) and Group 5: Prophylactic FEO + Infected (n = 5). In both treatment and prophylactic groups, the FEO's significantly reduced the lung injury investigated histo-pathologically and viral load expression measured by real time PCR in comparison to infected hamsters. Furthermore, cytokines expression analysis clearly highlighted the efficacy of FEO's due to its anti-inflammatory activity and overall protection in treatment groups. In conclusion, the FEO (SAV & EAV) seem to be potent in both prevention and treatment of COVID-19 and related lung injury.

In vitro Screening of Herbal Medicinal Products for Their Supportive Curing Potential in the Context of SARS-CoV-2.

COVID-19 herbal medicinal products may have the potential for symptom relief in nonsevere or moderate disease cases. In this in vitro study we screened the five herbal medicinal products Sinupret extract (SINx), Bronchipret thyme-ivy (BRO-TE), Bronchipret thyme-primula (BRO TP), Imupret (IMU), and Tonsipret (TOP) with regard to their potential to (i) interfere with the binding of the human angiotensin-converting enzyme 2 (ACE2) receptor with the SARS-CoV-2 spike S1 protein, (ii) modulate the release of the human defensin HBD1 and cathelicidin LL-37 from human A549 lung cells upon spike S1 protein stimulation, and (iii) modulate the release of IFN-γ from activated human peripheral blood mononuclear cells (PBMC). The effect of the extracts on the interaction of spike S1 protein and the human ACE2 receptor was measured by ELISA. The effects on the intracellular IFN-γ expression in stimulated human PBMC were measured by flow cytometry. Regulation of HBD1 and LL-37 expression and secretion was assessed in 25 d long-term cultured human lung A549 epithelial cells by RT-PCR and ELISA. IMU and BRO-TE concentration-dependently inhibited the interaction between spike S1 protein and the ACE2 receptor. SINx, TOP, and BRO-TE significantly upregulated the intracellular expression of anti-viral IFN-γ from stimulated PBMC. Cotreatment of A549 cells with IMU or BRO TP together with SARS-CoV-2 spike protein significantly upregulated mRNA expression (IMU) and release of HBD1 (IMU and BRO TP) and LL-37 (BRO TP). The in vitro screening results provide first evidence for an immune-activating potential of some of the tested herbal medicinal extracts in the context of SARS-CoV-2. Whether these could be supportive in symptom relief or curing from SARS-CoV-2 infection needs deeper understanding of the observations.

Electrostatic Features for the Receptor Binding Domain of SARS-COV-2 Wildtype and Its Variants. Compass to the Severity of the Future Variants with the Charge-Rule.

Electrostatic intermolecular interactions are important in many aspects of biology. We have studied the main electrostatic features involved in the interaction of the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein with the human receptor Angiotensin-converting enzyme 2 (ACE2). As the principal computational tool, we have used the FORTE approach, capable to model proton fluctuations and computing free energies for a very large number of protein-protein systems under different physical-chemical conditions, here focusing on the RBD-ACE2 interactions. Both the wild-type and all critical variants are included in this study. From our large ensemble of extensive simulations, we obtain, as a function of pH, the binding affinities, charges of the proteins, their charge regulation capacities, and their dipole moments. In addition, we have calculated the pKas for all ionizable residues and mapped the electrostatic coupling between them. We are able to present a simple predictor for the RBD-ACE2 binding based on the data obtained for Alpha, Beta, Gamma, Delta, and Omicron variants, as a linear correlation between the total charge of the RBD and the corresponding binding affinity. This "RBD charge rule" should work as a quick test of the degree of severity of the coming SARS-CoV-2 variants in the future.

SARS-CoV-2 spike N-terminal domain modulates TMPRSS2-dependent viral entry and fusogenicity.

SummaryThe severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike N-terminal domain (NTD) remains poorly characterized despite enrichment of mutations in this region across variants of concern (VOCs). Here, we examine the contribution of the NTD to infection and cell-cell fusion by constructing chimeric spikes bearing B.1.617 lineage (Delta and Kappa variants) NTDs and generating spike pseudotyped lentivirus. We find that the Delta NTD on a Kappa or wild-type (WT) background increases S1/S2 cleavage efficiency and virus entry, specifically in lung cells and airway organoids, through use of TMPRSS2. Delta exhibits increased cell-cell fusogenicity that could be conferred to WT and Kappa spikes by Delta NTD transfer. However, chimeras of Omicron BA.1 and BA.2 spikes with a Delta NTD do not show more efficient TMPRSS2 use or fusogenicity. We conclude that the NTD allosterically modulates S1/S2 cleavage and spike-mediated functions in a spike context-dependent manner, and allosteric interactions may be lost when combining regions from more distantly related VOCs.

Predictive Models of within- and between-Species SARS-CoV-2 Transmissibility.

Viruses from the Coronaviridae family have been reported to infect a large range of hosts, including humans. The latest human-infecting coronavirus, SARS-CoV-2, turned into a pandemic and subtypes with different transmissibility have appeared since then. The SARS-CoV-2 Spike (S) protein interacts with the angiotensin-converting enzyme 2 (ACE2) host receptor, and thus, in silico models, based on the structural features of the SARS-CoV-2 S protein–ACE2 receptor complex, as well as ACE2 amino acid patterns, may be used to predict the within- and between-species transmissibility of SARS-CoV-2 subtypes. Here, it is shown that at the beginning of the pandemic, the SARS-CoV-2 S protein was, as expected for a virus that just jumped the species barrier, ill-adapted to the human ACE2 receptor, and that the replacement of one SARS-CoV-2 variant by another is partially due to a better fitting of the S protein–human ACE2 complex. Moreover, it is shown that mutations that are predicted to lead to a better fit have increased in the population due to positive selection. It is also shown that the number of ACE2-interfacing residues is positively correlated with the transmissibility rate of SARS-CoV-2 variants. Finally, it is shown that the number of species that are susceptible to infection by SARS-CoV-2, and that could be a reservoir for this virus, is likely higher than previously thought.

Epigallocatechin gallate (EGCG) attenuates severe acute respiratory coronavirus disease 2 (SARS-CoV-2) infection by blocking the interaction of SARS-CoV-2 spike protein receptor-binding domain to human angiotensin-converting enzyme 2.

The outbreak of the coronavirus disease 2019 caused by the severe acute respiratory syndrome coronavirus 2 triggered a global pandemic where control is needed through therapeutic and preventive interventions. This study aims to identify natural compounds that could affect the fusion between the viral membrane (receptor-binding domain of the severe acute respiratory syndrome coronavirus 2 spike protein) and the human cell receptor angiotensin-converting enzyme 2. Accordingly, we performed the enzyme-linked immunosorbent assay-based screening of 10 phytochemicals that already showed numerous positive effects on human health in several epidemiological studies and clinical trials. Among these phytochemicals, epigallocatechin gallate, a polyphenol and a major component of green tea, could effectively inhibit the interaction between the receptor-binding domain of the severe acute respiratory syndrome coronavirus 2 spike protein and the human cell receptor angiotensin-converting enzyme 2. Alternately, in silico molecular docking studies of epigallocatechin gallate and angiotensin-converting enzyme 2 indicated a binding score of −7.8 kcal/mol and identified a hydrogen bond between R393 and angiotensin-converting enzyme 2, which is considered as a key interacting residue involved in binding with the severe acute respiratory syndrome coronavirus 2 spike protein receptor-binding domain, suggesting the possible blocking of interaction between receptor-binding domain and angiotensin-converting enzyme 2. Furthermore, epigallocatechin gallate could attenuate severe acute respiratory syndrome coronavirus 2 infection and replication in Caco-2 cells. These results shed insight into identification and validation of severe acute respiratory syndrome coronavirus 2 entry inhibitors.

Bioactive components of different nasal spray solutions may defeat SARS-Cov2: repurposing and in silico studies.

The recent outbreak "Coronavirus Disease 2019 (COVID-19)" is caused by fast-spreading and highly contagious severe acute respiratory syndrome coronavirus 2 (SARS-CoV2). This virus enters into the human respiratory system by binding of the viral surface spike glycoprotein (S-protein) to an angiotensin-converting enzyme2 (ACE2) receptor that is found in the nasal passage and oral cavity of a human. Both spike protein and the ACE2 receptor have been identified as promising therapeutic targets to develop anti-SARS-CoV2 drugs. No therapeutic drugs have been developed as of today except for some vaccines. Therefore, potent therapeutic agents are urgently needed to combat the COVID-19 infections. This goal would be achieved only by applying drug repurposing and computational approaches. Thus, based on drug repurposing approach, we have investigated 16 bioactive components (1-16) from different nasal spray solutions to check their efficacies against human ACE2 and SARS-CoV2 spike proteins by performing molecular docking and molecular dynamic (MD) simulation studies. In this study, three bioactive components namely ciclesonide (8), levocabastine (13), and triamcinolone acetonide (16) have been found as promising inhibitory agents against SARS-CoV2 spike and human ACE2 receptor proteins with excellent binding affinities, comparing to reference drugs such as nafamostat, arbidol, losartan, and benazepril. Furthermore, MD simulations were performed (triplicate) for 100ns to confirm the stability of 8, 13, and 16 with said protein targets and to compute MM-PBSA-based binding-free energy calculations. Thus, bioactive components 8, 13, and 16 open the door for researchers and scientist globally to investigate them against SARS-CoV2 through in vitro and in vivo analysis.

Third Dose of the BNT162b2 Vaccine Results in Sustained High Levels of Neutralizing Antibodies Against SARS-CoV-2 at 6 Months Following Vaccination in Healthy Individuals.

Third Dose of the BNT162b2 Vaccine Results in Sustained High Levels of Neutralizing Antibodies Against SARS-CoV-2 at 6 Months Following Vaccination in Healthy Individuals.

Singular Interface Dynamics of the SARS-CoV-2 Delta Variant Explained with Contact Perturbation Analysis.

Emerging SARS-CoV-2 variants raise concerns about our ability to withstand the Covid-19 pandemic, and therefore, understanding mechanistic differences of those variants is crucial. In this study, we investigate disparities between the SARS-CoV-2 wild type and five variants that emerged in late 2020, focusing on the structure and dynamics of the spike protein interface with the human angiotensin-converting enzyme 2 (ACE2) receptor, by using crystallographic structures and extended analysis of microsecond molecular dynamics simulations. Dihedral angle principal component analysis (PCA) showed the strong similarities in the spike receptor binding domain (RBD) dynamics of the Alpha, Beta, Gamma, and Delta variants, in contrast with those of WT and Epsilon. Dynamical perturbation networks and contact PCA identified the peculiar interface dynamics of the Delta variant, which cannot be directly imputable to its specific L452R and T478K mutations since those residues are not in direct contact with the human ACE2 receptor. Our outcome shows that in the Delta variant the L452R and T478K mutations act synergistically on neighboring residues to provoke drastic changes in the spike/ACE2 interface; thus a singular mechanism of action eventually explains why it dominated over preceding variants.

Probing structural basis for enhanced binding of SARS-CoV-2 P.1 variant spike protein with the human ACE2 receptor.

The initial step of infection by severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) involves the binding of receptor binding domain (RBD) of the spike protein to the angiotensin converting enzyme 2 (ACE2) receptor. Each successive wave of SARS‐CoV‐2 reports emergence of many new variants, which is associated with mutations in the RBD as well as other parts of the spike protein. These mutations are reported to have enhanced affinity towards the ACE2 receptor as well as are also crucial for the virus transmission. Many computational and experimental studies have demonstrated the effect of individual mutation on the RBD‐ACE2 binding. However, the cumulative effect of mutations on the RBD and away from the RBD was not investigated in detail. We report here a comparative analysis on the structural communication and dynamics of the RBD and truncated S1 domain of spike protein in complex with the ACE2 receptor from SARS‐CoV‐2 wild type and its P.1 variant. Our integrative network and dynamics approaches highlighted a subtle conformational changes in the RBD as well as truncated S1 domain of spike protein at the protein contact level, responsible for the increased affinity with the ACE2 receptor. Moreover, our study also identified the commonalities and differences in the dynamics of the interactions between spike protein of SARS‐CoV‐2 wild type and its P.1 variant with the ACE2 receptor. Further, our investigation yielded an understanding towards identification of the unique RBD residues crucial for the interaction with the ACE2 host receptor. Overall, the study provides an insight for designing better therapeutics against the circulating P.1 variants as well as other future variants.

Assessment of mutations on RBD in the Spike protein of SARS-CoV-2 Alpha, Delta and Omicron variants

The severe acute respiratory syndrome (SARS) coronavirus 2 (CoV-2) variant Omicron spread more rapid than the other variants of SARS-CoV-2 virus. Mutations on the Spike (S) protein receptor-binding domain (RBD) are critical for the antibody resistance and infectivity of the SARS-CoV-2 variants. In this study, we have used accelerated molecular dynamics (aMD) simulations and free energy calculations to present a systematic analysis of the affinity and conformational dynamics along with the interactions that drive the binding between Spike protein RBD and human angiotensin-converting enzyme 2 (ACE2) receptor. We evaluate the impacts of the key mutation that occur in the RBDs Omicron and other variants in the binding with the human ACE2 receptor. The results show that S protein Omicron has stronger binding to the ACE2 than other variants. The evaluation of the decomposition energy per residue shows the mutations N440K, T478K, Q493R and Q498R observed in Spike protein of SARS-CoV-2 provided a stabilization effect for the interaction between the SARS-CoV-2 RBD and ACE2. Overall, the results demonstrate that faster spreading of SARS-CoV-2 Omicron may be correlated with binding affinity of S protein RBD to ACE2 and mutations of uncharged residues to positively charged residues such as Lys and Arg in key positions in the RBD.

Structure-Based Development of SARS-CoV-2 Spike Interactors.

Coronaviruses, including SARS-CoV-2 (the etiological agent of the current COVID-19 pandemic), rely on the surface spike glycoprotein to access the host cells, mainly through the interaction of their receptor-binding domain (RBD) with the human angiotensin-converting enzyme 2 (ACE2). Therefore, molecular entities able to interfere with the binding of the SARS-CoV-2 spike protein to ACE2 have great potential to inhibit viral entry. Starting from the available structural data on the interaction between SARS-CoV-2 spike protein and the host ACE2 receptor, we engineered a set of soluble and stable spike interactors, here denoted as S-plugs. Starting from the prototype S-plug, we adopted a computational approach by combining stability prediction, associated to single-point mutations, with molecular dynamics to enhance both S-plug thermostability and binding affinity to the spike protein. The best developed molecule, S-plug3, possesses a highly stable α-helical con-formation (with melting temperature Tm of 54 °C) and can interact with the spike RBD and S1 domains with similar low nanomolar affinities. Importantly, S-plug3 exposes the spike RBD to almost the same interface as the human ACE2 receptor, aimed at the recognition of all ACE2-accessing coronaviruses. Consistently, S-plug3 preserves a low nanomolar dissociation constant with the delta B.1.617.2 variant of SARS-CoV-2 spike protein (KD = 29.2 ± 0.6 nM). Taken together, we provide valid starting data for the development of therapeutical and diagnostic tools against coronaviruses accessing through ACE2.