Insights into the Binding of Receptor-Binding Domain (RBD) of SARS-CoV-2 Wild Type and B.1.620 Variant with hACE2 Using Molecular Docking and Simulation Approaches

Recently, a new variant, B.1620, with mutations (S477N-E484K) in the spike protein’s receptor-binding domain (RBD) has been reported in Europe. In order to design therapeutic strategies suitable for B.1.620, further studies are required. A detailed investigation of the structural features and variations caused by these substitutions, that is, a molecular level investigation, is essential to uncover the role of these changes. To determine whether and how the binding affinity of ACE2–RBD is affected, we used protein–protein docking and all-atom simulation approaches. Our analysis revealed that B.1.620 binds more strongly than the wild type and alters the hydrogen bonding network. The docking score for the wild type was reported to be −122.6 +/− 0.7 kcal/mol, while for B.1.620, the docking score was −124.9 +/− 3.8 kcal/mol. A comparative binding investigation showed that the wild-type complex has 11 hydrogen bonds and one salt bridge, while the B.1.620 complex has 14 hydrogen bonds and one salt bridge, among which most of the interactions are preserved between the wild type and B.1.620. A dynamic analysis of the two complexes revealed stable dynamics, which corroborated the global stability trend, compactness, and flexibility of the three essential loops, providing a better conformational optimization opportunity and binding. Furthermore, binding free energy revealed that the wild type had a total binding energy of −51.14 kcal/mol, while for B.1.628, the total binding energy was −68.25 kcal/mol. The current findings based on protein complex modeling and bio-simulation methods revealed the atomic features of the B.1.620 variant harboring S477N and E484K mutations in the RBD and the basis for infectivity. In conclusion, the current study presents distinguishing features of B.1.620, which can be used to design structure-based drugs against the B.1.620 variant.

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Destabilizing the Structural Integrity of SARS-CoV2 Receptor Proteins by Curcumin Along with Hydroxychloroquine: An Insilco Approach for a Combination Therapy

10.26434/chemrxiv.12090438.v1 ◽

2020 ◽

Cited By ~ 1

Author(s):

Akhileshwar Srivastava ◽

Divya Singh

Keyword(s):

Binding Energy ◽

Combination Therapy ◽

Receptor Binding ◽

Structural Integrity ◽

Binding Domain ◽

Receptor Binding Domain ◽

Receptor Proteins ◽

Main Protease ◽

The Stability ◽

Therapeutic Activities

Presently, an emerging disease (COVID-19) has been spreading across the world due to coronavirus (SARS-CoV2). For treatment of SARS-CoV2 infection, currently hydroxychloroquine has been suggested by researchers, but it has not been found enough effective against this virus. The present study based on in silico approaches was designed to enhance the therapeutic activities of hydroxychloroquine by using curcumin as an adjunct drug against SARS-CoV2 receptor proteins: main-protease and S1 receptor binding domain (RBD). The webserver (ANCHOR) showed the higher protein stability for both receptors with disordered score (<0.5). The molecular docking analysis revealed that the binding energy (-24.58 kcal/mol) of hydroxychloroquine was higher than curcumin (-20.47 kcal/mol) for receptor main-protease, whereas binding energy of curcumin (<a>-38.84</a> kcal/mol) had greater than hydroxychloroquine<a> (-35.87</a> kcal/mol) in case of S1 receptor binding domain. Therefore, this study suggested that the curcumin could be used as combination therapy along with hydroxychloroquine for disrupting the stability of SARS-CoV2 receptor proteins

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Correlation of the commercial anti-SARS-CoV-2 receptor binding domain antibody test with the chemiluminescent reduction neutralizing test and possible detection of antibodies to emerging variants

10.1101/2021.05.25.21257828 ◽

2021 ◽

Author(s):

Yoshitomo Morinaga ◽

Hideki Tani ◽

Yasushi Terasaki ◽

Satoshi Nomura ◽

Hitoshi Kawasuji ◽

...

Keyword(s):

Receptor Binding ◽

Disease Onset ◽

Antibody Test ◽

Binding Domain ◽

Receptor Binding Domain ◽

Healthy Individuals ◽

Wild Type ◽

Serum Samples ◽

Serological Tests ◽

Convalescent Phase

Background Serological tests are beneficial for recognizing the immune response against SARS-CoV-2. To identify protective immunity, optimization of the chemiluminescent reduction neutralizing test (CRNT), using pseudotyped SARS-CoV-2, is critical. Whether commercial antibody tests are comparably accurate is unknown. Methods Serum samples collected before variants were locally found were obtained from confirmed COVID-19 patients (n = 74), confirmed non-COVID-19 individuals (n = 179), and unscreened individuals (suspected healthy individuals, n = 229). The convalescent phase was defined as the period after day 10 from disease onset. The CRNT against pseudotyped viruses displaying the wild-type spike protein and a commercially available anti-receptor binding domain (RBD) antibody test were assayed. The CRNT was also assayed, using South African (SA) and United Kingdom (UK)-derived variants. Results The CRNT (cut off value, 50% inhibition) and the anti-RBD antibody test (cut off value, 0.8 U/mL) concurred regarding symptomatic COVID-19 patients in the convalescent phase and clearly differentiated between patients and suspected healthy individuals (sensitivity; 95.8% and 100%, specificity; 99.1% and 100%, respectively). Anti-RBD antibody test results correlated with neutralizing titer (r = 0.47, 95% CI 0.20-0.68). Compared with the wild-type, CRNT reduction was observed for the SA and UK-derived variants. Of the samples with ≥100 U/mL by the anti-RBD antibody test, 77.8% and 88.9% showed ≥50% neutralization against the UK and the SA variants, respectively. Conclusion The CRNT and commercial anti-RBD antibody test effectively classified convalescent COVID-19 patients. The strong positive results using the commercial antibody test can reflect neutralizing activity against emerging variants.

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Discovery of potential inhibitors of the receptor-binding domain (RBD) of pandemic disease-causing SARS-CoV-2 Spike Glycoprotein from Triphala through molecular docking

Current Chinese Chemistry ◽

10.2174/2666001601666210322121802 ◽

2021 ◽

Vol 01 ◽

Author(s):

Sharuk L. Khan ◽

Falak A. Siddiqui ◽

Mohd Sayeed Shaikh ◽

Nitin V. Nema ◽

Aijaz A. Shaikh

Keyword(s):

Molecular Docking ◽

Hydrogen Bonds ◽

Receptor Binding ◽

Chemical Constituents ◽

Antioxidant Properties ◽

Binding Domain ◽

Quality Data ◽

Receptor Binding Domain ◽

Spike Glycoprotein

Background: COVID-19 (SARS-CoV-2 infection) has affected almost every region of the world. Presently, there is no defined line of treatment available for it. Triphala is already proven to have a safe biological window and well known for its antioxidant and immunomodulatory properties. Objective: Present work has been carried out to study Triphala's effectiveness for the treatment of COVID-19. Methods: The Receptor-binding domain (RBD) of SARS-CoV-2 Spike Glycoprotein responsible for the invasion into the host cell, which leads to further infection. The molecular docking (MD) was performed to explore the binding affinities (kcal/mol) of Triphala's chemical constituents and compared them with the existing drugs under investigation for the treatment of COVID-19 epidemiology. Results: Chebulinic acid binding affinity -8.5 kcal/mol with the formation of 10 hydrogen bonds. Almost all the major chemical constituents have formed two or more hydrogen bonds with RBD of SARS-CoV-2 Spike Glycoprotein. Conclusion: The present study showed that Triphala might perform vital roles in the treatment of COVID-19 and expand its usefulness to physicians to treat this illness. There is a need to complete the in-vitro, in-vivo biological testing of Triphala on SARS-CoV-2 disease to create more quality data. The binding mode of Chebulinic acid in the allosteric cavity allows a better understanding of RBD of SARS-CoV-2 Spike Glycoprotein target and provides insight for the design of new inhibitors. Triphala is already proven to have a safe biological window, which indicates we can skip the pre-clinical trials. Apart from this, Triphala is well known for its antioxidant properties, which ultimately improves the immunity of the COVID-19 patient.

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Epistasis at the SARS-CoV-2 RBD Interface and the Propitiously Boring Implications for Vaccine Escape

10.1101/2021.08.30.458225 ◽

2021 ◽

Cited By ~ 1

Author(s):

Nash D. Rochman ◽

Guilhem Faure ◽

Yuri I. Wolf ◽

Peter L. Freddolino ◽

Feng Zhang ◽

...

Keyword(s):

Receptor Binding ◽

Neutralizing Antibodies ◽

Binding Domain ◽

Receptor Binding Domain ◽

Wild Type ◽

Protein Structure Modeling ◽

Escape Mutants ◽

Small Set ◽

Global Concern ◽

The Impact

AbstractAt the time of this writing, August 2021, potential emergence of vaccine escape variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a grave global concern. The interface between the receptor-binding domain (RBD) of SARS-CoV-2 spike (S) protein and the host receptor (ACE2) overlap with the binding site of principal neutralizing antibodies (NAb), limiting the repertoire of viable mutations. Nonetheless, variants with multiple mutations in the RBD have rose to dominance. Non-additive, epistatic relationships among RBD mutations are apparent, and assessing the impact of such epistasis on the mutational landscape is crucial. Epistasis can substantially increase the risk of vaccine escape and cannot be completely characterized through the study of the wild type (WT) alone. We employed protein structure modeling using Rosetta to compare the effects of all single mutants at the RBD-NAb and RBD-ACE2 interfaces for the WT, Gamma (417T, 484K, 501Y), and Delta variants (452R, 478K). Overall, epistasis at the RBD surface appears to be limited and the effects of most multiple mutations are additive. Epistasis at the Delta variant interface weakly stabilizes NAb interaction relative to ACE2, whereas in the Gamma variant, epistasis more substantially destabilizes NAb interaction. These results suggest that the repertoire of potential escape mutations for the Delta variant is not substantially different from that of the WT, whereas Gamma poses a moderately greater risk for enhanced vaccine escape. Thus, the modest ensemble of mutations relative to the WT shown to reduce vaccine efficacy might constitute the majority of all possible escape mutations.SignificancePotential emergence of vaccine escape variants of SARS-CoV-2 is arguably the most pressing problem during the COVID-19 pandemic as vaccines are distributed worldwide. We employed a computational approach to assess the risk of antibody escape resulting from mutations in the receptor-binding domain of the spike protein of the wild type SARS-CoV-2 virus as well as the Gamma and Delta variants. The results indicate that emergence of escape mutants is somewhat less likely for the Delta variant than for the wild type and moderately more likely for the Gamma variant. We conclude that the small set of escape-enhancing mutations already identified for the wild type is likely to include the majority of all possible mutations with this effect, a welcome finding.

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An Engineered Receptor-Binding Domain Improves the Immunogenicity of Multivalent SARS-CoV-2 Vaccines

mBio ◽

10.1128/mbio.00930-21 ◽

2021 ◽

Vol 12 (3) ◽

Author(s):

Yan Guo ◽

Wenhui He ◽

Huihui Mou ◽

Lizhou Zhang ◽

Jing Chang ◽

...

Keyword(s):

Amino Acid ◽

Receptor Binding ◽

Binding Domain ◽

Full Length ◽

Vaccine Antigen ◽

Receptor Binding Domain ◽

Protein Vaccine ◽

Wild Type ◽

S Protein ◽

Glycosylation Sites

ABSTRACT The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) protein mediates viral entry into cells expressing angiotensin-converting enzyme 2 (ACE2). The S protein engages ACE2 through its receptor-binding domain (RBD), an independently folded 197-amino-acid fragment of the 1,273-amino-acid S-protein protomer. The RBD is the primary SARS-CoV-2 neutralizing epitope and a critical target of any SARS-CoV-2 vaccine. Here, we show that this RBD conjugated to each of two carrier proteins elicited more potent neutralizing responses in immunized rodents than did a similarly conjugated proline-stabilized S-protein ectodomain. Nonetheless, the native RBD is expressed inefficiently, limiting its usefulness as a vaccine antigen. However, we show that an RBD engineered with four novel glycosylation sites (gRBD) is expressed markedly more efficiently and generates a more potent neutralizing responses as a DNA vaccine antigen than the wild-type RBD or the full-length S protein, especially when fused to multivalent carriers, such as a Helicobacter pylori ferritin 24-mer. Further, gRBD is more immunogenic than the wild-type RBD when administered as a subunit protein vaccine. Our data suggest that multivalent gRBD antigens can reduce costs and doses, and improve the immunogenicity, of all major classes of SARS-CoV-2 vaccines. IMPORTANCE All available vaccines for coronavirus disease 2019 (COVID-19) express or deliver the full-length SARS-CoV-2 spike (S) protein. We show that this antigen is not optimal, consistent with observations that the vast majority of the neutralizing response to the virus is focused on the S-protein receptor-binding domain (RBD). However, this RBD is not expressed well as an independent domain, especially when expressed as a fusion protein with a multivalent scaffold. We therefore engineered a more highly expressed form of the SARS-CoV-2 RBD by introducing four glycosylation sites into a face of the RBD normally occluded in the full S protein. We show that this engineered protein, gRBD, is more immunogenic than the wild-type RBD or the full-length S protein in both genetic and protein-delivered vaccines.

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An engineered receptor-binding domain improves the immunogenicity of multivalent SARS-CoV-2 vaccines

10.1101/2020.11.18.388934 ◽

2020 ◽

Author(s):

Brian D. Quinlan ◽

Wenhui He ◽

Huihui Mou ◽

Lizhou Zhang ◽

Yan Guo ◽

...

Keyword(s):

Amino Acid ◽

Receptor Binding ◽

Viral Entry ◽

Binding Domain ◽

Vaccine Antigen ◽

Receptor Binding Domain ◽

Protein Vaccine ◽

Wild Type ◽

S Protein ◽

H Pylori

ABSTRACTThe SARS-coronavirus 2 (SARS-CoV-2) spike (S) protein mediates viral entry into cells expressing the angiotensin-converting enzyme 2 (ACE2). The S protein engages ACE2 through its receptor-binding domain (RBD), an independently folded 197-amino acid fragment of the 1273-amino acid S-protein protomer. The RBD is the primary SARS-CoV-2 neutralizing epitope and a critical target of any SARS-CoV-2 vaccine. Here we show that this RBD conjugated to each of two carrier proteins elicited more potent neutralizing responses in immunized rodents than did a similarly conjugated proline-stabilized S-protein ectodomain. Nonetheless, the native RBD expresses inefficiently, limiting its usefulness as a vaccine antigen. However, we show that an RBD engineered with four novel glycosylation sites (gRBD) expresses markedly more efficiently, and generates a more potent neutralizing responses as a DNA vaccine antigen, than the wild-type RBD or the full-length S protein, especially when fused to multivalent carriers such as an H. pylori ferritin 24-mer. Further, gRBD is more immunogenic than the wild-type RBD when administered as a subunit protein vaccine. Our data suggest that multivalent gRBD antigens can reduce costs and doses, and improve the immunogenicity, of all major classes of SARS-CoV-2 vaccines.

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Design, synthesis and in silico studies of novel N-(2-aminophenyl)-2,3-diphenylquinoxaline-6-sulfonamide derivatives targeting receptor-binding domain (RBD) of SARS-CoV-2 Spike Glycoprotein and evaluation as antimicrobial and antimalarial agents

Letters in Drug Design & Discovery ◽

10.2174/1570180818666210427095203 ◽

2021 ◽

Vol 18 ◽

Author(s):

Falak A. Siddiqui ◽

Sharuk L. Khan ◽

Rajendra P Marathe ◽

Nitin V. Nemac

Keyword(s):

Hydrogen Bonds ◽

Receptor Binding ◽

Drug Repurposing ◽

Binding Domain ◽

Receptor Binding Domain ◽

Spike Glycoprotein ◽

Design Synthesis ◽

Angiotensin Converting Enzyme 2 ◽

In Silico Studies ◽

Novel Coronavirus

Background: Pneumonia induced by a novel coronavirus (SARS-CoV-2) was named as coronavirus disease 2019 (COVID-19). The Receptor-binding domain (RBD) of SARS-CoV-2 Spike Glycoprotein, causes invasion of the virus into the host cell by attaching with human angiotensin-converting enzyme-2 (hACE-2) which leads to further infection. Objectives: The novel N-(2-aminophenyl)-2,3-diphenylquinoxaline-6-sulfonamide derivatives were designed and synthesized to inhibit the RBD of SARS-CoV-2 Spike Glycoprotein by applying molecular docking tools. Methods: The synthesized products was characterized by Infrared Spectroscopy (IR), and 1H Nuclear Magnetic Resonance (NMR). Results: All the derivatives were found to have a very good binding affinity between -9 to -10.1 kcal/mol, better than the drugs which are under investigation for the treatment of SARS-CoV-2 infection. Compound F1 has formed 4 hydrogen bonds whereas, F4 and F10 formed two hydrogen bonds each with RBD of SARS-CoV-2 Spike Glycoprotein. All the derivatives were subjected to antimicrobial, antifungal, and antimalarial susceptibility. Conclusion: From the above-obtained results, we have concluded that novel N-(2-aminophenyl)-2,3-diphenylquinoxaline-6-sulfonamide derivatives have excellent potential to inhibit the receptor-binding domain (RBD) of SARS-CoV-2 Spike Glycoprotein, which is now an attentive target in designing SARS-CoV-2 inhibitors. This scaffold can hold an effective interest in the development of inhibitors for SARS-CoV-2 in the future if drug repurposing fails to serve the purpose.

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Synthesis of a Cys949Tyr α2-macroglobulin thiol ester mutant: co-transfection with wild-type α2-macroglobulin in an episomal expression system

Biochemical Journal ◽

10.1042/bj3120183 ◽

1995 ◽

Vol 312 (1) ◽

pp. 183-190 ◽

Cited By ~ 1

Author(s):

L Van Rompaey ◽

H Van den Berghe ◽

P Marynen

Keyword(s):

Receptor Binding ◽

Covalent Binding ◽

Binding Domain ◽

Expression Vectors ◽

Receptor Binding Domain ◽

Wild Type ◽

Thiol Ester ◽

Thiol Esters ◽

Α2 Macroglobulin ◽

K 562 Cells

A full-length alpha 2-macroglobulin (alpha 2M) cDNA was cloned into the episomal expression vectors pREP7 and pMEP4. Electroporation of the cell lines WI-L2-729HF2, U-937, K-562 and an Epstein-Barr virus-transformed cell line resulted in stable transfectants only with K-562 cells. Stable expression was obtained exclusively with pMEP4-alpha 2M and was driven from the inducible human metallothionein IIA promoter. Expression of the wild-type alpha 2M cDNA resulted in a recombinant protein (r alpha 2M) that could not be distinguished from plasma alpha 2M (p-alpha 2M): the transfected K-562 cells secreted tetrameric alpha 2M with intact internal thiol esters, a functional bait domain and a latent receptor-binding domain. r alpha 2M inhibited trypsin and elastase from cleaving a high-molecular-mass substrate. When the Cys-949 involved in the formation of the internal thiol ester was mutated to tyrosine (C949Y-r alpha 2M), a tetrameric alpha 2M was secreted, with the electrophoretic mobility of methylamine-treated p-alpha 2M (p-alpha 2M/MA) and with a functional receptor-binding domain. The C949Y-r alpha 2M did not possess proteinase-inhibiting capacity. Heterozygosity was mimicked by co-transfecting the K-562 cells with wild-type and mutant expression vectors. In this case, r alpha 2M was secreted with zero, one, two, three or four internal thiol esters. A comparison of the interaction of interleukin 1 beta and basic fibroblast growth factor with native p-alpha 2M, p-alpha 2M/MA and the mutant C949Y-r alpha 2M revealed that when assayed under nondenaturing conditions, no binding occurred to ‘slow’ p-alpha 2M whereas quantitatively similar binding was observed to ‘fast’ p-alpha 2M/MA and C949Y-r alpha 2M. Covalent binding, however, was essentially limited to p-alpha 2M/MA, suggesting the involvement of Cys-949 in the process. Covalent binding of insulin, on the contrary, was only observed when it was present during hydrolysis of the internal thiol esters of p-alpha 2M by trypsin treatment, and thus involves the activated Glx residue.

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SARS-CoV-2 Omicron Spike Glycoprotein Receptor Binding Domain Exhibits Super-Binder Ability with ACE2 but not Convalescent Monoclonal Antibody

10.1101/2021.12.09.471885 ◽

2021 ◽

Author(s):

Olaposi Idowu Omotuyi ◽

Olubiyi Olujide ◽

Oyekanmi Nash ◽

Elizabeth O Afolabi ◽

Babatunji Oyinloye ◽

...

Keyword(s):

Monoclonal Antibody ◽

Receptor Binding ◽

Md Simulation ◽

Center Of Mass ◽

Salt Bridge ◽

Binding Domain ◽

Receptor Binding Domain ◽

Spike Glycoprotein ◽

Neutralizing Monoclonal Antibody ◽

Complementarity Determining Region

Background: SARS-CoV-2, the causative virus for COVID-19 has now super-mutated into the Omicron (Om) variant. On its spike glycoprotein alone, more than 30 substitutions have been characterized with 15 within the receptor binding domain (RBD); It therefore calls to question the transmissibility and antibody escapability of Omicron. This study was setup to investigate the Omicron RBD interaction with ACE2 (host receptor) and a SARS-CoV-2 neutralizing monoclonal antibody (mAb). Methods: In-silico mutagenesis was used to generate the Om-RBD in complex with ACE2 or mAb from the wildtype. All-atom molecular dynamics (MD) simulation trajectories were analyzed for interaction. Results: MD trajectories showed that Omicron RBD has evolved into an efficient ACE2 binder, via pi-pi (Om-RBD-Y501/ACE2-Y41) and salt-bridge (Om-RBD-K493/ACE2-Y41) interactions. Conversely, in binding mAb, it has become less efficient (Center of mass distance of RBD from mAb complex, wildtype-RBD =30 A, Omicron-RBD= 41 A). Disruption of Om-RBD/mAb complex resulted from loose interaction between Om-RBD and the light chain complementarity-determining region residues. Conclusions: Omicron is expected to be better transmissible and less efficiently interacting with neutralizing convalescent mAbs. General significance: Our results elucidate the mechanisms for higher transmissibility in Omicron variant.

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Stabilization of the SARS-CoV-2 Spike receptor-binding domain using deep mutational scanning and structure-based design

10.1101/2021.05.15.444222 ◽

2021 ◽

Author(s):

Daniel Ellis ◽

Natalie Brunette ◽

Katherine H. D. Crawford ◽

Alexandra C. Walls ◽

Minh N. Pham ◽

...

Keyword(s):

Receptor Binding ◽

Large Scale ◽

Protein S ◽

Binding Pocket ◽

Binding Domain ◽

Effective Vaccine ◽

Receptor Binding Domain ◽

Wild Type ◽

Vaccine Candidates ◽

Generation Vaccine

The unprecedented global demand for SARS-CoV-2 vaccines has demonstrated the need for highly effective vaccine candidates that are thermostable and amenable to large-scale manufacturing. Nanoparticle immunogens presenting the receptor-binding domain (RBD) of the SARS-CoV-2 Spike protein (S) in repetitive arrays are being advanced as second-generation vaccine candidates, as they feature robust manufacturing characteristics and have shown promising immunogenicity in preclinical models. Here, we used previously reported deep mutational scanning (DMS) data to guide the design of stabilized variants of the RBD. The selected mutations fill a cavity in the RBD that has been identified as a linoleic acid binding pocket. Screening of several designs led to the selection of two lead candidates that expressed at higher yields than the wild-type RBD. These stabilized RBDs possess enhanced thermal stability and resistance to aggregation, particularly when incorporated into an icosahedral nanoparticle immunogen that maintained its integrity and antigenicity for 28 days at 35-40°C, while corresponding immunogens displaying the wild-type RBD experienced aggregation and loss of antigenicity. The stabilized immunogens preserved the potent immunogenicity of the original nanoparticle immunogen, which is currently being evaluated in a Phase I/II clinical trial. Our findings may improve the scalability and stability of RBD-based coronavirus vaccines in any format and more generally highlight the utility of comprehensive DMS data in guiding vaccine design.

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