Exploration of surface glycoprotein to design multi-epitope vaccine for the prevention of Covid-19.
ABSTRACT: Stimulation and generation of T and B cell-mediated long-term immune response are essential for the curbing of a deadly virus such as SAR-CoV-2 (Severe Acute Respiratory Corona Virus 2). Immunoinformatics approach in vaccine design takes advantage of antigenic and non-allergenic epitopes present on the spike glycoprotein of SARS-CoV-2 to elicit immune responses. T cells and B cells epitopes were predicted, and the selected residues were subjected to allergenicity, antigenicity and toxicity screening which were linked by appropriate linkers to form a multi-epitope subunit vaccine. The physiochemical properties of the vaccine construct were analyzed, and the molecular weight, molecular formula, theoretical isoelectric point value, half-life, solubility score, instability index, aliphatic index and GRAVY were predicted. The vaccine structure was constructed, refined, validated, and disulfide engineered to get the best model. Molecular binding simulation and molecular dynamics simulation were carried out to predict the stability and binding affinity of the vaccine construct with TLRs. Codon acclimatization and in silico cloning were performed to confirm the vaccine expression and potency. Results obtained indicated that this novel vaccine candidate is non-toxic, capable of initiating the immunogenic response and will not induce an allergic reaction. The highest binding energy was observed in TLR4 (Toll-like Receptor 4) (-1398.1), and the least is TLR 2 (-1479.6). The steady rise in Th (T-helper) cell population with memory development was noticed, and IFN-g (Interferon gamma) was provoked after simulation. At this point, the vaccine candidate awaits animal trial to validate its efficacy and safety for use in the prevention of the novel COVID-19 (Coronavirus Disease 2019) infections.
Project description:BACKGROUND:The novel SARS-CoV-2 coronavirus, the causative agent of the ongoing pandemic COVID-19 disease continues to infect people globally and has infected millions of humans worldwide. However, no effective vaccine against this virus exists. METHOD:Using Immunoinformatics, epitopic sequences from multiple glycoproteins that play crucial role in pathogenesis were identified. Particularly, epitopes were mapped from conserved receptor-binding domain of spike protein which have been experimentally validated in SARS-CoV-1 as a promising target for vaccine development. RESULTS:A multi-epitopic vaccine construct comprising of B-cell, CTL, HTL epitopes was developed along with fusion of adjuvant and linkers. The epitopes identified herein are reported for the first time and were predicted to be highly antigenic, stable, nonallergen, nontoxic and displayed conservation across several SARS-CoV-2 isolates from different countries. Additionally, the epitopes associated with maximum HLA alleles and population coverage analysis shows the proposed epitopes would be a relevant representative of large proportion of the world population. A reliable three-dimensional structure of the vaccine construct was developed. Consequently, docking and molecular-dynamics simulation ensured the stable interaction between vaccine and innate-immune receptor.
Project description:Middle East respiratory syndrome coronavirus (MERS-CoV) is an emerging health concern due to its high mortality rate of 35%. At present, no vaccine is available to protect against MERS-CoV infections. Therefore, an in silico search for potential antigenic epitopes in the non-redundant proteome of MERS-CoV was performed herein. First, a subtractive proteome-based approach was employed to look for the surface exposed and host non-homologous proteins. Following, immunoinformatics analysis was performed to predict antigenic B and T cell epitopes that were used in the design of a multi-epitopes peptide. Molecular docking study was carried out to predict vaccine construct affinity of binding to Toll-like receptor 3 (TLR3) and understand its binding conformation to extract ideas about its processing by the host immune system. We identified membrane protein, envelope small membrane protein, non-structural protein ORF3, non-structural protein ORF5, and spike glycoprotein as potential candidates for subunit vaccine designing. The designed multi-epitope peptide then linked to ?-defensin adjuvant is showing high antigenicity. Further, the sequence of the designed vaccine construct is optimized for maximum expression in the Escherichia coli expression system. A rich pattern of hydrogen and hydrophobic interactions of the construct was observed with the TLR3 allowing stable binding of the construct at the docked site as predicted by the molecular dynamics simulation and MM-PBSA binding energies. We expect that the panel of subunit vaccine candidates and the designed vaccine construct could be highly effective in immunizing populations from infections caused by MERS-CoV and could possible applied on the current pandemic COVID-19.
Project description:BACKGROUND:The coronavirus (CoV) spike (S) protein is critical for receptor binding, membrane fusion and internalization of the virus into the human cells. We have tried to search the epitopic component of the S-protein that might be served as crucial targets for the vaccine development and also tried to understand the molecular mechanism of epitopes and TLR4/MD-2 complex for adaptive immunity. MATERIAL AND METHODS:Here we identified the antigenicity and the epitopic divergence of S-protein via immunoinformatics approach. The study was performed to identify the epitopes, composition of amino acids and its distribution in epitopic regions, composition of amino acid between the identified epitopes, secondary structure architecture of epitopes, physicochemical and biochemical parameters and molecular interaction between the identified epitope and TLR4/MD-2 complex. The SARS-CoV-2 can be possibly recognised by TLR4 of host immune cells that are responsible for the adaptive immune response. RESULTS:We identified four SARS-CoV-2 S-protein 9mer antigenic epitopes and observed that they bind with the TLR4/MD-2 complex by varied stable molecular bonding interactions. Molecular interaction between these characterized epitopes with TLR4/MD-2 complex might be indicated the binding affinity and downstream signalling of adaptive immune response. Different physicochemical and biochemical parameters such as O-glycosylation and N-glycosylation, Hydrophobicity, GRAVY were identified within epitopic regions of S-protein. These parameters help to understand the protein-protein interaction between epitopes and TLR4/MD-2 complex. The study also revealed different epitopic binding pockets of TLR4/MD-2 complex. CONCLUSIONS:The identified epitopes impart suitable prospects for the development of novel peptide-based epitopic vaccine for the control of COVID-19 infection.
Project description:Ongoing COVID-19 outbreak has raised a drastic challenge to global public health security. Most of the patients with COVID-19 suffer from mild flu-like illnesses such as cold and fever; however, few percentages of the patients progress from severe illness to death, mostly in an immunocompromised individual. The causative agent of COVID-19 is an RNA virus known as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Despite these debilitating conditions, no medication to stop the disease progression or vaccination is available till now. Therefore, we aimed to formulate a multi-epitope vaccine against SARS-CoV-2 by utilizing an immunoinformatics approach. For this purpose, we used the SARS-CoV-2 spike glycoprotein to determine the immunodominant T- and B-cell epitopes. After rigorous assessment, we designed a vaccine construct using four potential epitopes from each of the three epitope classes such as cytotoxic T-lymphocytes, helper T-lymphocyte, and linear B-lymphocyte epitopes. The designed vaccine was antigenic, immunogenic, and non-allergenic with suitable physicochemical properties and has higher solubility. More importantly, the predicted vaccine structure was similar to the native protein. Further investigations indicated a strong and stable binding interaction between the vaccine and the toll-like receptor (TLR4). Strong binding stability and structural compactness were also evident in molecular dynamics simulation. Furthermore, the computer-generated immune simulation showed that the vaccine could trigger real-life-like immune responses upon administration into humans. Finally, codon optimization based on Escherichia coli K12 resulted in optimal GC content and higher CAI value followed by incorporating it into the cloning vector pET28+(a). Overall, these results suggest that the designed peptide vaccine can serve as an excellent prophylactic candidate against SARS-CoV-2. Communicated by Ramaswamy H. Sarma.
Project description:The COVID-19 disease is caused by SARS-CoV-2 and spreading rapidly worldwide with extremely high infection rate. Since effective and specific vaccine is not available to combat the deadly COVID-19, the objective of our study was to design a multi-epitope vaccine using immunoinformatics approach with translational implications. Nucleocapsid (N) protein of SARS-CoV-2 is stable, conserved and highly immunogenic along with being less prone to mutations during infection, which makes it a suitable candidate for designing vaccine. In our study, B- and T-cells epitopes were identified from N protein and screened based on crucial parameters to design the multi-epitope vaccine construct. Additionally, human beta-defensin-2 was incorporated into the vaccine construct as an adjuvant along with suitable linkers followed by its further evaluation based on crucial parameters including allergenicity, antigenicity, stability etc. Combined major histocompatibility complexes (MHC-I and MHC-II) binding epitopes presented broader population coverage of the vaccine throughout the world. The three-dimensional structure of vaccine candidate implied strong interaction with toll-like receptor 3 (TLR3) using molecular docking. The vaccine-TLR3 complex was observed to be highly stable during simulation and electrostatic free energy was foremost contributor for stabilization of the structure. Subsequently, in silico cloning of vaccine candidate was carried out to generate the construct into pET-28a(+) expression vector succeeded by its virtual confirmation. Altogether, our results advocate that the designed vaccine candidate could be an effective and promising weapon to fight with COVID-19 infection worldwide.
Project description:As the number of infections and deaths caused by the recent COVID-19 pandemic is increasing dramatically day-by-day, scientists are rushing towards developing possible countermeasures to fight the deadly virus, SARS-CoV-2. Although many efforts have already been put forward for developing potential vaccines; however, most of them are proved to possess negative consequences. Therefore, in this study, immunoinformatics methods were exploited to design a novel epitope-based subunit vaccine against the SARS-CoV-2, targeting four essential proteins of the virus i.e., spike glycoprotein, nucleocapsid phosphoprotein, membrane glycoprotein, and envelope protein. The highly antigenic, non-allergenic, non-toxic, non-human homolog, and 100% conserved (across other isolates from different regions of the world) epitopes were used for constructing the vaccine. In total, fourteen CTL epitopes and eighteen HTL epitopes were used to construct the vaccine. Thereafter, several <i>in silico</i> validations i.e., the molecular docking, molecular dynamics simulation (including the RMSF and RMSD studies), and immune simulation studies were also performed which predicted that the designed vaccine should be quite safe, effective, and stable within the biological environment. Finally, <i>in silico</i> cloning and codon adaptation studies were also conducted to design an effective mass production strategy of the vaccine. However, more <i>in vitro</i> and <i>in vivo</i> studies are required on the predicted vaccine to finally validate its safety and efficacy.
Project description:The COVID-19 pandemic caused by SARS-CoV-2 is a public health emergency of international concern and thus calling for the development of effective and safe therapeutics and prophylactics particularly a vaccine to protect against the infection. SARS-CoV-2 spike glycoprotein is an attractive candidate for a vaccine, antibodies, and inhibitors development because of the many roles it plays in attachment, fusion and entry into the host cell. In the present investigation, we characterized the SARS-CoV-2 spike glycoprotein by immunoinformatics techniques to put forward potential B and T cell epitopes, followed by the use of epitopes in construction of a multi-epitope peptide vaccine construct (MEPVC). The MEPVC revealed robust host immune system simulation with high production of immunoglobulins, cytokines and interleukins. Stable conformation of the MEPVC with a representative innate immune TLR3 receptor was observed involving strong hydrophobic and hydrophilic chemical interactions, along with enhanced contribution from salt-bridges towards inter-molecular stability. Molecular dynamics simulation in aqueous milieu aided further in interpreting strong affinity of the MEPVC for TLR3. This stability is the attribute of several vital residues from both TLR3 and MEPVC as shown by radial distribution function (RDF) and a novel axial frequency distribution (AFD) analytical tool. Comprehensive binding free energies estimation was provided at the end that concluded major domination by electrostatic and minor from van der Waals. Summing all, the designed MEPVC has tremendous potential of providing protective immunity against COVID-19 and thus could be considered in experimental studies.
Project description:SARS-CoV-2 is the deadly virus behind COVID-19, the disease that went on to ravage the world and caused the biggest pandemic 21st century has witnessed so far. On the face of ongoing death and destruction, the urgent need for the discovery of a vaccine against the virus is paramount. This study resorted to the emerging discipline of immunoinformatics in order to design a multi-epitope mRNA vaccine against the spike glycoprotein of SARS-CoV-2. Various immunoinformatics tools were utilized to predict T and B lymphocyte epitopes. The epitopes were channeled through a filtering pipeline comprised of antigenicity, toxicity, allergenicity, and cytokine inducibility evaluation with the goal of selecting epitopes capable of generating both T and B cell-mediated immune responses. Molecular docking simulation between the epitopes and their corresponding MHC molecules was carried out. 13 epitopes, a highly immunogenic adjuvant, elements for proper sub-cellular trafficking, a secretion booster, and appropriate linkers were combined for constructing the vaccine. The vaccine was found to be antigenic, almost neutral at physiological pH, non-toxic, non-allergenic, capable of generating a robust immune response and had a decent worldwide population coverage. Based on these parameters, this design can be considered a promising choice for a vaccine against SARS-CoV-2.
Project description:Coronavirus disease 2019 (COVID-19) is an acute pneumonic disease, with no prophylactic or specific therapeutical solution. Effective and rapid countermeasure against the spread of the disease's associated virus, SARS-CoV-2, requires to incorporate the computational approach. In this study, we employed various immunoinformatics tools to design a multi-epitope vaccine polypeptide with the highest potential for activating the human immune system against SARS-CoV-2. The initial epitope set was extracted from the whole set of viral structural proteins. Potential non-toxic and non-allergenic T-cell and B-cell binding and cytokine inducing epitopes were then identified through a priori prediction. Selected epitopes were bound to each other with appropriate linkers, followed by appending a suitable adjuvant to increase the immunogenicity of the vaccine polypeptide. Molecular modelling of the 3D structure of the vaccine construct, docking, molecular dynamics simulations and free energy calculations confirmed that the vaccine peptide had high affinity for Toll-like receptor 3 binding, and that the vaccine-receptor complex was highly stable. As our vaccine polypeptide design captures the advantages of structural epitopes and simultaneously integrates precautions to avoid relevant side effects, it is suggested to be promising for elicitation of an effective and safe immune response against SARS-CoV-2 in vivo.
Project description:COVID-19, caused by SARS-CoV-2, is severe respiratory illnesses leading to millions of deaths worldwide in very short span. The high case fatality rate and the lack of medical counter measures emphasize for an urgent quest to develop safe and effective vaccine. Receptor-binding domain (RBD) of spike protein of SARS-CoV-2 binds to the ACE2 receptor on human host cell for the viral attachment and entry, hence considered as a key target to develop vaccines, antibodies and therapeutics. In this study, immunoinformatics approach was employed to design a novel multi-epitope vaccine using RBD of SARS-CoV-2 spike protein. The potential B- and T-cell epitopes were selected from RBD sequence using various bioinformatics tools to design the vaccine construct. The in silico designed multi-epitope vaccine encompasses 146 amino acids with an adjuvant (human beta-defensin-2), which was further computationally evaluated for several parameters including antigenicity, allergenicity and stability. Subsequently, three-dimensional structure of vaccine construct was modelled and then docked with various toll-like receptors. Molecular dynamics (MD) study of docked TLR3-vaccine complex delineated it to be highly stable during simulation time and the stabilization of interaction was majorly contributed by electrostatic energy. The docked complex also showed low deformation and increased rigidity in motion of residues during dynamics. Furthermore, in silico cloning of the multi-epitope vaccine was carried out to generate the plasmid construct for expression in a bacterial system. Altogether, our study suggests that the designed vaccine candidate containing RBD region could provide the specific humoral and cell-mediated immune responses against SARS-CoV-2. Communicated by Ramaswamy H. Sarma.