An Innovative Approach towards Construction, Evaluation, Simulations and Cloning of a Multi-epitope Vaccine against Ebola Virus, through Immunoinformatics-based Study Design

Abstract

Introduction Ebola virus has been preying on human populations for a long time. But recent outbreaks in Africa with symptoms such as severe haemorrhagic fever urge the immediate need for research on an Ebola vaccine as an initial step. Hence, in this study, an in silico vaccine is designed against the Ebola virus, with epitopes representing all three strains: Zaire, Sudan, and Bundibugyo. Methodologies After initial sequence retrieval, they are prioritized based on physicochemical analyses and other suitable parameters. Epitopes are then predicted from selected sequences and subjected to stringent analyses – antigenicity, allergenicity, toxicity, conservation, solubility, among other parameters. An in silico vaccine is constructed using epitopes, incorporating Beta-defensin 3 at the C-terminus and a His6 Tag at the N-terminus. After a stringent set of analyses, the protein structure is subjected to Secondary and Tertiary structure prediction. After refinement of the structure, validation procedures are performed using PROCHECK, PRO-SA, and Molprobity. Simulation methods such as GROMACS and Immune Simulations, as implemented in C-IMMISIM, are used. Thereafter, docking with TLR4 (in PATCHDOCK software) , interaction analyses of the docking mechanism , by PISA software, Codon Optimization by JCAT and cloning into the pET28a(+) with SnapGene , are achieved. Results After applying stringent analytical methods, epitopes are selected for in silico vaccine construction against Ebola virus strains – Zaire, Sudan, and Bundibugyo. The constructed vaccine is subjected to similar analyses again, yielding a Vaxijen score of 0.6853. Secondary structure prediction by PSIPRED ensures reliable conformity, while tertiary structure prediction by Colab AlphaFold2 further reveals the vaccine structure. After refinement, the validation process reveals that the protein vaccine structure contains over 98% Ramachandran-favoured regions and an ERRAT score of 71%. Docking of the vaccine against the TLR4 immune receptor by PATCHDOCK is achieved, and PISA interaction analyses reveal a binding energy of -14.5kcal/mol. Again, Molecular Dynamics simulations reveal its structural integrity and conformity amid solvation analyses. Then, for C-IMMISIM results, both humoral and cell-mediated immunity are activated, along with memory cell propagation. Again, in JCAT, the CAI score is 0.9572, and a GC content of 52.95% is achieved, indicating high expression potential. Conclusion The immunoinformatics tools applied suggest its conformational integrity, compactness, and safety for further wet-lab research. World -class population coverage of 87.44% , along with approx 70% coverge for West, East and Central Africa , reflects the need for further labotory-based experiments with this vaccine candidate.