SmartVax-Bp: An Immunoinformatic guided Multi-Epitope Vaccine Blueprint Against Burkholderia pseudomallei.

Burkholderia pseudomallei (B. pseudomallei) is the causative agent of melioidosis, a potentially serious and life-threatening infectious disease in humans. This pathogen invades the bloodstream and infects multiple organs, including the lungs, skin, liver, spleen, bones, and brain. The enzymes polyphosphate kinase 1 (Ppk1), type III secretion system cluster 3 (TTSS3)-effector protein (BopA), and trehalose-phosphate phosphatase (TPP) are vital for the survival and pathogenicity of B. pseudomallei, playing central roles in oxidative phosphorylation, stress responses, metabolism, biofilm formation, autophagy evasion, and motility. The normal function of these enzymes enables B. pseudomallei to produce energy and sustain fatal infections in affected organs. Due to their critical roles, these enzymes were identified as key targets for multi-epitope vaccine (MEV) design, with the aim of suppressing their expression in infected individuals by provoking specific immune responses. After retrieving the virulence factors Ppk1, BopA, and TPP, they were processed for B-cell and T-cell epitope prediction. Afterwards, predicted epitopes were filtered based on acceptable properties, including antigenicity, allergenicity, solubility, and toxicity. The filtered epitopes were linked using specific linkers and an adjuvant to create a finalized MEV construct. The structure of MEV was predicted and refined, demonstrating significant stability with a verify3D score of 90.28%. The MEV was further docked with immune receptors MHC-I, MHC-II, and TLR4, showing binding potentials of -1191.5 Kcal/mol, -1131.7 Kcal/mol, and - 1135.7 Kcal/mol, and maximum interactions, respectively. Molecular dynamics simulations of the MEV-docked complexes revealed a significant increase in B-factor and lower eigenvalues, indicating favorable and high-affinity interactions with the immune receptors. Furthermore, immune simulation analyses demonstrated a substantial production of immunoglobulins against the MEV, confirming its potential as an ideal antigen with low toxicity, optimal solubility, and no allergenicity. Based on these promising properties, the MEV was virtually cloned into a pET-28a(+) vector and expressed in Escherichia coli (E. coli). This suggests that the data can be used by experimentalists to produce the MEV in the lab and make it available to the community after clinical trials.