Multi-strain dengue antigen design: from epitope prediction to monoclonal antibody binding and dynamics

The present study describes the computational design and in silico evaluation of a multi-strain dengue antigen candidate intended for potential application in diagnostic platforms and as a vaccine-related component. Sixteen B-cell epitopes were initially retrieved from the Immune Epitope Database (IEDB) and screened for antigenicity and toxicity, leading to the selection of eight epitopes. These epitopes were joined using KK linkers and optimized to generate a 17 kDa construct exhibiting favourable stability and hydrophilicity. Immunogenicity predictions suggested a robust in silico antibody response. Molecular docking with the dengue-specific monoclonal antibody 1A1D-2 yielded a stable antigen-antibody complex, and molecular dynamics simulations demonstrated enhanced stability of the antigen in the complexed state compared to the unbound form. These observations were further supported by MM/PBSA binding free-energy calculations. Collectively, these findings support the designed construct as a computationally validated multi-strain dengue antigen candidate. However, extensive experimental validation will be required to establish its suitability for diagnostic or vaccine-related applications, which will be addressed in future studies.

Reference sequences of all four dengue virus serotypes were retrieved from NCBI. Dengue-specific B-cell epitopes were identified from IEDB. Immunogenicity was simulated using C-IMMSIM. Secondary and tertiary structures were predicted with PSIPRED and Phyre2. Protein-protein docking with the monoclonal antibody Fab 1A1D-2 (PDB 2R29) was performed on the HDOCK server. Molecular dynamics simulations totalling 1200 ns; four 300 ns replicate simulations of antigen-antibody complex were carried out in GROMACS 2023.8 using the CHARMM27 force field, TIP3P water model, and Particle Mesh Ewald electrostatics. Analyses were performed using GROMACS utilities, and MM/PBSA binding free energy calculations were conducted with gmx_MMPBSA to evaluate antigen-antibody complex stability.