Unveiling the Role of ZIKV NS4A Mutants F4L and E8D through Molecular Docking and Dynamics Simulation: Implications for MAVS-Mediated Immune Evasion

Zika virus (ZIKV), a mosquito-borne flavivirus, has emerged as a global health concern due to its association with congenital microcephaly and neurological disorders. The non-structural protein NS4A plays a pivotal role in viral replication and immune evasion by antagonizing the mitochondrial antiviral signaling protein (MAVS). In this study four NS4A mutations (L48M, K42E, F4L, and E8D) were originally evaluated using structural stability and interaction analyses; however, only F4L and E8D showed destabilizing effects that required further examination. We used molecular docking, 100 ns molecular dynamics simulations, and binding free energy calculations to assess their effects on NS4A-MAVS binding. Stability investigations (RMSD, RMSF, and Rg) revealed that both mutations changed the conformational dynamics of NS4A-MAVS complexes, with F4L displaying transitory fluctuations and E8D exhibiting long-term structural flexibility. Hydrogen bond research revealed that both mutants had stronger interaction networks with MAVS compared to the natural type. MM/PBSA computations showed that F4L and E8D had higher binding affinities, with ΔG values of -54.05 kcal/mol and -56.25 kcal/mol, respectively, compared to -61.73 kcal/mol in the wild type. The stronger electrostatic contributions observed in the E8D complex highlight its potential to further disrupt MAVS-mediated interferon induction. Collectively, these results suggest that the F4L and particularly E8D mutations enhance the immune-evasive capacity of ZIKV by stabilizing NS4A–MAVS interactions, offering insights into viral pathogenesis and providing a computational basis for therapeutic targeting of NS4A.