Computational Insights into Mechanostability and Dissociation Dynamics of the Dengue Virus Envelope Protein Ectodomain Dimer Across pH and Temperature Gradients

Abstract

Dengue virus (DENV) is an enveloped flavivirus of major public health importance. Its envelope (E) protein mediates viral entry through homodimer dissociation and subsequent membrane fusion, making it one of the principal targets for antiviral strategies. To investigate the molecular determinants of this process, we performed steered molecular dynamics (SMD) simulations of the E protein ectodomain (ecE) dimer from DENV-2 and DENV-3 under variable pH and temperature conditions. Force-extension analyses revealed a highly stable interface for both serotypes, with rupture forces exceeding 1000 pN. pH and temperature had modest effects on overall mechanical resistance. However, DENV-3 displayed a distinct sensitivity to thermal stress compared to DENV-2. We identified a robust, asymmetric dissociation pathway across all conditions, characterized by a metastable intermediate state involving partial dimer opening and exposure of the fusion loop (FL). This intermediate exposes an immunodominant epitope and persists under physiological conditions, suggesting it as a viable target for therapeutic intervention. The primary contributions to the interfacial interaction network were found to arise from van der Waals interactions, followed by hydrogen bonds, salt bridges, and pi-cation interactions. DENV-3 exhibited a slightly greater contribution from polar interactions involving domains EDI, EDII, and EDIII. Furthermore, pairwise occupancy analysis identified pH-sensitive contacts that are disrupted under acidic conditions, particularly in DENV-3, providing mechanistic insight into the early stages of ecE dissociation. Together, these findings provide structural insights into the dissociation mechanism, identifying key metastable states and pH-sensitive interactions that could be exploited to develop antivirals that stabilize the dimer and prevent viral fusion.