BPS2026 – Prediction and characterization of HIV-1 fusion peptide variants that modulate viral entry via conformational plasticity

HIV-1 infection remains a major global health challenge with more than 40 million people living with HIV and over one million new cases reported in 2024. Although antiretroviral therapy suppresses viral replication, it cannot eradicate latent reservoirs, underscoring the need for deeper mechanistic understanding of the virus to aid development of curative strategies. HIV-1 viral entry is mediated by the envelope glycoprotein (Env), which assembles as a trimer on the virus surface. Each Env subunit comprises gp120, the receptor-binding protein, and gp41, the fusion protein. Following receptor/coreceptor engagement and activation by gp120, gp41 undergoes a conformational change that drives membrane fusion and viral entry. One key step involves insertion of short N-terminal fusion peptides (FPs) from gp41 into the host membrane. These FPs exhibit a conformational plasticity, adopting either β-sheet or α-helical structures depending on membrane cholesterol content. This plasticity of FPs is essential for viral entry but remains poorly understood. Here, we combine molecular dynamics simulations, machine learning, and spectroscopy to dissect the molecular drivers of gp41 FP conformational flexibility. We perform extensive simulations of the FP in both α-helical and β-sheet states and apply statistical analysis to identify key residues that modulate conformational switching. These residues were further analyzed using an embedding-space representation of all combinatorial mutants to identify variants that are stable and compatible with viral infection. The mutations predicted to influence conformational switching were first probed through simulations and will subsequently be characterized by experimental validation. Our integrated framework reveals specific mutational changes that govern HIV-1 FP conformational switching and differentiates these effects from cholesterol-driven plasticity. These studies provide new insights into viral entry mechanisms and raise the question of how intrinsic conformational plasticity and cholesterol-induced plasticity interplay during HIV-1 entry.