INTERACTION OF SARS-COV-2 FUSION PEPTIDES WITH MICELLAR MEMBRANE MODELS: INSIGHTS FROM SPECTROSCOPIC AND COMPUTATIONAL APPROACHES

Coronavirus membrane fusion is mediated by the Spike glycoprotein and involves membrane-active regions known as fusion peptides. Understanding how these peptides interact with lipid membranes is essential for elucidating the molecular mechanisms underlying viral membrane fusion. Here, we investigated the interaction of two SARS-CoV and SARS-CoV-2 fusion peptides, the upstream fusion peptide (FP_u) and internal fusion peptide (IFP), with zwitterionic lysophosphatidylcholine (LPC) and anionic lysophosphatidylglycerol (LPG) micelles at pH 5.0 and 7.4. Steady-state and time-resolved fluorescence spectroscopy, circular dichroism (CD), electron spin resonance (ESR), and molecular dynamics (MD) simulations were combined to characterize peptide partitioning, conformational organization, and effects on membrane dynamics. Both peptides displayed moderate interactions with lipid micelles, with IFP exhibiting higher partition coefficients than FP_u under all conditions and a preferential association with negatively charged LPG. Fluorescence measurements revealed membrane- and pH-dependent changes in the local environment of the peptide fluorophores, with longer average fluorescence lifetimes generally observed in LPC than in LPG micelles. CD spectroscopy demonstrated that both peptides adopted ordered secondary structures in membrane-mimetic environments. FP_u exhibited a mixture of α-helical and β-strand structures, whereas IFP showed a predominantly α-helical conformation. ESR analysis revealed that both peptides restricted acyl-chain mobility and increased lipid ordering within the micelles. These effects were detected at different depths of the hydrophobic core and were generally more pronounced in LPG, particularly under acidic conditions. MD simulations further showed that both peptides penetrated lyso-myristoyl phosphatidylglycerol (LMPG) and dodecylphosphocholine (DPC) micelles and maintained organized secondary structures during membrane association, supporting the experimental observations. Overall, our results demonstrate that lipid composition and pH modulate the membrane interaction, structural organization, and membrane-ordering effects of SARS-CoV and SARS-CoV-2 fusion peptides. The stronger membrane association of IFP, together with its preferential interaction with anionic lipids and predominantly α-helical structure, highlights the importance of membrane composition in regulating fusion-peptide behavior and provides further insight into the molecular determinants of coronavirus membrane engagement. Together, these findings indicate that lipid composition and pH can enhance peptide-membrane association while altering lipid dynamics, suggesting that peptide sequence and membrane properties contribute to the conformations required for membrane perturbation and fusion.