Heterogeneous binding of SARS-CoV2 fusion peptide on complex cellular membranes enhances its fusogenicity

There is a large class of enveloped viruses that utilize sophisticated fusion mechanisms as a precursor to enter host cells. In SARS-Cov-2 the S2 domain of the S protein contains the fusion peptide (FP), which is believed to be central to the inter-membrane fusion machinery. However, the microscopic parameters that drive enhanced fusogenicity of the SARS-CoV-2 FP on realistic complex cellular membranes, leading to the observed virulence and fatality due to SARS-CoV-2 infection, remain unclear. In this report, we identify the correlation between SARS-CoV-2 FP conformational and multiphase cellular membrane dynamical heterogeneity, using existing and new membranotropic parameters, that drives enhanced SARS-CoV-2 FP fusogenicity. Combining high-resolution fluorescence microscopy and time-domain spectroscopy along with atomic molecular dynamics simulations, we demonstrate significantly enhanced membranotropy of SARS-CoV-2 FP in phase-separated model host cell membranes compared with their homogeneous counterparts. We observe dynamic phase homogenization and strongly correlated peptide-lipid diffusion, which correlates with the broader spectrum of interactions of SARS-CoV-2 FP with both the L_o and L_d phases in the multiphase complex model cellular membranes. Significantly, we correlate SARS-CoV-2 binding heterogeneity with lipid-mixing data to demonstrate how the FP's conformational binding landscape modulates key fusogenic parameters such as membrane fluidity and dehydration, leading to enhanced macroscopic fusion. Our findings offer a mechanistic framework that extends existing paradigms of viral FP activity to heterogeneous membrane environments, potentially informing the development of broadly acting antiviral strategies targeting the fusion machinery.