The Interplay of Furin Cleavage and D614G in Modulating SARS-CoV-2 Spike Protein Dynamics

Summary We report a detailed analysis of the full-length SARS-CoV-2 spike dynamics within a native-like membrane environment and variants inaccessible to studies on soluble constructs by conducting hydrogen-deuterium exchange mass spectrometry (HDX-MS) on enveloped virus-like particles (eVLPs) displaying various spike constructs. We find that the previously identified open-interface trimer conformation is sampled in all eVLP-displayed spike variants studied including sequences from engineered vaccine constructs and native viral sequences. The D614G mutation, which arose early in the pandemic, favors the canonical ‘closed-interface’ prefusion conformation, potentially mitigating premature S1 shedding in the presence of a cleaved furin site and providing an evolutionary advantage to the virus. Remarkably, furin cleavage at the S1/S2 boundary allosterically increases the flexibility of the S2’ site, which may facilitate increased TMPRSS2 processing, enhancing viral infectivity. The use of eVLPs in HDX-MS studies provides a powerful platform for studying viral and membrane proteins in near-native environments. Full Text Availability The license terms selected by the author(s) for this preprint version do not permit archiving in PMC. The full text is available from the preprint server.