The role of conformational dynamics of enveloped virus spike proteins in cell entry

Research Abstract: The spike/fusion glycoproteins residing on the surface of human enveloped viruses are targets of immune response and are the focus of researchers developing vaccines and antiviral treatments. The conformational dynamics of spike proteins drive the entry of enveloped viruses into cells via viral membrane fusion and facilitate antibody recognition. However, the lack of deep insights into dynamics has prevented a complete elucidation of the molecular mechanism by which spike proteins promote virus entry. Many viruses, such as coronavirus SARS-CoV-2, respiratory syncytial virus (RSV), and HIV-1, share a similar viral fusion mechanism (type-I) mediated by their respective spike proteins. These spikes undergo dramatic structural changes, and the energy released from conformational transitions overcomes the fusion kinetic barriers. However, our understanding of the multi-step fusion process mainly relies on individual structural snapshots of spike proteins at fusion endpoints. How these endpoints are correlated in a time-resolved manner and the order and frequency of conformational events underlying virus entry remain largely elusive. The proposed research extends our efforts to explore viral membrane fusion and is supported by our experience probing the conformational dynamics of the SARS-CoV-2 spike (S) and HIV-1 envelope (Env) glycoproteins. The overarching goal of this project is to integrate our knowledge into a generic working model that will describe a time-resolved stepwise framework of the type-I fusion mechanism, in which the conformational trajectories of fusion proteins are explicitly defined in space and time. We pioneered the use of the single-molecule Förster resonance energy transfer (smFRET) to study S and revealed multiple S conformations on the virus. We delineated sequential transitions of S from closed to open conformations upon activation by cellular receptors. We provided the first experimental evidence of decelerated transition dynamics from open states, suggesting increased stability of the fusion-reactive open state to be part of the SARS-CoV-2 adaption strategies. Here, we will use an integrated platform of smFRET and virus-to-cell fusion in combination with computational and structural tools to reveal the conformational plasticity S adapts during virus evolution and to visualize the conformational trajectory S undergoes during fusion. We will perform comparative studies on another respiratory virus - RSV fusion (F) protein, with interest in other type-I spike proteins of newly emerging viruses. We will elucidate conformational events and transition dynamics of F-mediated viral membrane fusion and evaluate whether conformation- presentation of F-based vaccine candidates represents the predominant state exposed to the host. The studies are expected to allow us to identify the common and divergent traits of the S- and F-mediated fusion processes that will advance our knowledge and help us define the common theme of the type-I fusion mechanism. We envision that this program of research using different advanced technologies will reveal unrecognized insights into virus entry that lay the foundation for advances in anti-viral interventions, in line with the mission of NIGMS.