SARS-CoV-2 VARIANTS INCREASE KINETIC STABILITY of OPEN SPIKE CONFORMATIONS

ABSTRACT

Background: SARS-CoV-2 variants of concern harbor mutations in the Spike (S) glycoprotein that confer more efficient transmission and dampen the efficacy of COVID-19 vaccines and antibody therapies. S mediates virus entry and is the primary target for antibody responses, with structural studies of soluble S variants revealing an increased propensity towards conformations accessible to the human Angiotensin-Converting Enzyme 2 (hACE2) receptor. However, real-time observations of conformational dynamics that govern the structural equilibriums of the S variants have been lacking. Methods: Here, we report single-molecule Förster Resonance Energy Transfer (smFRET) studies of S variants of concern containing critical mutations, including D614G and E484K, in the context of virus particles. Results: Investigated variants were shown by smFRET to predominantly occupy more open hACE2-accessible conformations, agreeing with predictions from structures of soluble trimers. Additionally, S variants exhibited decelerated transitions from hACE2-accessible/bound states. Conclusion: Here, we provide the real-time dimension to distinct structures of Spikes in the context of virus particles and present the first experimental evidence of increased stability of Spike variants. Our finding of increased S kinetic stability in the open conformation provides a new perspective on SARS-CoV-2 adaptation to the human population.