ABSTRACT
SARS-CoV-2 spike glycoprotein mediates receptor binding and subsequent membrane fusion. It exists in a range of conformations, including a closed state unable to bind the ACE2 receptor, and an open state that does so but displays more exposed antigenic surface. Spikes of variants of concern (VOCs) acquired amino acid changes linked to increased virulence and immune evasion. Here, using HDX-MS, we identified changes in spike dynamics that we associate with the transition from closed to open conformation, to ACE2 binding, and to specific mutations in VOCs. We show that the RBD-associated subdomain plays a role in spike opening, whereas the NTD acts as a hotspot of conformational divergence of VOC spikes driving immune evasion. Alpha, beta and delta spikes assume predominantly open conformations and ACE2 binding increases the dynamics of their core helices, priming spikes for fusion. Conversely, substitutions in omicron spike lead to predominantly closed conformations, presumably enabling it to escape antibodies. At the same time, its core helices show characteristics of being pre-primed for fusion even in the absence of ACE2. These data inform on SARS-CoV-2 evolution and omicron emergence.