SARS-CoV-2 Spike DMS

We developed a Deep Mutational Scanning (DMS) platform to investigate SARS-CoV-2 mutations and their impact on immune evasion and infectivity. Our platform focuses on the spike protein and utilizes an inverted infection assay to measure spike expression, ACE2 affinity, and viral infectivity in human cells. Surprisingly, our analysis reveals that spike protein expression, rather than ACE2 affinity, is the primary factor affecting viral infectivity. Notably, within the N-terminal domain (NTD), spike expression and infectivity-enhancing mutations are concentrated in flexible loops. We also observed that Omicron variants BA.1 and BA.2 exhibit immune evasion through receptor binding domain (RBD) mutations, although these mutations reduce structural stability. Interestingly, the NTD has evolved increased stability, compensating for RBD instability and resulting in heightened overall infectivity. Our findings, available in SpikeScanDB, emphasize the importance of spike expression levels and compensatory mutations in both the NTD and RBD domains for shaping Omicron variant infectivity.