Fig. 4: Sialic acids present on ACE2 precluded perfect spike/ACE2-interaction.

a Schematic of the N-linked and O-linked ACE2 glycosylation mutants. b Western blot of ACE2 glycosylation mutants overexpressed in BHK21 cells. c BHK21 cells were transfected with the ACE2 mutants for 24 h before inoculating the cells with SARS-CoV-2-S-pseudovirus. Pseudovirus entry was determined at 24 h post inoculation (n = 3). d Entry of VSV-G-pseudovirus in BHK21 cells transfected with the ACE2 mutants (n = 3). e BHK21 cells expressing the ACE2 mutants were infected with SARS-CoV-2. Virus replication in the cell lysate samples at 24hpi was evaluated with qRT-PCR (n = 8 for EV, n = 12 for del N + O, n = 14 for other groups). f Virus replication in the supernatant samples at 24hpi was evaluated with qRT-PCR (n = 8 for EV, n = 12 for N690Q, n = 14 for other groups). g The binding affinity between SARS-CoV-2-S RBD and human ACE2 was determined with SPR. h The binding affinity between SARS-CoV-2-S RBD and NA-pretreated human ACE2 was determined with SPR. Data represented mean and standard deviations from the indicated number of biological repeats. The experiments in b, g, and h were repeated three times independently with similar results. Statistical significance between groups was determined with one way-ANOVA. * represented p < 0.05 and ** represented p < 0.01. ns not significant, EV empty vector. Source data are provided as a Source Data file.