Comprehensive Analysis of the Glycan Complement of SARS-CoV‑2 Spike Proteins Using Signature Ions-Triggered Electron-Transfer/Higher-Energy Collisional Dissociation (EThcD) Mass Spectrometry

Severe acute respiratory syndrome
coronavirus 2 (SARS-CoV-2) has
led to a global pandemic of coronavirus disease 2019 (COVID-19). The
spike protein expressed on the surface of this virus is highly glycosylated
and plays an essential role during the process of infection. We conducted
a comprehensive mass spectrometric analysis of the N-glycosylation
profiles of the SARS-CoV-2 spike proteins using signature ions-triggered
electron-transfer/higher-energy collision dissociation (EThcD) mass
spectrometry. The patterns of N-glycosylation within the recombinant
ectodomain and S1 subunit of the SARS-CoV-2 spike protein were characterized
using this approach. Significant variations were observed in the distribution
of glycan types as well as the specific individual glycans on the
modification sites of the ectodomain and subunit proteins. The relative
abundance of sialylated glycans in the S1 subunit compared to the
full-length protein could indicate differences in the global structure
and function of these two species. In addition, we compared N-glycan
profiles of the recombinant spike proteins produced from different
expression systems, including human embryonic kidney (HEK 293) cells
and Spodoptera frugiperda (SF9) insect
cells. These results provide useful information for the study of the
interactions of SARS-CoV-2 viral proteins and for the development
of effective vaccines and therapeutics.