Quantitative profiling of N-glycosylation of SARS-CoV-2 spike protein variants.

With the global spread of the corona virus disease-2019 pandemic, new spike variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continuously emerge due to increased possibility of virus adaptive amino acid mutations. However, the N-glycosylation profiles of different spike variants are yet to be explored extensively, although the spike protein is heavily glycosylated and surface glycans are well-established to play key roles in viral infection and immune response. Here, we investigated quantitatively the N-glycosylation profiles of seven major emerging spike variants including Original, Alpha (B.1.1.7), Beta (B.1.351), Gamma (P.1), Kappa (B.1.671.1), Delta (B.1.671.2), and Omicron (B.1.1.529). The aim was to understand the changing pattern of N-glycan profiles in SARS-CoV-2 evolution in addition to the widely studied amino acid mutations. Different spike variants exhibit substantial variations in the relative abundance of different glycan peaks and subclasses, although no specific glycan species are exclusively present in or absent from any specific variant. Cluster analysis shows that the N-glycosylation profiles may hold the potential for SARS-CoV-2 spike variants classification. Alpha and Beta variants exhibit the closest similarity to the Original, and the Delta variant displays substantial similarity to Gamma and Kappa variants, while the Omicron variant is significantly different from its counterparts. We demonstrated that there is a quantifiable difference in N-glycosylation profiles among different spike variants. The current study and observations herein provide a valuable framework for quantitative N-glycosylation profiling of new emerging viral variants and give us a more comprehensive picture of COVID-19 evolution.