New Publication in the journal Viruses

New publication in the journal Viruses: Site-Specific O-Glycosylation Analysis of SARS-CoV-2 Spike Protein Produced in Insect and Human Cells

Authors: Ieva Bagdonaite, Andrew J. Thompson, Xiaoning Wang, Max Søgaard, Cyrielle Fougeroux, Martin Frank, Jolene K. Diedrich, John R. Yates III, Ali Salanti, Sergey Y. Vakhrushev, James C. Paulson, and Hans H. Wandall 

Abstract:
Enveloped viruses hijack not only the host translation processes, but also its glycosylation machinery, and to a variable extent cover viral surface proteins with tolerogenic host-like structures. SARS-CoV-2 surface protein S presents as a trimer on the viral surface and is covered by a dense shield of N-linked glycans, and a few O-glycosites have been reported. The location of O-glycans is controlled by a large family of initiating enzymes with variable expression in cells and tissues and hence is difficult to predict. Here, we used our well-established O-glycoproteomic workflows to map the precise positions of O-linked glycosylation sites on three different entities of protein S—insect cell or human cell-produced ectodomains, or insect cell derived receptor binding domain (RBD). In total 25 O-glycosites were identified, with similar patterns in the two ectodomains of different cell origin, and a distinct pattern of the monomeric RBD. Strikingly, 16 out of 25 O-glycosites were located within three amino acids from known N-glycosides. However, O-glycosylation was primarily found on peptides that were unoccupied by N-glycans, and otherwise had low overall occupancy. This suggests possible complementary functions of O-glycans in immune shielding and negligible effects of O-glycosylation on subunit vaccine design for SARS-CoV-2.