PT  - JOURNAL ARTICLE
AU  - Zhang, Shijian
AU  - Go, Eden P.
AU  - Ding, Haitao
AU  - Anang, Saumya
AU  - Kappes, John C.
AU  - Desaire, Heather
AU  - Sodroski, Joseph
TI  - Analysis of glycosylation and disulfide bonding of wild-type SARS-CoV-2 spike glycoprotein
AID  - 10.1101/2021.04.01.438120
DP  - 2021 Jan 01
TA  - bioRxiv
PG  - 2021.04.01.438120
4099  - http://biorxiv.org/content/early/2021/04/01/2021.04.01.438120.short
4100  - http://biorxiv.org/content/early/2021/04/01/2021.04.01.438120.full
AB  - The SARS-CoV-2 coronavirus, the etiologic agent of COVID-19, uses its spike (S) glycoprotein anchored in the viral membrane to enter host cells. The S glycoprotein is the major target for neutralizing antibodies elicited by natural infection and by vaccines. Approximately 35% of the SARS-CoV-2 S glycoprotein consists of carbohydrate, which can influence virus infectivity and susceptibility to antibody inhibition. We found that virus-like particles produced by coexpression of SARS-CoV-2 S, M, E and N proteins contained spike glycoproteins that were extensively modified by complex carbohydrates. We used a fucose-selective lectin to enrich the Golgi-resident fraction of a wild-type SARS-CoV-2 S glycoprotein trimer, and determined its glycosylation and disulfide bond profile. Compared with soluble or solubilized S glycoproteins modified to prevent proteolytic cleavage and to retain a prefusion conformation, more of the wild-type S glycoprotein N-linked glycans are processed to complex forms. Even Asn 234, a significant percentage of which is decorated by high-mannose glycans on soluble and virion S trimers, is predominantly modified in the Golgi by processed glycans. Three incompletely occupied sites of O-linked glycosylation were detected. Viruses pseudotyped with natural variants of the serine/threonine residues implicated in O-linked glycosylation were generally infectious and exhibited sensitivity to neutralization by soluble ACE2 and convalescent antisera comparable to that of the wild-type virus. Unlike other natural cysteine variants, a Cys15Phe (C15F) mutant retained partial, but unstable, infectivity. These findings enhance our understanding of the Golgi processing of the native SARS-CoV-2 S glycoprotein carbohydrates and could assist the design of interventions.