Host Cell Glycocalyx Remodeling Reveals SARS-CoV-2 Spike Protein Glycomic Binding Sites

Abstract

Glycans on the host cell membrane and viral proteins play critical roles in pathogenesis. Highly glycosylated epithelial cells represent the primary boundary separating embedded host tissues from pathogens within the respiratory and intestinal tracts. SARS-CoV-2, the causative agent for the COVID-19 pandemic, reaches into the respiratory tract. We found purified human milk oligosaccharides (HMOs) inhibited the viral binding on cells. Spike (S) protein receptor binding domain (RBD) binding to host cells were partly blocked by co-incubation with exogenous HMOs, most by 2-6-sialyl-lactose (6′SL), supporting the notion that HMOs can function as decoys in defense against SARS-Cov2. To investigate the effect of host cell glycocalyx on viral adherence, we metabolically modified and confirmed with glycomic methods the cell surface glycome to enrich specific N-glycan types including those containing sialic acids, fucose, mannose, and terminal galactose. Additionally, Immunofluorescence studies demonstrated that the S protein preferentially binds to terminal sialic acids with α-(2,6)-linkages. Furthermore, site-specific glycosylation of S protein RBD and its human receptor ACE2 were characterized using LC-MS/MS. We then performed molecular dynamics calculations on the interaction complex to further explore the interactive complex between ACE2 and the S protein. The results showed that hydrogen bonds mediated the interactions between ACE2 glycans and S protein with desialylated glycans forming significantly fewer hydrogen bonds. These results supported a mechanism where the virus binds initially to glycans on host cells preferring α-(2,6)-sialic acids and finds ACE2 and with the proper orientation infects the cell.

Proteomics‐based mass spectrometry profiling of SARS‐CoV‐2 infection from human nasopharyngeal samples

“…Interestingly, some studies have indicated that altering the host glycosylation can effectively inhibit viral binding to the host. Particularly, human milk oligosaccharides that are heavily decorated with α−2,6‐sialyl‐lactose are capable of preventing SARS‐CoV‐2 infection (Moore et al, 2021 ; Sheng et al, 2022 ). In‐depth characterization of the viral glycoproteome is therefore vital to understand the roles of SARS‐CoV‐2 in host‐pathogen mechanisms.…”

Absolute Affinities from Quantitative Shotgun Glycomics Using Concentration-Independent (COIN) Native Mass Spectrometry

“…According to the pseudotyped lentivirus and authentic virus infection data, acidic host N -glycans may serve as viral attachment factors. Contrary to previously reported findings, oligomannose N -glycans are found not to be RBD ligands . Finally, the screening data provide irrefutable evidence that SARS-CoV-2 RBD recognizes sialoglycans.…”

“…63 Based on this finding and the observation that RBD binding to host cells is reduced upon incubation with oligomannose glycans, it was concluded that cell surface oligomannose glycans increase adherence of RBD. 63 However, in the current study, as well as previous screening studies, oligomannose N-glycans are not detected as RBD ligands. 57 To gain further insight into the host cell Nglycans on SARS-CoV-2 infection, we performed pseudotyped lentivirus infection of ACE2-expressing HEK293T cells.…”

Integrating Computational Methods in Network Pharmacology and In Silico Screening to Uncover Multi-targeting Phytochemicals against Aberrant Protein Glycosylation in Lung Cancer

“…Targeting glycoproteins is an underexploited strategy for the development of cancer therapeutics. Carbohydrate-active enzymes such as glycosidases and glycosyltransferases are attractive drug targets due to their involvement in the biosynthesis of glycan structures. , Relatively few studies have investigated potential small molecule inhibitors with drug-like properties against these enzymes. − These include plant alkaloids that inhibit glycosidases by inducing inhibition of trimming reactions after the attachment of Glc 3 Man 9 GlcNAc 2 oligosaccharide. An example of a plant alkaloid is swainsonine from an Astralagus species known as locoweeds.…”