6.2. SARS-CoV-2 Recognizes 9-O-Acetyl-SAs and MERS-CoV Recognizes α2,3-SAs as Attachment Receptors
The S glycoprotein SARS-CoV-2 initiates infection of the host cells. The molecular basis of CoV attachment to sugar/glycan receptors is an important issue, as demonstrated by recent cryo-EM defining the structure of the CoV-OC43 S glycoprotein trimer complexed with a 9-O-acetylated SA [56]. Cryo-EM structures of the trimeric ectodomain of S glycoprotein were observed using forms complexed with Neu5Ac, Neu5Gc, sialyl–LewisX (SLeX), α2,3-sialyl-N-acetyl-lactosamine (α2,3-SLacNAc) and α2,6-SLacNAc, respectively. The receptor-binding site is commonly conserved in all CoV S glycoproteins, which attach to 9-O-Ac-SA species with similar ligand-binding pockets to the CoV HEs and influenza virus C/D HEF glycoproteins, indicating conserved recognizing structures [25]. The S glycoprotein-9-O-acetyl-SA interaction resembles the ligand-binding pockets of CoV HEs and influenza virus C/D HE fusion glycoproteins. HCoV-OC43 and BCoV recognize 9-O-Ac-SA. S glycoproteins engage 9-O-acetyl-SAs. The 9-O-acetyl SAs are the binding site for HCoV-OC43 S glycoprotein and related β-1 CoV S glycoproteins, however SA-binding sites on the 9-O-acetyl sialyl receptors of MERS-CoV S glycoprotein and HCoV-OC43 S glycoprotein are different [71]. Thus, CoVs use two different entry and attachment receptors. Therefore, S glycoproteins of CoVs are distinct from influenza virus A HAs, which bind to the Neu5Ac species by conserved binding sites. The ligand-binding sites of BCoV HE enzyme, influenza HEF enzyme and CoV S glycoprotein have evolved 9-O-Ac-SA binding through hydrogen bonding with the 9-O-acetyl carbonyl group and hydrophobic pocket formation with the 9-O-acetyl methyl group [71,72]. However, influenza HA cannot bind to 9-O-acetyl-SAs but can bind to NeuGcs [73]. The HCoV-OC43 S glycoprotein, HCoV-HKU1 S glycoprotein, BCoV S glycoprotein and PHEV S glycoprotein, therefore, share the ligand-binding specificity of influenza C/D HEF enzyme, although they are functionally more similar to influenza virus A/B HA, whereas CoV HE or influenza virus A/B NA have RDE activities
CoV HEs are functionally similar to influenza virus C/D HEF glycoproteins. In CoV, the S glycoprotein recognizes the 9-O-Ac-SA sugar, while the HE acts as the RDE enzyme with SA-O-acetyl-esterase activity to release virions from infected host cells. For example, HCoV-OC43 also has a similar HE as an RDE [71]. In influenza C and D viruses, HEF glycoproteins act similarly to the CoV HE [74]. In influenza A virus, RDE NA releases virions from host cells. However, MERS-CoV does not have a similar enzyme and thus MER-CoV binding to SA receptors is mediated by energetically reversible interactions of the lipid rafts with increased SA receptors [75], thus enhancing dipeptidyl peptidase 4 (DPP4) or carcinoembryonic antigen-related cell adhesion molecule 5 (CEACAM5) recognition power and viral entry [76] and membrane-associated 78-kDa glucose-regulated protein (GRP78) [77].
MERS-CoV S glycoprotein can hemagglutinate human erythrocytes and mediates virus entry into human respiratory epithelial cells. MERS-CoV S glycoprotein attachment is not observed for 9-O-acetylated or 5-N-glycolyl SAs, but is observed for α2,3-SA linkage over α2,6-SA linkages. SA-binding sites of MERS-CoV S glycoprotein and HCoV-OC43 S glycoprotein are not conserved [78], although they engage α2,3-SAs on the avian host cell surface [79]. MERS-CoV recognizes α2,3-SA and to a lesser extent the α2,6-SAs and sulfated SLeX for binding preference. Thus, S glycoproteins may have independently evolved SA recognition. The acquisition of SA-binding ability of MERS-CoV S seems to be an evolutionarily recent event, because HKU4 S1 and HKU5 S1 cannot hemagglutinate human erythrocytes [75], indicating flexible evolutionary exchange allowing cross-species transmission towards host cell tropism of CoVs. In conclusion, CoV recognition of 9-O-Ac-SAs for infection is based on a conserved sequence for engagement of SA-related carbohydrate ligands across CoVs and orthomyxoviruses.