1.G.18 The SARS-CoV Fusion Peptide in the Spike Glycoprotein Precursor (SARS-FP) Family

The severe acute respiratory syndrom (SARS) coronavirus (CoV) fusion peptide is within the spike glycoprotein precursor.  The fusion peptide is a C-terminal 19 aas peptide, in the spike glycoprotein precursor of 1255 aas (Apellániz et al. 2014). Members of this family have a central CDD/Pfam Spike_rec_bind domain and a large C-terminal Corona_S2 domain. 120 references have been published on this and related proteins as of 3/2020. Generally, a stretch of 20-25 amino acids located at the N-terminus of the fusion protein, known as a fusion peptide, plays a decisive role in the fusion process. The stalk model of membrane fusion postulated a common route of bilayer transformation for stalk, transmembrane contact, and pore formation, and the fusion peptide is believed to facilitate bilayer transformation to promote membrane fusion (Meher and Chakraborty 2021). Single-domain antibodies (nanobodies) bind tightly to Spike and neutralize SARS-Co/V-2 (Schoof et al. 2020; Xiang et al. 2020. A cell-based system combined with flow cytometry has been used to evaluate antibody responses against SARS-CoV-2 transmembrane proteins in patients with COVID-19 (Martin et al. 2022).

The coronavirus spike protein (S) forms the distinctive virion surface structures that are characteristic of this viral family, appearing in negatively stained electron microscopy as stems capped with spherical bulbs. These structures are essential for the initiation of infection through attachment of the virus to cellular receptors followed by fusion to host cell membranes (Ye et al. 2004). The S protein can also mediate the formation of syncytia in infected cells. It is a large type I (1 TMS) protein, and all except a short carboxy-terminal segment of S constitutes the ectodomain. For the prototype coronavirus mouse hepatitis virus (MHV), S protein assembly into virions is specified by the carboxy-terminal segment, which comprises the transmembrane domain and the endodomain. Ye et al. 2004 genetically dissected these domains in the MHV S protein to localize the determinants of S incorporation into virions. Assembly competence maps to the endodomain of S, which is sufficient to target the protein for incorporation into the virion. The charge-rich carboxy-terminal region of the endodomain plays a major role. The adjacent cysteine-rich region of the endodomain is critical for fusion.

Wrapp et al. 2020 determined a 3.5-Å-resolution cryo-EM structure of the 2019-nCoV S trimer in the prefusion conformation. The predominant state of the trimer has one of the three receptor-binding domains (RBDs) rotated up in a receptor-accessible conformation. They also provided biophysical and structural evidence that the 2019-nCoV S protein binds angiotensin-converting enzyme 2 (ACE2) with higher affinity than does severe acute respiratory syndrome (SARS)-CoV S. They tested several published SARS-CoV RBD-specific monoclonal antibodies and found that they did not have appreciable binding to 2019-nCoV S (Wrapp et al. 2020). By using cryo-electron tomography, Tai et al. 2021 observed both prefusion and postfusion spikes in beta-propiolactone-inactivated SARS-CoV-2 virions and solved the in situ structure of the postfusion spike at nanometer resolution. Compared to previous reports, the six-helix bundle fusion core, the glycosylation sites, and the location of the transmembrane domain were clearly resolved. They observed oligomerization patterns of the spikes on the viral membrane, likely suggesting a mechanism of fusion pore formation (Tai et al. 2021).