Domains and Functions of Spike Protein in Sars-Cov-2 in the Context of Vaccine Design

Abstract

The spike protein in SARS-CoV-2 (SARS-2-S) interacts with the human ACE2 receptor to gain entry into a cell to initiate infection. Both Pfizer/BioNTech's BNT162b2 and Moderna's mRNA-1273 vaccine candidates are based on stabilized mRNA encoding prefusion SARS-2-S that can be produced after the mRNA is delivered into the human cell and translated. SARS-2-S is cleaved into S1 and S2 subunits, with S1 serving the function of receptor-binding and S2 serving the function of membrane fusion. Here, I dissect in detail the various domains of SARS-2-S and their functions discovered through a variety of different experimental and theoretical approaches to build a foundation for a comprehensive mechanistic understanding of how SARS-2-S works to achieve its function of mediating cell entry and subsequent cell-to-cell transmission. The integration of structure and function of SARS-2-S in this review should enhance our understanding of the dynamic processes involving receptor binding, multiple cleavage events, membrane fusion, viral entry, as well as the emergence of new viral variants. I highlighted the relevance of structural domains and dynamics to vaccine development, and discussed reasons for the spike protein to be frequently featured in the conspiracy theory claiming that SARS-CoV-2 is artificially created.

Keywords: COVID-19; S-2P; SARS-CoV-2; cleavage; hydrophobicity; isoelectric point; protein structure; spike protein; vaccine.

Figure 1

Domain structure of SARS-S and SARS-2-S. (A) Key domains in SARS-S and SARS-2-S. SP, signal peptide; NTD, N-terminal domain; RBD, receptor-binding domain; FP, fusion peptide; IFP, internal fusion peptide; HR, heptad repeats; TM, transmembrane domain; CT, cytoplasmic tail. The top and bottom numbers in each domain pertain to SARS-S and SARS-2-S, respectively. The red arrows indicate cleavage sites, and their numbers pertain to SARS-2-S; (B) Alignment of SP between SARS-S (top) and SARS-2-S (bottom); (C,D) Alignment of two inter-domain segments; (E) HR1 in SARS-S and SARS-2-S, together with the top view of a helix showing hydrophobic positions a and d on the same side; (F) Hydrophobicity plot generated from DAMBE [16].

Figure 2

Cleavage sites at the S1/S2 boundary. (A) An insertion of 12 nt in SARS-CoV-2 results in a new polybasic furin cleavage site, resulting in two cleavage sites indicated by the red downward arrows. “*” indicates sites that are identical among the six viral strains. Numbers follow (B) Schematic domain structure of S protein, with the same abbreviation as in Figure 1A; (C) Tissue-specific mRNA distribution of human trypsin-like protease TMPRESS11D and FURIN, derived from [30]; (D) Cleavage site for splitting S2 into FP and S2′ domains.

Figure 3

Hydrophobicity (A) and protein isoelectric point (B) plots of spike protein from SARS-CoV-2 and its close relatives over sliding windows. For window-specific calculation of isoelectric point (pI), the N-terminus amino group is added to the first window and the C-terminus carboxyl added to the last window. Generated from DAMBE [49].

Figure 4

Transmembrane (TM) domain with its tripartite structure (juxtamembrane aromatic part in blue, central hydrophobic part in pink, and cysteine-rich part in purple) and the cytoplasmic tail that anchors inside the viral membrane.

Figure 5

Two amino acid replacements that stabilize the spike protein at the prefusion state. (A) Amino acids KY in the native state of SARS-2-S is replaced by PP spike variant S-2P used in the FDA-approved Pfizer/BioNTech and Moderna vaccine; (B) Partial structure from 6VSB showing the two proline residues stabilizing the structural bend.