The Case for S2: The Potential Benefits of the S2 Subunit of the SARS-CoV-2 Spike Protein as an Immunogen in Fighting the COVID-19 Pandemic

Abstract

As COVID-19 cases continue to rise, it is imperative to learn more about antibodies and T-cells produced against the causative virus, SARS-CoV-2, in order to guide the rapid development of therapies and vaccines. While much of the current antibody and vaccine research focuses on the receptor-binding domain of S1, a less-recognized opportunity is to harness the potential benefits of the more conserved S2 subunit. Similarities between the spike proteins of both SARS-CoV-2 and HIV-1 warrant exploring S2. Possible benefits of employing S2 in therapies and vaccines include the structural conservation of S2, extant cross-reactive neutralizing antibodies in populations (due to prior exposure to common cold coronaviruses), the steric neutralization potential of antibodies against S2, and the stronger memory B-cell and T-cell responses. More research is necessary on the effect of glycans on the accessibility and stability of S2, SARS-CoV-2 mutants that may affect infectivity, the neutralization potential of antibodies produced by memory B-cells, cross-reactive T-cell responses, antibody-dependent enhancement, and antigen competition. This perspective aims to highlight the evidence for the potential advantages of using S2 as a target of therapy or vaccine design.

Keywords: COVID-19; S2 subunit; SARS-CoV-2; SARS-CoV-2 vaccine; antibodies; coronavirus; immunity; spike protein.

Figure 1

SARS-CoV-2 spike glycoprotein monomer representation showing (A) functional domains and (B) comparison of amino acid sequence identity with SARS-CoV and related isolates in the wild. (A) Functional domain S1 mediates binding of the receptor binding domain (RBD) to the angiotensin converting enzyme 2 (ACE2), the host cell receptor that is specifically recognized by the receptor binding motif (RBM) interface, is cleaved (S1/S2) and shed. Shedding exposes the S2 domain. Cleavage at S2' triggers spike trimer activation, release of the fusion peptide (FP), heptad repeat 1 (HR1) and heptad repeat 2 (HR2); the membrane proximal external region (MPER) is sometimes considered part of HR2 and with a cholesterol recognition/interaction amino acid consensus (CARC) sequence, potentially participating in membrane lipid fusion. The transmembrane domain (TM) and a short cytoplasmic tail (CyT) are indicated. Cleavage sites that drive host-cell infection are shown in red. (B) Phylogenetic analysis of SARS-CoV-2 domain sequences identity among SARS-CoV-2, bat and pangolin's isolates, and SARS-CoV are tabulated for each domain. A high degree of sequence identity for the RBD in SARS-CoV correlates well with ACE2 receptor recognition. However, the RBM-ACE2 interface is 50% identical and may account for the increased ACE2 binding affinity. Sequence identities are high for all functional segments of S2, the Type I metastable domain that induces viral-host cell membrane fusion, suggesting an optimum sequence-structure-function relationship. Metastability is a functional requirement, allowing these proteins to refold into a lower energy conformation while transferring the difference in energy to catalyze the membrane fusion reaction. Structural studies have shown that stable immunogens presenting the same antigenic sites as the labile wild-type proteins efficiently elicit potent neutralizing antibodies. In the alternative endosomal pathway that is apparently mediated by NTD, the fusion machine of S2 is equally required to infect the host cells (9, 10). Furthermore, detrimental amino acid substitutions found in the S2 domain of Hepatitis Mouse Virus, but not in SARS-CoV-2, affected the post fusion conformational stability, explaining a reported reduction of S-mediated membrane fusion (11).