Targeted N vs. S protein in COVID-19 Antibody Testing

Throughout the COVID-19 pandemic, researchers have been working tirelessly to understand the human immune response to SARS-CoV-2, including the duration and level of protection that antibodies may provide against re-infection.
Several tests have already been applied which can detect antibodies against SARS-CoV-2 and identify individuals who have experienced prior infection with the virus or vaccination.
Some tests detect the presence of antibodies against the spike (S) protein, while others detect antibodies against the nucleocapsid (N) protein.

About S and N proteins

Coronavirus has four main structural proteins: nucleocapsid (N), spike (S), membrane (M) and envelope (E). The S protein consists of the S1 and S2 subunits. The S protein is highly immunogenic since it is located on the surface of the virus.

Picture from Cell, 2020, doi:10.1016/j.cell.2020.04.011

The N protein plays an important role in the transcription and replication of viral RNA, packaging the encapsidated genome into virions and inhibits the cell cycle process of the host cells. The N protein is the most abundant protein in virions, and also has high immunogenic activity. Both N and S protein could be potential targets for the antibody-based detection of SARS-CoV-2. However, the N protein homology between SARS-CoV-2 and SARS-CoV is 90 percent, compared with the S protein (77 percent), especially the S1 subunit including the RBD (66 percent).

Similar to other coronaviruses, SARS-CoV-2 entry into host cells is mediated by the transmembrane spike (S) glycoprotein, which forms prominent homotrimers protruding from the viral surface.

S1 subunit also has evolutionary low protein homologies within the coronavirus family suggesting that it could potentially demonstrate less cross-reactivities among the endemic coronaviruses. Therefore, the S1 subunit could be the specific target antigen for detecting SARS-CoV-2 antibodies.

Although coronavirus infections induce a strong antibody (Ab) response against N, these Abs are not neutralizing.

S protein comprises an S1 subunit, which recognizes host cell receptors (and is divided into A, B, C, and D domains), and an S2 subunit that promotes fusion of the viral and cellular membranes to initiate infection

The S1 subunit domain B (so-called receptor-binding domain or RBD) binds to angiotensin-converting enzyme 2 (ACE2), which serves as an entry receptor. The SARS-CoV-2 RBD is the main target of neutralizing Abs, which was demonstrated on a research published on Cell.

Therefore, the presence of antibodies alone does not indicate that the person is immune against potential re-infection. It is important to determine whether the antibodies can confer protective immunity long-term, providing virus-neutralizing antibodies that block viral infection, and help in clearing viral infection. Neutralizing antibodies primarily target S protein in coronaviruses, in particular the S1 subunit and the RBD contained within the S1 subunit, preventing viral entry into the host cell.

A research published on Cell shows that in a cohort of 647 SARS-CoV-2-infected subjects, both the magnitude of Ab responses to SARS-CoV-2 spike (S) and nucleoprotein and nAb titers correlate with clinical scores. The receptor-binding domain (RBD) is immunodominant and the target of 90% of the neutralizing activity present in SARS-CoV-2 immune sera.

 

Reference

[1] Luca Piccoli et al. Mapping neutralizing and immunodominant sites on the SARS-CoV-2 spike receptor-binding domain by structure-guided high-resolution serology. Cell, 2020, doi:10.1016/j.cell.2020.09.037.

[2] https://www.technologynetworks.com/diagnostics/blog/covid-19-antibody-testing-s-vs-n-protein-340327