How natural alterations inside the SARS-CoV-2 glycan barrier affect spike protein dynamics

By Shanet Susan AlexAug 22 2022Reviewed by Danielle Ellis, B.Sc.

In a recent article published in the bioRxiv* preprint server, researchers explored how the natural alterations inside the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) glycan barrier affect the spike (S) protein dynamics.

This news article was a review of a preliminary scientific report that had not undergone peer-review at the time of publication. Since its initial publication, the scientific report has now been peer reviewed and accepted for publication in a Scientific Journal. Links to the preliminary and peer-reviewed reports are available in the Sources section at the bottom of this article. View Sources

Background

The SARS-CoV-2 pandemic, which has caused a long-lasting global medical crisis, has led to the death of over 6.4 million people worldwide. The effectiveness of current protection, whether it develops naturally or as a result of vaccination, is altered by the emergence of SARS-CoV-2 variants. 

The analysis of the structure of the SARS-CoV-2 S glycoprotein will aid in comprehending the impacts of antigenic surface mutations. One type of mutation affects the attachment sites for glycosylation, which can impact the antigenic architecture further than the primary attachment site.

About the study

The present study aimed to comprehend potential SARS-CoV-2 immune evasion pathways involving glycan shield modifications.

The researchers interrogated the glycan barrier of the SARS-CoV-2 variant of concern (VOC) S protein. Further, they investigated their influence on angiotensin-converting enzyme 2 (ACE2) adhesion. For this, the investigators engineered the sequences for the SARS-CoV-2 P.1 (Gamma), B.1.617.2 (Delta), and B.1.351 (Beta) variants to generate soluble recombinant native-like trimers. They analyzed the compositional variations in the site-specific glycan shield of SARS-CoV-2 VOCs employing liquid chromatography-mass spectrometry (LC-MS).

The team produced glycopeptides with a single asparagine (N)-linked glycosylation site (PNGS). For this, they used chymotrypsin, alpha-lytic, and trypsin protease. In addition, the glycopeptide compounds were exposed to higher energy collision-induced dissociation (HCD) fragmentation.

The authors sought to comprehend how the landscape of the protein might affect the N188 glycosylation of the Gamma S glycoprotein. For this, they used conventional molecular dynamics (MD) to conduct comprehensive sampling on two Gamma S models, one of which had Man5GlcNAc2 (Man5) at N188 and the other did not have glycosylation at this site. Moreover, the researchers recreated the ectodomain of the P.1 S glycoprotein from the structure obtained by cryogenic electron microscopy (cryo-EM), i.e., PDB 7SBS.

Results and discussions

The study results offer a molecular understanding of how the SARS-CoV-2 Gamma variant with the extra N188 glycan site exhibits superior shielding of the S glycoprotein surface by fusing compositional glycan analyses with MD simulations. Additionally, the team found minimal alterations across the glycan barrier of other VOCs, potentially indicating that SARS-CoV-2 has not yet fully utilized the capacity of glycan-facilitated immune escape like other viruses have. 

Despite having more than sixty N-glycan sites throughout the trimer, the SARS-CoV-2 S protein's glycan barrier density was low compared to that of influenza, human immunodeficiency virus 1 (HIV-1), and Lassa virus (LASV). Hence, a significant portion of the immunogenic protein surface was left accessible to the antibody-mediated immune responses.

The scientists then stated that SARS-CoV-2, a virus that has just recently started to circulate in humans, has so far taken advantage of simple adaptations to increase its infectivity and bypass host immunity, like modifications to the receptor binding domain (RBD).

The authors pointed out that altering the glycan barrier probably negatively influences viral infectiousness. Earlier studies have shown a buildup of N-linked glycan sites upon the haemagglutinin ectodomain, and a corresponding rise in oligomannose-type glycan aggregation in influenza H3N2 has been attributed to recurrent endemic circulation and developing immunity among humans. Further, the gradual plateau of PNGS accumulation shows that increasing glycan shielding adversely affects viral fitness.

SARS-CoV-2 does not have N370, an RBD glycosylation site, compared to other coronaviruses (CoVs), possibly enhancing SARS-CoV2's infectiousness. The investigators stated that presumably, as infections and vaccines persist, SARS-CoV-2 evolution will run out of simple amino acid deletions and substitutions and start to alter the glycan shield, with the associated fitness costs that may be involved.

Conclusions

Overall, the study findings indicated that the N188 site in SARS-CoV-2 exhibited very little glycan maturation, which was congruent with the accessibility of a few enzymes. MD and structural modeling revealed that N188 was located inside a cavity in the RBD, affecting the dynamics of these adhesion domains. The study observations illustrate a mechanism via mutations to the SARS-CoV-2's glycosylation sites that affect the antigenic surface's structural integrity.

Besides, the scientists mentioned that other CoVs circulating among humans for longer, like NL63, the common cold CoV, were significantly more densely glycosylated and harbored about twice as many glycan domains per protomer versus SARS-CoV-2. Thus, to better understand how the glycans of SARS-CoV-2 S glycoproteins manipulate the host immune system when new VOCs arise, methods like those presented in the present manuscript would be helpful.

This news article was a review of a preliminary scientific report that had not undergone peer-review at the time of publication. Since its initial publication, the scientific report has now been peer reviewed and accepted for publication in a Scientific Journal. Links to the preliminary and peer-reviewed reports are available in the Sources section at the bottom of this article. View Sources