As the ongoing coronavirus disease 2019 (COVID-19) pandemic continues to take lives worldwide, leaving more than 2.27 million dead, the use of vaccines and therapeutic antibodies remains a matter of urgency. Even as vaccines are starting to roll out in many parts of the world, new and more infectious variants are emerging, raising concerns about the long-term efficacy of the present models. Three important new variants of concern all possess the N501Y mutation.
A new preprint recently released on the bioRxiv* preprint server reports the underlying basis of such an increase in infectiousness associated with the N501Y, a mutation at the site of virus-host receptor interaction. This mutation has been found in the UK, South African and Brazilian variants. These are also called B.1.1.7, 20I/501Y.V1, B.1.351, 20H/501Y.V2, and P1, 20J/501Y.V3, respectively.
The virus engages with the host cell via its spike protein, binding through the receptor-binding domain (RBD), with the host cell receptor, the angiotensin-converting enzyme 2 (ACE2).
Antibodies target spike RBD
The vast majority, about 90%, of neutralizing antibodies against the virus target the RBD. Regeneron and Eli Lilly have both produced monoclonal antibodies that are directed specifically against the native form of the spike protein, and both have been approved by the US Food and Drug Administration (FDA) for emergency use.
There are now several vaccines that have received emergency use approval, including the two mRNA vaccines, from Pfizer-BioNTech and Moderna, and the adenoviral vectored vaccine from Oxford Astra-Zeneca, all of which elicit antibodies against the native spike and RBD.
However, new variants like the UK and South African variants continue to emerge, raising doubts about whether the antibodies elicited by these vaccines, or therapeutic antibodies raised against earlier variants, will continue to be effective against them. This is a matter of concern, especially given their increased transmissibility.
Tenfold rise in binding affinity
The researchers used Surface Plasmon Resonance binding assays to measure the binding affinity of the wildtype and mutant spike for ACE2. They found that the 501Y mutation in the RBD is associated with a tenfold rise in binding affinity for ACE2, relative to the original N501-RBD, at ~0.566 nM vs. ~5.76 nM, respectively.
A similar increase in binding affinity has been reported for SARS-CoV-2-ACE2 binding, compared to SARS-CoV-ACE2, respectively, which could be a substantial reason for the higher transmissibility of SARS-CoV-2.
Mechanism of increased binding affinity
The reason could lie in the structural differences. No current structural model shows the effect of the isolated N501Y mutation on the binding affinity of the spike for ACE2. The researchers, therefore, constructed computational models of the structure of the RBD in these two forms of the spike protein.
This showed the potential for additional interactions between aromatic residues, as well as additional hydrogen bonds with the ACE2. Though the RBD-N501 shows a hydrogen bond with the Y41 of the ACE2 – that is abolished with the Y501 substitution – the latter introduces new hydrogen bonds and hydrophobic interaction.
The new hydrogen bonds were formed between Y501 and ACE2 residues D38 and K353. The substitution also introduces an aromatic ring that causes a strong aromatic ring or pi stacking with that of the ACE2 Y41 residue.
The potential interference with such stacking by the side chain of the residue R498 could be prevented by the formation of a negatively charged cage to contain this side chain. Such a cage can be formed by two aromatic rings and the D38 side chain, but further study is required to confirm its existence.
This leads to much stronger and more stable spike-ACE2 interactions.
The current mutation of N501 to Y501 and consequent ~10 times greater affinity for ACE2 may therefore account for the increased rate of infections in the United Kingdom and, likewise, for the increased transmission rate of both the South African (20H/501Y.V2) and Brazil variants (20J/501Y.V3), although their K417N, E484K besides N501Y changes may also contribute.”
Neutralization of variants by vaccine-induced/therapeutic antibodies
Immunization with the Pfizer-BioNTech vaccine generated high titers of antibodies, indicating a robust immune response. However, when tested with sera from vaccinated individuals at different dilutions, the researchers observed efficient neutralization of both N501-RBD and Y501-RBD binding to the ACE2 receptor.
High levels of neutralization were observed with undiluted serum, and at up to fivefold dilutions, but slightly less for the Y501 variant.
The same was true of the Eli Lilly monoclonal antibody Bamlanivimab (LY-CoV555), which was isolated from a convalescent COVID-19 patient. It was reported to inhibit the N501-RBD-ACE2 binding. Structural studies showed that it binds to the RBD near the site of the Y501 mutation. The researchers found the antibody bound with equivalent efficiency of binding to Y501-RBD and N501-RBD.
The binding affinity of the N501Y mutant for ACE2 is higher than its affinity for the antibody, however, even while the mutant strain spike protein shows tenfold higher affinity for ACE2 relative to the wildtype spike protein.
What are the implications?
These findings suggest that both the Pfizer-BioNTech vaccine and Bamlanivimab “should be still effective for COVID-19 patients contracting either with wildtype SARS-COV-2 or with the 501Y.V1 variant.”
However, since the binding affinity of the Y501-RBD UK spike variant for the antibody is lower than its affinity for ACE2, neutralization may be achieved only with higher concentrations of Bamlanivimab in patients infected with this strain.
The South African and Brazil variants have K417N and E484K mutations in addition to the N501Y mutation, both at the RBD. The E484 forms a salt bridge with the R50 residue in the antibody, and this may be abolished or drastically reduced with the K484 variant. If so, this could lead to loss of efficacy of Bamlanivimab against these variants.
From this regard, therapeutic antibodies to treat COVID-19 patients should be adjusted for new emerging variants.”
bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.