Naturally mutated spike proteins of SARS-CoV-2 variants show differential levels of cell entry

Abstract

1The causative agent of the coronavirus disease 2019 (COVID-19) pandemic, severe 2 acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is steadily mutating during continuous 3 transmission among humans. Such mutations can occur in the spike (S) protein that binds to the 4 angiotensin-converting enzyme-2 (ACE2) receptor and is cleaved by transmembrane protease 5 serine 2 (TMPRSS2). However, whether S mutations affect SARS-CoV-2 infectivity remains 6 unknown. Here, we show that naturally occurring S mutations can reduce or enhance cell entry 7 via ACE2 and TMPRSS2. A SARS-CoV-2 S-pseudotyped lentivirus exhibits substantially lower 8 entry than SARS-CoV S. Among S variants, the D614G mutant shows the highest viral entry, as 9 supported by structural observations. Nevertheless, the D614G mutant remains susceptible to 10 neutralization by antisera against prototypic viruses. Taken together, these data indicate that the 11 D614G mutation enhances viral infectivity while maintaining neutralization susceptibility. 12 13 3

Structural insights into spike protein and its natural variants of SARS-CoV-2 found on Mexican population.

“…5D), this modi cation not only increases the exibility of the hinge but also it changes the direction which might contribute to the increase virulency of this mutant because of might alter the binding to ACE protein 13 . Previously, it was found that D614 formed a hydrogen bond with T859 and a salt bridge with K854 placed on the subunit two of the other protomer, thus the change of D by G might provide a exible space between the monomers and more conformational exibility as our MD simulations suggest which might lead to improved accessibility to ACE2, which are consistent with more cell entry of the virus that carries this mutation 8,34 . This motion perturbation is explained because D614 residue is located on a loop exposed side chain, has a negative charge, and that will be a loss on the variant G614, this causes loss of interactions with other molecules or residues.…”

“…D614G change is caused by an A/G nucleotide mutation at position 23,403 in the reference strain 32 , this variant is being associated with higher viral loads and enhances viral effectivity in patients 8,32,33 despite the higher viral loads, G614 is neutralizing similarly as D614 by polyclonal antibody 32 . To date, this variant becomes the dominant form displacing the WT, according to the levels of mutation in the world, presented on the nextstrain database (www.nextstrain.org).…”

“…It was signi cantly decreased on S2 region (816-1106), suggesting that this mutant not only confers more structural stability as we found but also energetic stability as it was suggested by other authors 13 . Aside, it was found that the H49Y mutant virus has increased cell entry in comparison to WT S protein 8 .…”

“…Recent studies show SARS-CoV-2 is mutating during the continuous transmissions among the population 8,9 . To date, the Mexican population is circulating two of three viral lineages reported 10 .…”

Nanoluciferase complementation-based biosensor reveals the importance of N- linked glycosylation of SARS-CoV-2 Spike for viral entry

“…Interestingly, two N-linked glycan modi cations occur within the conserved Receptor Binding Domain or RBD of the S protein. The RBD mediates the binding of the S protein to ACE2 7,8 and while there have been a number of documented polymorphisms in the amino acid sequence of the RBD from clinical isolates around the world 9 , these two glycosylation sites are uniformly conserved. This suggested to us the possibility that glycosylation of the RBD is important for its binding to the cellular ACE2 receptor or as suggested earlier inhibits immune recognition.…”