Exploring Highly Conserved Regions of SARS-CoV-2 Spike S2 Subunit as Targets for Fusion Inhibition Using Chimeric Proteins

Abstract

Since the beginning of the COVID-19 pandemic, considerable efforts have been made to develop protective vaccines against SARS-CoV-2 infection. However, immunity tends to decline within a few months, and new virus variants are emerging with increased transmissibility and capacity to evade natural or vaccine-acquired immunity. Therefore, new robust strategies are needed to combat SARS-CoV-2 infection. The viral spike composed of S1 and S2 subunits mediates viral attachment and membrane fusion to infect the host cell. In this process, interaction between the highly conserved heptad repeat 1 and 2 regions (HR1 and HR2) of S2 is crucial and for this reason; these regions are promising targets to fight SARS-CoV-2. Here, we describe the design and characterization of chimeric proteins that structurally imitate the S2 HR1 region in a trimeric coiled-coil conformation. We biophysically characterized the proteins and determined their capacity to bind the HR2 region, as well as their inhibitory activity of SARS-CoV-2 infection in vitro. HR1 mimetic proteins showed conformational heterogeneity and a propensity to form oligomers. Moreover, their structure is composed of subdomains with varied stability. Interestingly, the full HR1 proteins showed high affinity for HR2-derived peptides and SARS-CoV-2 inhibitory activity, whereas smaller proteins mimicking HR1 subdomains had a decreased affinity for their complementary HR2 region and did not inhibit the virus. The results provide insight into effective strategies to create mimetic proteins with broad inhibitory activity and therapeutic potential against SARS-CoV-2.

Global genomic diversity and conservation of SARS-CoV-2 since the COVID-19 outbreak

“…There are one and three different amino acid sequences in Nsp3 and Nsp8, which can be used as important sites for virus identification. S protein contains multiple domains; S1 plays a role in the binding of receptor ACE2 ( 21 ), whereas S2 mediates the membrane fusion process ( 22 ). Partial domains and sequence identity are highly correlated; the NTD domain in S1 and HR1 in S2 showed a significantly reduced (16.9% and 52.3%), while the consistency of other parts was 69.2%; they showed clear demarcation ( Fig.…”

Broad Epitope Coverage of Therapeutic Multi-Antibody Combinations Targeting SARS-CoV-2 Boosts In Vivo Protection and Neutralization Potency to Corner an Immune-Evading Virus

“…In this study, two epitope bins (C and S2) outside the RBD of the S-protein were included in TATX-03 to engage other MoAs. Among the selected non-RBD-blockers, Ab 2-A6 binds to the S2 region, which is considered a highly conserved domain and therefore a promising target for a sustainable therapeutic [ 62 , 63 , 64 , 65 ]. However, Abs recognizing this region of the spike trimer are rarely reported.…”

“…A combination of two of Abs discovered from B cells harvested from vaccinated convalescents neutralized Omicron variants BA.2.75, BA.2.75.2, BF.7, BQ.1, BQ.1.1, CH.1.1, XBB, and XBB.1 [ 8 ]. In TATX-03, clones 2-A6 and, to a lesser extent, 22-E7 bind to regions relatively less prone to mutations, with 2-A6 binding to the S2 subunit involved in the membrane fusion process of the virus with the host cell [ 64 ].…”