Molecular dynamics simulations and functional studies reveal that hBD-2 binds SARS-CoV-2 spike RBD and blocks viral entry into ACE2 expressing cells

Abstract

New approaches to complement vaccination are needed to combat the spread of SARS-CoV-2 and stop COVID-19 related deaths and long-term medical complications. Human beta defensin 2 (hBD-2) is a naturally occurring epithelial cell derived host defense peptide that has antiviral properties. Our comprehensive in-silico studies demonstrate that hBD-2 binds the site on the CoV-2-RBD that docks with the ACE2 receptor. Biophysical and biochemical assays confirm that hBD-2 indeed binds to the CoV-2-receptor binding domain (RBD) (KD ∼ 300 nM), preventing it from binding to ACE2 expressing cells. Importantly, hBD-2 shows specificity by blocking CoV-2/spike pseudoviral infection, but not VSV-G mediated infection, of ACE2 expressing human cells with an IC50 of 2.4± 0.1 μM. These promising findings offer opportunities to develop hBD-2 and/or its derivatives and mimetics to safely and effectively use as novel agents to prevent SARS-CoV-2 infection.

Ramping Up Antimicrobial Peptides Against Severe Acute Respiratory Syndrome Coronavirus-2

“…Additionally, reduced levels of VREs (important in AMP induction (McMahon et al, 2011;Aguilar-Jimenez et al, 2013)) in cells isolated from bronchoalveolar lavage were found in patients infected with CoV-2 than healthy subjects (George et al, 2020). Moreover, the fact that LL-37 (Zhang et al, 2020) and hBD-2 (Zhang L. et al, 2021), both regulated through Vit D 3 , were able to block S protein-expressing pseudovirions from infecting vulnerable human cells supports the notion that healthy levels of Vit D 3 may be important in reducing the risk of acquiring SARS-CoV-2 infection. However, further studies on the direct correlation between Vit D 3 and AMP levels in relation to susceptibility to CoV-2 acquisition are required.…”

“…Recent in silico molecular docking studies predicted strong binding interactions of LL-37 (Lokhande et al, 2020) and hBD-2 (Zhang L et al, 2021) with the receptor-binding domain (RBD) of SARS-CoV-2, suggesting an RBD blocking potential for these two peptides. Biophysical assays using bio-layer interferometry (BLI) and microscale thermophoresis (MST) supported the in silico findings for LL-37 (Wang C. et al, 2021) and hBD-2 (Zhang L. •Aggregates IAV and enhances neutrophil-mediated clearance [Hartshorn et al (2006), Tecle et al (2007), Doss et al (2009)] •Inhibits IAV replication through the inhibition of protein kinase C (PKC) in infected cells [Salvatore et al (2007)] HD5…”

“…•Promotes reduction of the metabolic activity of infected cells [Sousa et al (2017)] et al, 2021), respectively. Additionally, biochemical studies with hBD-2 showed that it inhibited the RBD from binding ACE2 and prevented S protein-expressing pseudovirions from infecting ACE2-expressing human cells (Zhang L. et al, 2021). Interestingly, LL-37 has been found to suppress S pseudovirion infection in a dose-dependent manner with an IC 50 value of 1.05 μM (Wang C. et al, 2021).…”

“…Recent in silico molecular docking studies predicted strong binding interactions of LL-37 ( Lokhande et al, 2020 ) and hBD-2 ( Zhang L et al, 2021 ) with the receptor-binding domain (RBD) of SARS-CoV-2, suggesting an RBD blocking potential for these two peptides. Biophysical assays using bio-layer interferometry (BLI) and microscale thermophoresis (MST) supported the in silico findings for LL-37 ( Wang C. et al, 2021 ) and hBD-2 ( Zhang L. et al, 2021 ), respectively. Additionally, biochemical studies with hBD-2 showed that it inhibited the RBD from binding ACE2 and prevented S protein–expressing pseudovirions from infecting ACE2-expressing human cells ( Zhang L. et al, 2021 ).…”

“…FIGURE 1 | Schematic representation showing differential binding of human AMPs to SARS-CoV-2 S protein or ACE2 on cell surfaces. LL-37 and hBD-2 bind to SARS-CoV-2 S protein and inhibit its binding to ACE2 on the cell surface prior to entry(Lokhande et al, 2020;Roth et al, 2020;Wang C. et al, 2021;Zhang L. et al, 2021). In contrast, HD-5 binds to ACE2 to inhibit viral entry(Wang et al, 2020a) [LL-37 has also been shown to bind ACE2, however, with lower affinity than S-RBD(Wang C. et al, 2021)].…”