Molecular dynamics simulations and functional studies reveal that hBD-2 binds SARS-CoV-2 spike RBD and blocks viral entry into ACE2 expressing cells
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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) (K D ∼ 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 IC 50 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.
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SciScore for 10.1101/2021.01.07.425621: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
NIH rigor criteria are not applicable to paper type.Table 2: Resources
Antibodies Sentences Resources Membranes were blocked with 5% milk in TBST and then probed with goat anti-human BD2 antibody (0.2 µg/ml; Peprotech), followed by secondary antibody (1:5000) at room temperature, and visualized by enhanced chemiluminescence. anti-human BD2suggested: (GenWay Biotech Inc. Cat# GWB-BD2A5C, RRID:AB_10526593)Experimental Models: Cell Lines Sentences Resources Cells: HEK 293T and HEK 293T cells stably expressing ACE2 receptor (ACE2 HEK293T) were cultured in DMEM media containing 10% FBS, 100 U/ml penicillin/streptomycin and 4 mM L-Glutamine. HEK 293Tsuggested: NoneTo study the ability of hBD2 to compete with RBD … SciScore for 10.1101/2021.01.07.425621: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
NIH rigor criteria are not applicable to paper type.Table 2: Resources
Antibodies Sentences Resources Membranes were blocked with 5% milk in TBST and then probed with goat anti-human BD2 antibody (0.2 µg/ml; Peprotech), followed by secondary antibody (1:5000) at room temperature, and visualized by enhanced chemiluminescence. anti-human BD2suggested: (GenWay Biotech Inc. Cat# GWB-BD2A5C, RRID:AB_10526593)Experimental Models: Cell Lines Sentences Resources Cells: HEK 293T and HEK 293T cells stably expressing ACE2 receptor (ACE2 HEK293T) were cultured in DMEM media containing 10% FBS, 100 U/ml penicillin/streptomycin and 4 mM L-Glutamine. HEK 293Tsuggested: NoneTo study the ability of hBD2 to compete with RBD binding to ACE2, ACE2 HEK 293T cells were seeded in 6 cm plates. ACE2 HEK 293Tsuggested: NoneSoftware and Algorithms Sentences Resources After a brief energy minimization using the conjugate gradient and line search algorithm, 4 ps of dynamics was run at 50 K, and then the system was brought up to 310 K over an equilibration period of 1 ns using NAMD program version 2.12 (Phillips et al., 2005). NAMDsuggested: (NAMD, RRID:SCR_014894)Immunoprecipitation and Western blotting: To study interaction between hBD-2 and His-RBD, recombinant hBD-2 (Peprotech, Inc.) with or without recombinant HIS-tagged-RBD (Sino Biologicals, Inc.) were pre-incubated at room temperature for 1 h in binding buffer (30mM HEPES pH 7.6, 5mM MgCl2, 150mM NaCl, 0.5mM dithiothreitol, 1 % Triton X-100 and 1mM EDTA) and then incubated with washed Ni-NTA agarose resin beads (25 µl) overnight at 4°C. Sino Biologicalssuggested: NoneResults from OddPub: Thank you for sharing your data.
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
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