Lectin-like Intestinal Defensin Inhibits 2019-nCoV Spike binding to ACE2

Cheng Wang; Shaobo Wang; Daixi Li; Xia Zhao; Songling Han; Tao Wang; Gaomei Zhao; Yin Chen; Fang Chen; Jianqi Zhao; Liting Wang; Wei Sun; Yi Huang; Yongping Su; Dongqing Wei; Jinghong Zhao; Junping Wang

( Figure 5A ). The content of S1 binding to the cells preincuabted with HD5 was 3.4-fold lower than that binding to the cells without treatment ( Figure 5B) . Notably, the protection of HD5 was in a dose-dependent manner ( Figure 5C ). As shown in Figure 5D , HD5 could significantly decrease S1 binding to epithelial cells at concentrations as low as 10 µg/mL, which is within the range of HD5 concentration in intestine 22 . The protective effect of HD5 was also observed for human renal proximal tubular epithelial cells abundant in ACE2 ( Figure S4 ) 16, 17 . However, S1 pretreated with HD5 was still efficient to contact Caco-2, as revealed by either immunoblotting or immunofluorescence. These data are in line with the results shown in Figure 1E and 2B, demonstrating that HD5 inhibits 2019-nCoV S1 adhering to host cells possibly by blocking ACE2 but not S1. The copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/2020.03.29.013490 doi: bioRxiv preprint chromatography (RP-HPLC) and electrospray ionization mass spectrometry, respectively, were shown in Table S1 . BLI-based blocking assay was conducted by monitoring the binding thickness of S1-ACE2 interaction with the interference of HD5. S1 and ACE2 were loaded on HIS1K and AR2G biosensors, respectively, at a concentration of 15 μg/mL. The immobilized ACE2 was incubated with 200 and 50 nM HD5 for 300 s at 30 o C. After a 300 s of disassociation, the signals of 400 nM S1 binding to ACE2 were recorded for another 300 s. Otherwise, the immobilized S1 was covered with 100 nM HD5 for 300 s. The bindings of 100 nM ACE2 to S1 were recorded for 300 s after peptide disassociation. 44 ) and the LBD of ACE2 (PDB: 6ACG 45 ) was performed on ZDOCK server 46 . The Gromacs 2020 software package 47 , AMBER99SB-ILDN force field, and TIP3P water model were applied for the simulation with a time step of 2 fs. Firstly, 1000-step minimization was carried out. Then, for fully relaxation, four 1-ns pre-equilibration simulation with restrained coordinates of the atoms belonging to the heavy atoms, main chain, backbone, and C-α, respectively, were performed step by step. Finally, each of five production simulation with isothermal-isobaric (NPT) ensemble at 1 atm and 298 K was performed for 20 ns. The binding free energy was author/funder. All rights reserved. No reuse allowed without permission.