Phase III: MD simulation of the spike protein-ACE2 complex in the absence and presence of calcitriol

With regards to the absence of calcitriol (calcitriol: the only steroid molecules which could interact with spike contacting site), MD simulation of spike-ACE2 started from the crystallography structure (6LZG). In this case, spike and ACE2 were glycosylated with N-acetyl-D-glucosamine at position N343 in RBD of spike and N53, N90 and N322 in ACE2. Protein–protein docking simulation approach was conducted on the spike protein-ACE2 complex in the presence of calcitriol using the HADDOCK 2.4 web server⁵⁵. The proteins applied to the HADDOCK 2.4 web server were the complex of spike protein with steroid molecules (from phase II MD simulation, spike protein-calcitriol) and optimized ACE2. The active residues at the contacting sites of two proteins are listed as follows: spike protein (Lys417, Tyr449, Leu455, Phe456, Ala475, Gly476, Phe486, Asn487, Tyr489, Gln493, Gly496, Gln498, Thr500, Asn501, Gly502, Tyr505, Gln506) and ACE2 (Gln24, Thr27, Phe28, Lys31, His34, Glu35, Glu37, Asp38, Tyr41, Gln42, Met82, Tyr83, Lys353, Asp355, Arg357)^30,31. The residues shown in bold are involved in the formation of hydrogen bonds between spike protein and ACE2. Interactive residues prepared by DIMPLOT were provided in Fig. ^S4A. The results obtained from the docking of spike protein-calcitriol-ACE2 via HADDOCK had 12 clusters and 166 conformers. The average HADDOCK score for the best cluster (top score) was − 122.0 ± 4.1 (a. u.). The portion of electrostatic energy (− 159.6 ± 24.6 kcal/mol) was greater than the Van der Waals energy (− 71.8 ± 3.4 kcal/mol) in the interaction between spike protein and ACE2 in the presence of calcitriol. The MD simulation and the MM/PBSA approach were then used to calculate the binding free energy of coronavirus spike protein to its receptor, ACE2, in the absence or presence of calcitriol^56,57.