3D Structures of Vitamin D (25-hydroxy cholecalciferol) (Pubchem id-5280453) and its analogues such as Paricalcitol (Pubchem id-5281104), Doxercalciferol (Pubchem id-5281107), Falecalciferol (Pubchem id-5282190), Maxacalcitol (Pubchem id-6398761), Tacalcitol (Pubchem id-5283734), Calcipotriol (Pubchem id-5288783), Alfacalcidol (Pubchem id-66577031), Eldecalitol (Pubchem id-6918141), Selocalcitol (Pubchem id-5288149), Lexacalcitol (Pubchem id-5288670) and Inecalcitol (Pubchem id-6915835) were obtained from Pubchem database (https://pubchem.ncbi.nlm.nih.gov/). The structures of ACE2 (PDB id-6LZG) and VDR (PDB id-1DB1) proteins were downloaded from Protein Data Bank (PDB) (https://www.rcsb.org/). To screen the interactions at first vitamin D and the aforementioned analogues were docked using AUTODOCK 4.2.6 with the known ligand binding sites of ACE2. The analogue that performed better in terms of binding energy was chosen for further studies.
The screened analogue and vitamin D were docked with VDR and ACE2 in separate runs for 6 times using AUTODOCK 4.2.6 and GOLD 5.3.0 software to obtain free energy of binding and the binding score. The interaction of the analogue at a known ligand-binding site is validated by Molecular Dynamics simulation run for 10 ns using GROMACS 2018.4 software following the instruction of the software manufacturer. Lys31 of ACE2 is essential for binding to SARS-CoV2. In VDR Tyr 143 and His305 are critical for ligand binding. [29-33]
The NZ atom of Lys31 from ACE2 and OH & NE2 of Tyr143 & His305 from VDR were chosen as the grid centre for docking.
The docked structures exhibiting the highest negative binding energy and best GOLD score with ACE2 was considered for MD simulation. In the case of VDR, docked structures at His305 were chosen. Drug topology files were generated using the SwissParam tool, and CHARMM 27 force-field was used throughout the simulation. The Protein-drug complexes were solvated in 1.2nm cubic boxes of water using the TIP3P water model, and neutralization was performed using required sodium ions (Na+). Energy minimization was performed for 50,000 steps using the steepest descent method. The system was then equilibrated using NVT and NPT protocols for 50,000 steps each. Then the entire system was considered for MD simulation of 10ns runs at 300K temperature and 1 bar pressure and results were visualized using Pymol. Xmgrace plotting tool was used to analyze the graphs of potential energy, root mean square deviation (RMSD), radius of gyration, hydrogen bonds, and other parameters as discussed in the results.