Maestro 13

The crystal structure of human NR4A1 LBD domain (PDB: 4KZI) was utilized for the modeling. The protein preparation workflow in Maestro 13.3 (Schrödinger) was used to preprocess and optimize H-bond assignment, minimize energy, and delete water. Celastrol was prepared by the LigPrep module with the OPLS4 force field. Grid box was generated centroid of Cys551 with 20 Å. Standard precision mode was selected with default parameters. The final disposition with low-energy conformation was selected. The result was visualized by PyMOL software.

Docking studies were performed using the Schrödinger Maestro 13.3 software package. The conformation of compounds was generated and energy-minimized by the LigPrep tool using force field OPLS4. The 3D crystal structure of P-gp (PDB code: 6QEE and 6QEX) was obtained from the RCSB Protein Data Bank (https://www.rcsb.org/) (Alam et al. 2019 (link)). The obtained receptor was prepared (involving optimization, charge calculation, deletion of co-crystal ligand, and addition of hydrogen, etc.) with Protein Preparation Workflow. The GLIDE tool was employed to dock ligands at the substrate and inhibitor binding sites with a high degree of accuracy (Brožová et al. 2023 (link)). The proposed binding modes were visualized using a combination of software tools, including the Maestro workspace, and PyMOL to generate high-quality images. The obtained docking results of the P-gp/drug complexes were used to predict the values of the binding affinity (ΔG) (kcal/mol) using the PROtein binding enerGY prediction (PRODIGY-LIGAND) online server tool (Vangone et al. 2019 (link)).

Autodock Vina provides better ligand-protein binding poses and accuracy than AutoDock 4.2. Hence, we have used AutoDock Vina [62 (link)] for this study's docking experiments to dock the selected ligand molecule G-CK and seven control drugs against ACE2 and TMPRSS2. The Supplementary File (S2) contains the gird size and exhaustiveness number information for all the docking procedures. All Compounds were further docked to obtain docking scores and the binding sites. Finally, we have utilized the academic version of PyMOL [63 ] MAESTRO 13.3 (Schrödinger, LLC, NY, USA), and BIOVIA Discovery Studio Visualizer (BIOVIA, Dassault Systèmes) [64 ] to visualize the ligand interactions with the active sites of the receptors.