The dockings were performed in a 64 bit PC. The receptor design was made by using SWISS-MODEL, a fully automated protein structure homology-modeling server. In this tool, energy minimization and simulated annealing are done with the GROMOS96 forcefield [13]. The 2D structures of the ligands were drawn, optimized with full hydrogen LEE011 bonds and saved as .sk2 format using ChemSketch tool from Advanced Chemistry Development, Inc. (ACD/ChemSketch, [14]) and the 3D structures were obtained using PRODRG server [15]. Receptors The wild type receptor was derived from

the crystal structure of deazaflavin dependent nitroreductase (3R5W) [16]. The mutant receptor was designed by introducing A76E mutation [9], in the sequence of Ddn and modeling it using SWISS MODEL without the presence

of its cofactor F420. Ligands The ligands were derived from the structure of PA-824 by removing the trifluoromethyl group (CF3) and replacing it with key anti-M. tuberculosis drugs such as isoniazid, moxifloxacin, gatifloxacin etc., through ester linkages. The removal of trifluoromethyl group was done on the basis to reduce the toxicity [17]. The designed combinatorial ligands are listed in Table 2. Table 2 Docking values of PA-824 and its novel analogues with the wild and mutant Ddn receptor S.no. Drug Docking score with wild type receptor (kcal/mol) Docking score with A76E mutant receptor (kcal/mol) Structure of the analogues 1 PA-824 −6.9 https://www.selleckchem.com/products/pifithrin-alpha.html −6.7 2 Ligand 1 (glucose) −7.6 −7.0 3 Ligand 2 (Nitroglucose) −7.6 −7.2 4 Ligand 3 (Hydroxyl modification) −6.3 −6.3 5 Ligand 4 (Biotin) −7.1 −6.7 6 Ligand 5 (Cholestryl ester) −8.3 −6.9 7 Ligand 6 (gati) −8.4 −8.1 8 Ligand 7 (isoniazid) −7.8 −7.5 9 Ligand 8 (Moxi) −7.7 −8.5 10 Ligand 9

(PZA) −7.2 −7.2 11 Ligand 10 (Trehalose) −8.0 −7.7 Analysis of binding The binding sites for the docking were designed such that the entire receptor molecule was included within the selection grid. The highest Masitinib (AB1010) binding energy values corresponding to the RMSD value of zero were considered as the binding affinity value of the ligands for each docking. The Hydrogen bond interactions were obtained using Molegro molecular viewer (molegro.com) [18]. Results Bactericidal activity The results of the bactericidal activity of different drugs from the two sets of experiments are given in Figure 1. PA-824 at lower concentration of 3 μg/ml (P1) had less activity on all the days resulting with a log of 4.9 CFU/ml on the 21st day. Rifampicin (1 μg/ml) showed slightly increased activity than PA-824 at a lower concentration of 3 μg/ml, with a reduction in the count of 1.42 log cfu/ml on the 7th day, whereas for PA-824 at a concentration of 12.5 μg/ml (P2), showed a decrease in the count to log of 2.49 CFU/ml on the same day. A small reduction in RIF activity was seen on the 7th day, and on 14th day reduction of 2.