High-Throughput Virtual Screening for a New Class of Antagonist Targeting LasR of Pseudomonas aeruginosa

Abstract

Pseudomonas aeruginosa, an opportunistic human pathogen, causes fatal effects in patients with cystic fibrosis and immunocompromised individuals and leads to around 1000 deaths annually. The quorum sensing mechanism of P. aeruginosa plays a major role in promoting biofilm formation and expression of virulent genes. Hence, quorum sensing inhibition is a promising novel approach to treat these bacterial infections as these organisms show a wide range of antibiotic resistance. Among the interconnected quorum sensing network of P. aeruginosa, targeting the las system is of increased interest as its principal receptor protein LasR is the earliest activated gene. It is also shown to be involved in the regulation of other virulence-associated genes. In this study, we have applied high-throughput virtual screening, an in silico computational method to identify a new class of LasR inhibitors that could serve as potent antagonists to treat P. aeruginosa-associated infections. Three-tire structure-based virtual screening was performed on the Schrödinger small molecule database, which resulted in 12 top hit compounds with docking scores lesser than −11.0 kcal/mol. Three of these best-scored compounds CACPD2011a-0001928786 (C1), CACPD2011a-0001927437 (C2), and CACPD2011a-0000896051 (C3) were further analyzed. The binding free energies of these compounds in complex with the target protein LasR (3IX4) were evaluated, and the pharmacokinetic properties were determined. The stability of the docked complexes was assessed by running a molecular dynamics simulation for 100 ns. Molecular dynamics simulation analysis revealed that all three compounds were found to be in stable contact with the protein over the entire simulation period. The antagonistic effect of these compounds was validated using the LasR reporter gene assay in the presence of acyl homoserine lactone. Significant reduction in the β-galactosidase enzyme activity was achieved at 100 nM concentration for all three compounds pursued. Hence, the present study provides strong evidence that these three compounds could serve as quorum sensing inhibitors of P. aeruginosa LasR protein and can be a probable candidate to treat Pseudomonas-associated infections.

Computational and Biological Evaluation of β-Adrenoreceptor Blockers as Promising Bacterial Anti-Virulence Agents

“…Significant π-π hydrophobic interaction with Tyr66 was also depicted stable for all sulphonamide antibacterial agents across the simulation studies. The above hydrogen bonding with Try58, Trp62, Asp75, and Ser129 was also considered significant for stabilizing a series of triphenyl-structured antagonists within the ligand binding domain of P. aeruginosa QS protein at preferentially higher binding energies than a synthetic triphenyl mimic super-inducer, TP-1 [55]. The triple aromatic-based pharmacophores of these ligands further permitted favored ligand anchoring within the binding site through face-to-face hydrophobic interactions with Tyr66, Trp63, Trp90, and/or Phe101 residues.…”

“…Moreover, residues around 80-90, 135-140, 150-165, 180-195, and 210-225 ranges were of the most flexible pattern (∆RMSF down to the highest negative values~−5.00 Å). Unlike the previously described QS proteins, the simulated CviR proteins depicted an extra stabilized residue region (45)(46)(47)(48)(49)(50)(51)(52)(53)(54)(55) near the N-terminus showing a significant immobility profile (∆RMSF~1.20 Å). Compounds 2, 11, 13, and 17 depicted high negative ∆RMSF values across the flexible regions; 180-195 and 210-225 residue range.…”

Repurposing α-Adrenoreceptor Blockers as Promising Anti-Virulence Agents in Gram-Negative Bacteria

“…Several FDA-approved sulfonamide and carboxamide-based antimicrobial analogues predicted favorable hydrogen bond pairing with Tyr58, Trp62, Asp75, and Ser129, as well as π–π hydrophobic interaction with Tyr66 within computational studies [ 56 ]. Finally, both in vitro LasR reporter gene evaluation and in silico investigations revealed potential QscR antagonistic activity of triphenyl-structured compounds, highlighting the ligand–protein binding with Try58, Trp62, Tyr66, Trp63, Asp75, Trp90, Phe101, and/or Ser129 amino acids [ 57 ].…”

The role of eugenol and ferulic acid as the competitive inhibitors of transcriptional regulator RhlR in P. aeruginosa

“…The ligand-binding domain selection box (grid-box) was generated by locating the residues of the AHL binding domain. For each transcriptional regulator was obtained different coordinates: LasR (X: 40.491, Y: 9.8455, and Z: 7.345 Å), QscR (X: -77.418, Y: -4.841 and Z: 9.181 Å) and RhlR (X: 4.83, Y: 64.94, and Z: 40.89 Å) [37] . The interaction of each transcriptional regulator with the corresponding functional groups from the aromatic compounds was described for the bonds formed between the ligand and each regulatory protein.…”