Network-guided functional enrichment and interface-focused prioritization of quorum sensing targets in Pseudomonas aeruginosa for anti-virulence drug discovery

Pseudomonas aeruginosa persistence is driven by quorum sensing (QS) that coordinates biofilm formation and virulence. Single-node QS inhibitors, often focused on LasR, underperform clinically. We built a statistically supported QS–virulence interactome (STRING 11.5, BioGRID, IntAct; combined score ≥0.9, non–text-mining evidence) and prioritized targets using centrality and clustering metrics in Cytoscape. Functional enrichment (GO/KEGG/InterPro/Pfam) and infection-stage transcriptomes (GSE119103, GSE99816) were used to validate biological relevance. Structure-based screening of FDA drugs and natural products was performed with Glide; top poses underwent 100-ns MD (OPLS4, 310 K, 1 atm), with RMSD/RMSF, hydrogen-bond occupancy and MM/GBSA used to assess stability. PqsE and rhlR ranked in the top 5% for degree and betweenness, indicating non-redundant bottlenecks within a densely connected network (empirical PPI enrichment p=3.1 × 10⁻⁴). Enrichment highlighted QS, phenazine biosynthesis and biofilm formation. Docking and MD identified honokiol—a low-toxicity biphenyl—as an interface-modulating scaffold at the PqsE–RhlR complex. Honokiol engaged a shallow, hydrophobic pocket with a single persistent H-bond (ASP196) and hydrophobic contacts (LYS70, TYR72, PHE195), producing ligand relocation consistent with adaptive, allosteric binding. Although pimozide scored stronger in docking, its CNS liabilities make honokiol the safer lead. Prioritizing network bottlenecks and protein–protein interfaces reveal the PqsE–RhlR axis as a resistance-robust QS target. Honokiol destabilizes the interface without bactericidal pressure, nominating an interface-focused route for anti-virulence design against P. aeruginosa and related ESKAPE pathogens. A reproducible workflow advances interface-specific QS inhibition and points to honokiol-like scaffolds as clinically tractable leads.