OLIVE POMACE-DERIVED PHYTOCHEMICALS AS MODULATORS OF BACTERIAL EFFLUX PUMP RESISTANCE: AN INTEGRATED NETWORK PHARMACOLOGY, MOLECULAR DOCKING, AND MOLECULAR DYNAMICS APPROACH

Abstract

Background: Antimicrobial resistance (AMR) represents one of the most formidable threats to global public health, with over 4.71 million deaths associated with bacterial AMR recorded in 2021 alone. Efflux pump-mediated resistance, particularly via the AcrAB-TolC and MexAB-OprM systems belonging to the Resistance-Nodulation-Division (RND) superfamily, constitutes a primary mechanism of multidrug resistance in ESKAPE pathogens. Olive pomace, a voluminous agro-industrial by-product, is enriched with bioactive phenolic compounds including hydroxytyrosol (HT) and oleuropein (OLE), which possess documented antimicrobial properties; however, their potential as efflux pump inhibitors (EPIs) has not been systematically evaluated through integrated computational approaches. Methods: Network pharmacology analysis was employed to identify shared molecular targets between olive pomace phytochemicals and AMR-associated pathways. Molecular docking simulations against AcrB (PDB: 4DX5) and MexB (PDB: 2V50) were performed using AutoDock Vina, followed by 100 ns molecular dynamics (MD) simulations using OpenMM to assess binding stability. ADMET profiling was conducted using pkCSM and SwissADME. Results: HT and OLE demonstrated significant binding affinities toward the deep binding pocket of AcrB, with estimated ΔG values of −6.8 and −9.2 kcal/mol, respectively, outperforming the reference EPI phenylalanine-arginine β-naphthylamide (PAβN). MD simulations confirmed stable protein-ligand complexes over 100 ns, evidenced by low RMSD values (<2 Å). ADMET analysis revealed favourable drug-likeness profiles for HT. Conclusion: This study provides the first comprehensive computational evidence for HT and OLE as promising natural efflux pump inhibitors derived from olive agro-waste, offering a sustainable strategy to re-sensitise MDR ESKAPE pathogens to conventional antibiotics. These findings warrant experimental validation through in vitro synergy assays.