Integrative in silico and in vitro approaches to validate the efflux pump inhibitory potential of plant compounds targeting AdeABC efflux system of Acinetobacter baumannii.

Multi-drug resistance in Acinetobacter baumannii has emerged as a major public health threat, owing to its capability to cause severe, invasive infections, predominantly in hospital settings, resulting in high mortality and morbidity. Targeting Resistance Nodulation and Cell Division (RND) efflux pumps could be a prospective strategy to restore antibiotic efficacy against multidrug-resistant (MDR) bacterial infections. Therefore, in this study, we explored the efflux pump inhibitory potential of phytocompounds targeting AdeB (an RND protein) of Acinetobacter baumannii. The efflux pump inhibition potential of phytocompounds targeting AdeB (7KGE) was assessed using HTVS, MM/GBSA, and MD simulations. Among the 6,31,226 compounds docked against the AdeB protein, crocin and naringenin exhibited confined binding at proximal and distal binding sites of the drug binding pocket. Furthermore, detailed analysis of the AdeB switch loop identified key phenylalanine residues Phe136, Phe612, and Phe623 as critical contributors to substrate binding and the efflux transport mechanism. Furthermore, the efflux pump inhibitory potential of crocin and naringenin against selected strains of MDR A. baumannii was analysed. Furthermore, the synergistic potential of the phytocompounds with the antibiotic was evaluated. The results of the EtBr efflux assay, antibiotic accumulation assays, and checkerboard assay have affirmed the efflux pump inhibitory potential of naringenin. Lastly, the RT-qPCR data have revealed that naringenin has downregulated the expression of the AdeB gene (p < 0.05). By integrating structure-based ligand screening, molecular mechanics, molecular dynamics simulations, and experimental validation, the study confirmed that the interactions had contributed to efflux inhibition. Overall, naringenin had demonstrated the significant efflux pump inhibitory potential, thereby restoring the efficacy of antibiotics against multidrug-resistant pathogens.