RSC Advances | Issue 36, 2025 | https://doi.org/10.1039/D5RA03958D
Kiran Harikumar, Udisha Singh,Subakamakshi Krishnaswamy Ramaraj, Lakshmi Ajayakumar Rekha, Adyasha Nayak, Poulomi Ghosh, Roshny Prasad, Alwin Philip, Saloo Sahuab and Shijulal Nelson-Sathi *
Antibiotic resistance in Gram-positive priority pathogens is mediated by a diverse set of mechanisms such as target protection, antibiotic inactivation, decreased uptake, antibiotic efflux, etc. In Staphylococcus aureus, efflux pumps of the major facilitator superfamily (MFS) expel various antibiotics and multiple efflux pumps are activated upon antibiotic exposure. Efflux pump inhibitors (EPIs) that can act as antibiotic adjuvants are proposed to be promising solutions to tackle antibiotic resistance. In this study, in silico screening of 17,967 phytochemical compounds from Indian medicinal plants (IMPPAT 2.0) against four key MFS efflux pumps activated by fluoroquinolone exposure (NorA, NorB, NorC, and SdrM) followed by in vitro validation identified a tannin derivative, pentagalloyl glucose (PGG), as a potential efflux pump inhibitor (EPI) with high binding affinity. Molecular docking scores (≤−16.383 kcal/mol) and MM/GBSA binding affinities (≤−100.62 kcal/mol) indicate a strong interaction between PGG and its targeted efflux pumps. PGG forms stable interactions via hydrogen bonding with key residues of NorA, including GLU222 and ASP307, which are crucial for proton-coupled transport. Likewise, it interacts with essential residues in NorB (SER147, ASN280), NorC (ASN276, LYS398), and SdrM (SER143, GLN283), forming strong hydrogen bonds that contribute to its inhibitory potential. The stability of PGG-bound complexes was confirmed through molecular dynamics simulations over 100 ns in triplicates, along with free energy landscape (FEL) and principal component analysis (PCA). Furthermore, PGG's synergistic action with ciprofloxacin, and effects on S. aureus growth dynamics were validated using the checkerboard assay, and time-kill kinetic studies, respectively. Following further structural optimization and in vivo studies, PGG can be considered a promising therapeutic candidate against multidrug-resistant S. aureus strains.
