Exploring the Structural and Functional role of α,β-unsaturated Ketoesters as Anti-Staphylococcal Agents Targeting Glutathione Peroxidase

Multiple-drug resistant (MDR) Staphylococcus aureus strains (like methicillin-resistant S. aureus or MRSA) uses an arsenal of antioxidant enzymes to mitigate host-induced oxidative stress. Among them the non-canonical Staphylococcal glutathione peroxidase (SaGpx) plays a crucial role in bacterial redox homeostasis by reducing peroxides via thioredoxin-dependent pathways. Thus, enabling oxidative stress mitigation during host infection. Despite its importance in S. aureus , its role in bacterial pathogenesis remains unexplored. This study aimed to elucidate the possible role of SaGpx in Staphylococcal virulence. First, we determined the high-resolution crystal structure of SaGpx (at 1.65 Å resolution) using X-ray crystallography. Guided by the catalytic cleft architecture of SaGpx, small-molecule based inhibitors were then rationally designed and synthesized. These inhibitors exhibited good binding affinity to SaGpx and complete enzymatic blockade. These inhibitors exhibited potent anti- S. aureus activity (MICs 6.25-31.25 μM) along with no cytotoxicity in L929 fibroblast wound-healing assays. Furthermore, the in vivo antibacterial ability of these inhibitors was evaluated using S. aureus -infected skin wound mouse model, where these compounds show potent antibacterial and wound healing ability supported by subsequent histological as well as immunohistochemical analysis. These findings suggest SaGpx as a possible virulence determinant in S. aureus and position these synthesized inhibitors as promising antivirulence therapeutics.