Reduced glutathione levels in <i>Enterococcus faecalis</i> trigger metabolic and transcriptional compensatory adjustments during iron exposure

<i>Enterococcus faecalis,</i> a facultative anaerobic pathogen and common constituent of the gastrointestinal microbiota, must navigate varying iron levels within the host. This study explores its response to iron supplementation in a glutathione-deficient mutant strain (Δ<i>gsh</i>). We examined the transcriptomic and metabolic responses of a glutathione synthetase mutant strain (Δ<i>gsh</i>) exposed to iron supplementation, integrating these data into a genome-scale metabolic model (GSMM). Our results show that under glutathione deficiency, <i>E. faecalis</i> reduces intracellular iron levels and shifts its transcriptional response to prioritize energy production genes. Notably, basal metabolites, including arginine, increase. The GSMM highlights the importance of arginine metabolism, particularly the <i>arc</i> operon (anaerobic arginine catabolism), as a presumed compensatory mechanism for glutathione deficiency generated during iron exposure. These findings provide insights into how <i>E. faecalis</i> adjusts metal homeostasis and transcriptional/metabolic processes to mitigate the effects of oxidative stress caused by iron.IMPORTANCEIron is essential for bacterial survival, yet its excess can be harmful due to its role in increasing oxidative stress. <i>Enterococcus faecalis</i>, a common member of the human gut microbiota, must carefully balance its iron levels to survive in changing environments. Here, we investigate how <i>E. faecalis</i> compensates for the reduced availability of glutathione, a key antioxidant, when exposed to high iron concentrations. We discovered that <i>E. faecalis</i> lowers its intracellular iron levels when glutathione biosynthesis is disrupted and reprograms its metabolism to prioritize energy production, potentially to fuel stress response mechanisms under iron-induced oxidative conditions. These findings enhance our understanding of bacterial adaptation under oxidative stress and suggest that interfering with arginine metabolic pathways could represent novel strategies to combat <i>E. faecalis</i> infections.