3 research outputs found

    Inhibiting the BfrB:Bfd interaction in: Pseudomonas aeruginosa causes irreversible iron accumulation in bacterioferritin and iron deficiency in the bacterial cytosol

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    © The Royal Society of Chemistry 2017. Iron is an essential nutrient for bacteria but the reactivity of Fe2+ and the insolubility of Fe3+ present significant challenges to bacterial cells. Iron storage proteins contribute to ameliorating these challenges by oxidizing Fe2+ using O2 and H2O2 as electron acceptors, and by compartmentalizing Fe3+. Two types of iron-storage proteins coexist in bacteria, the ferritins (Ftn) and the heme-containing bacterioferritins (Bfr), but the reasons for their coexistence are largely unknown. P. aeruginosa cells harbor two iron storage proteins (FtnA and BfrB), but nothing is known about their relative contributions to iron homeostasis. Prior studies in vitro have shown that iron mobilization from BfrB requires specific interactions with a ferredoxin (Bfd), but the relevance of the BfrB:Bfd interaction to iron homeostasis in P. aeruginosa is unknown. In this work we explore the repercussions of (i) deleting the bfrB gene, and (ii) perturbing the BfrB:Bfd interaction in P. aeruginosa cells by either deleting the bfd gene or by replacing the wild type bfrB gene with a L68A/E81A double mutant allele in the P. aeruginosa chromosome. The effects of the mutations were evaluated by following the accumulation of iron in BfrB, analyzing levels of free and total intracellular iron, and by characterizing the ensuing iron homeostasis dysregulation phenotypes. The results reveal that P. aeruginosa accumulates iron mainly in BfrB, and that the nutrient does not accumulate in FtnA to detectable levels, even after deletion of the bfrB gene. Perturbing the BfrB:Bfd interaction causes irreversible flow of iron into BfrB, which leads to the accumulation of unusable intracellular iron while severely depleting the levels of free intracellular iron, which drives the cells to an acute iron starvation response despite harboring normal levels of total intracellular iron. These results are discussed in the context of a dynamic equilibrium between free cytosolic Fe2+ and Fe3+ compartmentalized in BfrB, which functions as a buffer to oppose rapid changes of free cytosolic iron. Finally, we also show that P. aeruginosa cells utilize iron stored in BfrB for growth in iron-limiting conditions, and that the utilization of BfrB-iron requires a functional BfrB:Bfd interaction

    Small Molecule Inhibitors of the Bacterioferritin (BfrB)-Ferredoxin (Bfd) Complex Kill Biofilm-Embedded Pseudomonas aeruginosa Cells

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    © 2020 American Chemical Society. Bacteria depend on a well-regulated iron homeostasis to survive adverse environments. A key component of the iron homeostasis machinery is the compartmentalization of Fe3+ in bacterioferritin and its subsequent mobilization as Fe2+ to satisfy metabolic requirements. In Pseudomonas aeruginosa Fe3+ is compartmentalized in bacterioferritin (BfrB), and its mobilization to the cytosol requires binding of a ferredoxin (Bfd) to reduce the stored Fe3+ and release the soluble Fe2+. Blocking the BfrB-Bfd complex in P. aeruginosa by deletion of the bfd gene triggers an irreversible accumulation of Fe3+ in BfrB, concomitant cytosolic iron deficiency and significant impairment of biofilm development. Herein we report that small molecules developed to bind BfrB at the Bfd binding site block the BfrB-Bfd complex, inhibit the mobilization of iron from BfrB in P. aeruginosa cells, elicit a bacteriostatic effect on planktonic cells, and are bactericidal to cells embedded in mature biofilms
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