Recycling of pyoverdin on the FpvA receptor after ferric pyoverdin uptake and dissociation in Pseudomonas aeruginosa.
Under iron-limiting conditions, Pseudomonas aeruginosa secretes a fluorescent siderophore called pyoverdin (PaA), which, after complexing iron, is transported back into the cells via its outer membrane receptor FpvA. The recent finding that all FpvA receptors on the bacterial cell surface are loaded with iron-free PaA under iron limiting conditions has raised questions about the mechanism by which P. aeruginosa transports efficiently iron. We used [(3)H]PaA' [(55)Fe]PaA-Fe, and a kinetically stable chromium-PaA complex to show that iron loading of the receptor occurs through a siderophore displacement mechanism in vivo. Moreover, the fluorescence properties of iron-free PaA revealed that, after PaA-Fe uptake and dissociation, the PaA molecule is recycled into the extracellular medium. We used fluorescence resonance energy transfer (FRET) between the PaA chromophore and the FpvA tryptophans in vivo to monitor the kinetics of PaA displacement by PaA-Fe at the cell surface, the dissociation of iron from the siderophore, and the recycling of PaA back to the receptor on the outer membrane of the bacteria in real time. The loading status of FpvA (PaA versus PaA-Fe) was shown to depend on the relative concentration of the two forms of pyoverdin in the growth medium.
Schalk
Isabelle J
IJ
Département des Récepteurs et Protéines Membranaires, UPR 9050, CNRS, ESBS, Bld Sébastien Brant, F-67 400 Illkirch, Strasbourg, France. schalk@esbs.u-strasbg.fr
Abdallah
Mohamed A
MA
Pattus
Franc
F
D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D002857:Chromium / Q000378:metabolism; D004735:Energy Transfer / Q000235:genetics; D007501:Iron / Q000378:metabolism; D046911:Macromolecular Substances; D009842:Oligopeptides; D010860:Pigments, Biological / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011550:Pseudomonas aeruginosa / Q000235:genetics / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D017262:Siderophores / Q000235:genetics / Q000378:metabolism*; D013050:Spectrometry, Fluorescence / Q000379:methods
D001425:Bacterial Outer Membrane Proteins; C082326:FpvA protein, Pseudomonas aeruginosa; D046911:Macromolecular Substances; D009842:Oligopeptides; D010860:Pigments, Biological; D011956:Receptors, Cell Surface; D017262:Siderophores; C035902:siderophore receptors; D002857:Chromium; C042453:pyoverdin; D007501:Iron