Synthesis of the siderophore pyoverdine in Pseudomonas aeruginosa involves a periplasmic maturation
Pyoverdines, the main siderophores produced by fluorescent Pseudomonads, comprise a fluorescent dihydroxyquinoline chromophore attached to a strain-specific peptide. These molecules are thought to be synthesized as non-fluorescent precursor peptides that are then modified to give functional pyoverdines. Using the fluorescent properties of PVDI, the pyoverdine produced by Pseudomonas aeruginosa PAO1, we were able to show that PVDI was not present in the cytoplasm of the bacteria, but large amounts of a fluorescent PVDI precursor PVDIp were stored in the periplasm. Like PVDI, PVDIp is able to transport iron into P. aeruginosa cells. Mutation of genes encoding the periplasmic PvdN, PvdO and PvdP proteins prevented accumulation of PVDIp in the periplasm and secretion of PVDI into the growth medium, indicating that these three enzymes are involved in PVDI synthesis. Mutation of the gene encoding PvdQ resulted in the presence of fluorescent PVDI precursor in the periplasm and secretion of a functional fluorescent siderophore that had different isoelectric properties to PVDI, suggesting a role for PvdQ in the periplasmic maturation of PVDI. Mutation of the gene encoding the export ABC transporter PvdE prevented PVDI production and accumulation of PVDIp in the periplasm. These data are consistent with a model in which a PVDI precursor peptide is synthesized in the cytoplasm and exported to the periplasm by PvdE where siderophore maturation, including formation of the chromophore moiety, occurs in a process involving the PvdN, PvdO, PvdP and PvdQ proteins.
KeywordsSiderophore biosynthesis Pyoverdine Iron uptake Fluorescence Fluorescent peptide Pseudomonasaeruginosa
This work was supported by the Centre National de la Recherche Scientifique (CNRS) and by the association Mucoviscidose ABCF. E. Yeterian had a fellowship from the French Ministère de la Recherche et de la Technologie. The authros are grateful to Keith Poole for providing plasmid pJSS2, to Karla Mettrick for constructing the fpvA and fpvA fpvR mutant strains and to Dr Gaëtan L. A. Mislin for mass spectrum analyses and discussion.
- Abdallah MA, Pattus F (2000) Siderophores and iron-transport in microorganisms. J Chin Chem Soc 47:1–20Google Scholar
- Chuanchuen R, Narasaki CT, Schweizer HP (2002) Benchtop and microcentrifuge preparation of Pseudomonas aeruginosa competent cells. Biotechniques 33(760):762–763Google Scholar
- Mossialos D, Ochsner U, Baysse C, Chablain P, Pirnay JP, Koedam N, Budzikiewicz H, Fernandez DU, Schafer M, Ravel J, Cornelis P (2002) Identification of new, conserved, non-ribosomal peptide synthetases from fluorescent pseudomonads involved in the biosynthesis of the siderophore pyoverdine. Mol Microbiol 45:1673–1685CrossRefPubMedGoogle Scholar
- Sambrook J, Russel DW (2001) Molecular cloning: a laboratory manual. Cold Spring Harbor, NYGoogle Scholar
- Visca P (2004) Iron regulation and siderophore signalling in virulence by Pseudomonas aeruginosa. In: Juan-Luis R (ed) Pseudomonas, vol 2. Kluwer/Plenum Publishers, New York, pp 69–123Google Scholar