PvdP Is a Tyrosinase That Drives Maturation of the Pyoverdine Chromophore in Pseudomonas aeruginosa

Abstract

eThe iron binding siderophore pyoverdine constitutes a major adaptive factor contributing to both virulence and survival in fluorescent pseudomonads. For decades, pyoverdine production has allowed the identification and classification of fluorescent and nonfluorescent pseudomonads. Here, we demonstrate that PvdP, a periplasmic enzyme of previously unknown function, is a tyrosinase required for the maturation of the pyoverdine chromophore in Pseudomonas aeruginosa. PvdP converts the nonfluorescent ferribactin, containing two iron binding groups, into a fluorescent pyoverdine, forming a strong hexadentate complex with ferrous iron, by three consecutive oxidation steps. PvdP represents the first characterized member of a small family of tyrosinases present in fluorescent pseudomonads that are required for siderophore maturation and are capable of acting on large peptidic substrates.

PvdN Enzyme Catalyzes a Periplasmic Pyoverdine Modification

“…4). In a more recent study on PvdP, it was already suggested that PvdN, besides PvdM and PvdO, can only be involved in sidechain modifications, as PvdP promotes all steps of chromophore formation (8). Our study now shows that this suggestion was indeed correct.…”

“…Double crossover mutants were selected by successive counter-selection on sucrose-containing medium, and scar-and marker-less deletions were identified and confirmed by PCR and sequencing. We included a deletion of monocistronic pvdP as control, as PvdP has been biochemically confirmed to be essential for the fluorophore formation (8).…”

“…While significant progress has been made with PvdQ and PvdP (5,8,26), the function of any of the proteins encoded by the pvdMNO operon has remained unclear. As interposon mutagenesis suggested essential roles of these proteins for pyoverdine production, it was seemingly not possible to address the individual functions of these enzymes (6,10,11).…”

“…The cytoplasmic biosynthesis reactions are well established, but recently, the periplasmic maturation has gained interest. Of the five periplasmic enzymes, PvdM, PvdN, PvdO, PvdP, and PvdQ, which are found in all known pyoverdine-producing species, functions have been assigned so far only to PvdQ and PvdP, which are involved in a precursor deacylation step and the chromophore cyclization, respectively (5)(6)(7)(8). PvdQ has been identified as a potential novel drug target (5).…”

PvdO is required for the oxidation of dihydropyoverdine as the last step of fluorophore formation in Pseudomonas fluorescens

“…Ferribactins are transported into the periplasm in acylated form and deacylated thereafter by PvdQ (3)(4)(5). In the periplasm, a 5,6-dihydroxyquinoline fluorophore is generated from ferribactin and the conserved glutamic acid residue at the first position of the peptide is modified (6)(7)(8). Iron can then be chelated by the two hydroxyl groups of the fluorophore and four further ligands from side chains of the peptide moiety (9,10).…”