Nucleotide sequence of pvdD, a pyoverdine biosynthetic gene from Pseudomonas aeruginosa: PvdD has similarity to peptide synthetases

Abstract

Pseudomonas aeruginosa secretes a fluorescent siderophore, pyoverdine, when grown under iron-deficient conditions. Pyoverdine consists of a chromophoric group bound to a partly cyclic octapeptide. As a step toward understanding the molecular events involved in pyoverdine synthesis, we have sequenced a gene, pvdD, required for this process. The gene encodes a 2,448-residue protein, PvdD, which has a predicted molecular mass of 273,061 Da and contains two highly similar domains of about 1,000 amino acids each. The protein is similar to peptide synthetases from a range of bacterial and fungal species, indicating that synthesis of the peptide moiety of pyoverdine proceeds by a nonribosomal mechanism. The pvdD gene is adjacent to a gene, fpvA, which encodes an outer membrane receptor protein required for uptake of ferripyoverdine.Pseudomonas aeruginosa is an opportunistic pathogen which infects injured, immunodeficient, or otherwise compromised patients. The bacteria secrete a siderophore, pyoverdine, which is likely to play an important role in infection by competing with transferrin for iron in order to overcome the iron-withholding defense mechanism present in mammals (3,11,42). Almost all isolates of P. aeruginosa from infected patients secrete pyoverdine (9,22,29), and it has been shown that pyoverdine is present in the sputa of cystic fibrosis patients infected with P. aeruginosa (21). A mutant of P. aeruginosa which is unable to synthesize pyoverdine showed reduced virulence in an animal model of infection (24).Pyoverdine from P. aeruginosa PAO is a water-soluble, yellow-green fluorescent compound. It consists of a dihydroxyquinoline chromophoric group linked to an eight-residuevia the N-terminal serine, with a small dicarboxylic acid attached to the chromophore. Iron complexation is thought to occur through oxygen atoms present on the dihydroxyquinoline and two hydroxamate units supplied by the L-N 5 -OH-Orn residues. Several Pseudomonas species produce similar compounds, variously called pyoverdines or pseudobactins, all of which have the same dihydroxyquinoline group but differ in the nature of the attached peptide (reviewed in references 1 and 7), and it is likely that all of these are synthesized by similar mechanisms. Synthesis of the chromophoric group begins with condensation of D-tyrosine and L-2,4-diaminobutyric acid (7), with glutamic acid being the precursor of the amide-linked dicarboxylic acid (50). It has been suggested that biosynthesis of the peptide moiety of pyoverdine occurs by a nonribosomal mechanism (30).Little is known about the molecular nature of enzymes involved in the biosynthesis of pyoverdine in P. aeruginosa or any other pseudomonad. Recently, pvdA, which encodes an enzyme (L-ornithine N 5 -oxygenase) responsible for catalyzing the hydroxylation of ornithine, an early step in pyoverdine biosynthesis in P. aeruginosa, has been characterized at the molecular level (55). The pbsC gene, which is involved in synthesis of pseudobactin by Pseudomonas sp. strain M114, has also been seque...

Identification and characterization of novel pyoverdine synthesis genes in Pseudomonas aeruginosa

“…These genes are thought to encode peptide synthetases (Lehoux et al, 2000;Merriman et al, 1995), a family of proteins with many conserved sequence features . PvdD directs incorporation of two L-threonine residues into pyoverdine PAO (Ackerley et al, 2003) and pvdI and pvdJ are very likely to encode peptide synthetases that direct incorporation of the remaining six amino acids into the peptide of type I pyoverdines.…”

Co-ordination of iron acquisition, iron porphyrin chelation and iron–protoporphyrin export via the cytochrome c biogenesis protein CcmC in Pseudomonas fluorescens

“…PVDs are composed of a conserved dihydroxyquinoline chromophore, a variable peptide chain, comprising 6-12 amino acids, specific to a producing strain, and a side-chain, generally a dicarboxylic acid or an amide (Ravel & Cornelis, 2003). Both the chromophore and the peptide chain of PVDs are synthesized by non-ribosomal peptide synthetases (Merriman et al, 1995;Lehoux et al, 2000;Mossialos et al, 2002). Ferribactin, the precursor of PVD, is probably exported to the periplasm where it is matured into PVD with a chromophore .…”

Decreasing the Level of Ethyl Acetate in Ethanolic Fermentation Broths ofEscherichia coliKO11 by Expression ofPseudomonas putida estZEsterase

“…1). The third ORF (fpvA) was identified as a ferric pyoverdine receptor protein (39% identity with FpvA from P. aeruginosa PAO1) (32). Based on protein sequence similarity, the fourth ORF (pvdE) appears to encode the C-terminal portion of an ATPdependent ABC membrane transporter associated with pyoverdine biosynthesis (66% identity with PvdE in P. aeruginosa PAO1) (31).…”

“…These ORFs were identified by comparison to other known genes by using Blast (www .ncbi.nlm.nih.gov). One of the ORFs contained the C-terminal 594 amino acids of a siderophore synthase (nonribosomal peptide synthesis) resembling PvdD in P. aeruginosa PAO1 (32) and P. putida WCS358 (GenBank accession no. CAC32046), with 61 and 62% identity, respectively.…”