Structural genes for salicylate biosynthesis from chorismate in Pseudomonas aeruginosa

Abstract

Salicylate is a precursor of pyochelin in Pseudomonas aeruginosa and both compounds display siderophore activity. To elucidate the salicylate biosynthetic pathway, we have cloned and sequenced a chromosomal region of P. aeruginosa PAO1 containing two adjacent genes, designated pchB and pchA, which are necessary for salicylate formation. The pchA gene encodes a protein of 52 kDa with extensive similarity to the chorismate-utilizing enzymes isochorismate synthase, anthranilate synthase (component I) and p-aminobenzoate synthase (component I), whereas the 11 kDa protein encoded by pchB does not show significant similarity with other proteins. The pchB stop codon overlaps the presumed pchA start codon. Expression of the pchA gene in P. aeruginosa appears to depend on the transcription and translation of the upstream pchB gene. The pchBA genes are the first salicylate biosynthetic genes to be reported. Salicylate formation was demonstrated in an Escherichia coli entC mutant lacking isochorismate synthase when this strain expressed both the pchBA genes, but not when it expressed pchB alone. By contrast, an entB mutant of E. coli blocked in the conversion of isochorismate to 2,3-dihydro-2,3-dihydroxybenzoate formed salicylate when transformed with a pchB expression construct. Salicylate formation could also be demonstrated in vitro when chorismate was incubated with a crude extract of P. aeruginosa containing overproduced PchA and PchB proteins; salicylate and pyruvate were formed in equimolar amounts. Furthermore, salicylate-forming activity could be detected in extracts from a P. aeruginosa pyoverdin-negative mutant when grown under iron limitation, but not with iron excess. Our results are consistent with a pathway leading from chorismate to isochorismate and then to salicylate plus pyruvate, catalyzed consecutively by the iron-repressible PchA and PchB proteins in P. aeruginosa.

Salicylic Acid Produced by the Rhizobacterium Pseudomonas aeruginosa 7NSK2 Induces Resistance to Leaf Infection by Botrytis cinerea on Bean

“…Pyochelin is not exclusively essential for ISR because the pyochelin-deficient mutant KMPCH induces ISR, but it might play a role in ISR by 7NSK2 and MPFM1. This hypothesis, however, cannot be tested with siderophore-deficient mutants, because SA is a precursor for pyochelin biosynthesis (32). The conclusion that SA and maybe also pyochelin are responsible for ISR induction could explain the role of these siderophores in the biocontrol of the root pathogen Pythium splendens in tomato (3), but this statement needs confirmation in tomato.…”

“…SA-deficient mutants were made by gene replacement (32). The suicide plasmid pME3356, which contains a mutated pchA SA biosynthetic gene, was mobilized from E. coli S17-1 to P. aeruginosa 7NSK2 and to the pyoverdin-negative mutant MPFM1.…”

“…In iron-limiting conditions, this strain produces three siderophores: pyoverdin, pyochelin (12), and SA (3). The latter is also a precursor for pyochelin biosynthesis (32). In studies with siderophore-deficient mutants on tomato plants, pyoverdin or pyochelin proved necessary for wild-type biocontrol levels of Pythium damping-off (3).…”

Ferripyochelin uptake genes are involved in pyochelin-mediated signalling in Pseudomonas aeruginosa

“…During pyochelin biosynthesis, chorismate is converted to salicylate, which is then coupled to two molecules of cysteine by a thiotemplate mechanism (Serino et al, 1995;Gaille et al, 2002;Reimmann et al, 1998;Quadri et al, 1999). Dha, a byproduct of the pathway, is formed by P. aeruginosa in only small amounts and consists of a single cysteine moiety coupled to salicylate (Serino et al, 1997).…”

Stereospecific recognition of pyochelin and enantio-pyochelin by the PchR proteins in fluorescent pseudomonads

“…This is low compared to the iron affinity of pyoverdine, the primary siderophore of P. aeruginosa (dissociation constant~10 24 M 21 at pH 7.0; Visca et al, 2007), but the iron affinity of pyochelin in aqueous solution at physiological pH is unknown and may be higher. Pyochelin is a condensation product of salicylate and two cysteinyl residues and its biosynthesis in P. aeruginosa requires proteins encoded by the operons pchDCBA and pchEFGHI (Serino et al, 1995(Serino et al, , 1997Reimmann et al, 1998Reimmann et al, , 2001. Pyochelin contains three chiral centres (C49, C20 and C40) and exists in solution as a mixture of two interconvertible stereoisomers ( Fig.…”