Biosynthesis of the Pseudomonas aeruginosa siderophore pyochelin: Peptide synthesis and tailoring.

摘要

When bacteria are in iron-limited environments they often respond with the production of low molecular weight iron chelators known as siderophores, with moderate (pyochelin) to good (mycobactins) to remarkable (enterobactin) affinities for scavenging FeIII. Siderophores fall into three general types, with hydroxamates (mycobactin, aerobactin), catechols (enterobactin, vibriobactin) or thiazoline/oxazoline (pyochelin, yersiniabactin) as iron-coordinating functional groups, but all three types are assembled by polyketide synthases (PKS) and/or nonribosomal peptide synthetases (NRPS). PKSs and NRPSs comprise a class of large multimodular enzymes that catalyze the synthesis of biopolymers (e.g. vancomycin, penicillins, cyclosporins, bleomycin); through the utilization of a "thiotemplate" mechanism, these multimodular/multienzyme systems generate diverse structures derived from acyl-coenzyme A or amino acid monomers. New natural products can be generated by introducing variability into the structure through the incorporation of various monomers and the action of a diverse array of tailoring enzymes. Identification and characterization of novel peptide synthetases and tailoring enzymes allows for a better understanding of how NRPSs systems generate structural diversity. The opportunistic pathogen Pseudomonas aeruginosa makes both a high affinity peptide hydroxamate, pyoverdin and a low affinity peptide bithiazoline, pyochelin, as siderophores that contribute to pathogenicity.; Pyochelin (Pch), the tricyclic iron chelating siderophore produced as a virulence factor by Pseudomonas aeruginosa, is encoded by the NRPS Pch synthetase. The Pch system encodes for many intriguing catalytic steps including: (1) the heterocyclization of N-acyl-cysteinyl-S-enzyme intermediates to thiazolinyl rings; (2) the epimerization of the first thiazoline ring; (3) the regioselective redox adjustment of the terminal thiazoline of Pch to thiazolidine; and (4) the introduction of an N-methyl group on the terminal thiazoline ring. This work explores each of these modifications. We have fully reconstituted the pyochelin synthetase assigning all 14 chemical steps to their respective catalytic domains and have characterized three novel thiazoline tailoring activities found in PchE (epimerization), PchF (methylation), and PchG (reduction).