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A combinatorial approach to the structure elucidation of a pyoverdine siderophore produced by a Pseudomonas putida isolate and the use of pyoverdine as a taxonomic marker for typing P. putida subspecies

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Abstract

The structure of a pyoverdine produced by Pseudomonas putida, W15Oct28, was elucidated by combining mass spectrometric methods and bioinformatics by the analysis of non-ribosomal peptide synthetase genes present in the newly sequenced genome. The only form of pyoverdine produced by P. putida W15Oct28 is characterized to contain α-ketoglutaric acid as acyl side chain, a dihydropyoverdine chromophore, and a 12 amino acid peptide chain. The peptide chain is unique among all pyoverdines produced by Pseudomonas subspecies strains. It was characterized as –l-Asp-l-Ala-d-AOHOrn-l-Thr-Gly-c[l-Thr(O-)-l-Hse-d-Hya-l-Ser-l-Orn-l-Hse-l-Ser-O-]. The chemical formula and the detected and calculated molecular weight of this pyoverdine are: C65H93N17O32, detected mass 1624.6404 Da, calculated mass 1624.6245. Additionally, pyoverdine structures from both literature reports and bioinformatics prediction of the genome sequenced P. putida strains are summarized allowing us to propose a scheme based on pyoverdines structures as tool for the phylogeny of P. putida. This study shows the strength of the combination of in silico analysis together with analytical data and literature mining in determining the structure of secondary metabolites such as peptidic siderophores.

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Correspondence to Pierre Cornelis.

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Supplementary Figure 1

IEF gel image of pyoverdines produced by P. putida W15Oct28 and P. putida L1. Pyoverdine of P. putida W15Oct28 (left lane) shows white fluorescence under UV light due to the un-matured dihydropyoverdine chromophore, while the pyoverdine L1 shows blue fluorescence and two isoforms. (PPT 1276 kb)

Supplementary Figure 2

Amino acid analysis of fully hydrolyzed N-TFA-n-butylester derivates by GC/MS. (DOC 1155 kb)

Supplementary Fig. 3

Three dimensional structure model of P. putida W15Oct28 pyoverdine–Fe3+ complex based on the hexadentate character. Green: carbons, Red: oxygens, Blue: nitrogen, Brown: Iron ion, non-polar hydrogens not shown. (PPT 302 kb)

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Ye, L., Ballet, S., Hildebrand, F. et al. A combinatorial approach to the structure elucidation of a pyoverdine siderophore produced by a Pseudomonas putida isolate and the use of pyoverdine as a taxonomic marker for typing P. putida subspecies. Biometals 26, 561–575 (2013). https://doi.org/10.1007/s10534-013-9653-z

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