Project

Back to overview

Enantio-pyochelin: Biosynthesis, regulation, and natural occurrence

English title Enantio-pyochelin: Biosynthesis, regulation, and natural occurrence
Applicant Reimmann Cornelia
Number 132998
Funding scheme Project funding
Research institution Département de Microbiologie Fondamentale Faculté de Biologie et de Médecine Université de Lausanne
Institution of higher education University of Lausanne - LA
Main discipline Experimental Microbiology
Start/End 01.12.2010 - 31.07.2014
Approved amount 288'000.00
Show all

Keywords (7)

siderophore; pyochelin; enantio-pyochelin; Pseudomonas; Burkholderia; stereospecificity; Chirality

Lay Summary (English)

Lead
Lay summary
Bacterial siderophores (iron chelators) are essential for growth under iron limitation and often qualify as virulence factors. Their production is tightly regulated to assure that they are made only when needed and to avoid a deleterious iron accumulation in the cell. In the Gram-negative opportunistic human pathogen Pseudomonas aeruginosa the expression of the biosynthesis genes for the siderophore pyochelin requires the transcriptional regulator PchR together with pyochelin, which acts as an effector of PchR. Similarly enantio-pyochelin, the optical antipode of pyochelin made by certain plant-beneficial strains of Pseudomonas fluorescens, induces its biosynthesis genes by activating a PchR homolog. The interaction of PchR with its siderophore is stereospecific, meaning that PchR of P. aeruginosa is not activated by enantio-pyochelin, and pyochelin cannot activate PchR of P. fluorescens. Genetic experiments indicate that siderophore recognition occurs in the N-terminal domain of PchR. The objectives of this project are(i) to understand how the different stereochemistry in pyochelin/enantio-pyochelin is generated during biosynthesis(ii) to develop an in vitro assay to investigate binding of the siderophores pyochelin and enantio-pyochelin to their respective PchR proteins(ii) to study the natural occurrence of pyochelin and enantio-pyochelin in pseudomonads and class 2 members of the Burkholderia cepacia complexSignificanceThis project will allow us to understand the importance of chirality in siderophore-mediated iron acquisition.
Direct link to Lay Summary Last update: 21.02.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
Functional analysis of pyochelin/enantiopyochelin-related genes from a pathogenicity island of Pseudomonas aeruginosa strain PA14
Maspoli Alessandro, Wenner Nicolas, Mislin Gaetan L. A., Reimmann Cornelia (2014), Functional analysis of pyochelin/enantiopyochelin-related genes from a pathogenicity island of Pseudomonas aeruginosa strain PA14, in Biometals, 27(3), 559-573.
In vitro-binding of the natural siderophore enantiomers pyochelin and enantiopyochelin to their AraC-type regulators PchR in Pseudomonas
Lin Po-Chi, Youard Zeb A., Reimmann Cornelia (2013), In vitro-binding of the natural siderophore enantiomers pyochelin and enantiopyochelin to their AraC-type regulators PchR in Pseudomonas, in Biometals, 26(6), 1067-1073.
Inner-membrane transporters for the siderophores pyochelin in Pseudomonas aeruginosa and enantio-pyochelin in Pseudomonas fluorescens display different enantioselectivities
Reimmann Cornelia (2012), Inner-membrane transporters for the siderophores pyochelin in Pseudomonas aeruginosa and enantio-pyochelin in Pseudomonas fluorescens display different enantioselectivities, in Microbiology, 158(5), 1317-1324.
Iron acquisition with the natural siderophore enantiomers pyochelin and enantio-pyochelin in Pseudomonas species
Youard Zeb A, Wenner Nicolas, Reimmann Cornelia (2011), Iron acquisition with the natural siderophore enantiomers pyochelin and enantio-pyochelin in Pseudomonas species, in Biometals, 24(3), 513-522.
Pyochelin enantiomers and their outer-membrane siderophore transporters in fluorescent pseudomonads: structural bases for unique enantiospecific recognition
Brillet Karl, Reimmann Cornelia, Mislin Gaëtan L. A., Noël Sabrina, Rognan Didier, Schalk Isabelle J., Cobessi David (2011), Pyochelin enantiomers and their outer-membrane siderophore transporters in fluorescent pseudomonads: structural bases for unique enantiospecific recognition, in Journal of the American Chemical Society, 133(41), 16503-16509.
A cell biological view of the siderophore pyochelin iron uptake pathway in Pseudomonas aeruginosa
Cunrath Olivier, Gasser Véronique, Hoegy Françoise, Reimmann Cornelia, Guillon Laurent, Schalk Isabelle J., A cell biological view of the siderophore pyochelin iron uptake pathway in Pseudomonas aeruginosa, in Environmental Microbiology.

Collaboration

Group / person Country
Types of collaboration
Université de Strasbourg-CNRS France (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Université Joseph Fourier, CEA, CNRS France (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
72nd Annual Assembly of the SSM Poster Functional analysis of pyochelin/enantiopyochelin-related genes from a pathogenicity island of Pseudomonas aeruginosa strain PA14 19.06.2014 Fribourg, Switzerland Reimmann Cornelia; Maspoli Alessandro;
14th International Conference on Pseudomonas Poster A genomic island of Pseudomonas aeruginosa PA14 with pyochelin/enantiopyochelin biosynthesis genes 07.09.2013 Lausanne, Switzerland Reimmann Cornelia; Maspoli Alessandro;
Congress Biometals 2012 Talk given at a conference Iron uptake with pyochelin in Pseudomonas 15.07.2012 Brussels, Belgium Reimmann Cornelia;
70th Annual Assembly of the SSM Poster Siderophore binding to the transcriptional regulator PchR from Pseudomonas aeruginosa and Pseudomonas fluorescens 21.06.2012 St. Gallen, Switzerland Lin Po-Chi; Reimmann Cornelia;
4th Congress of European Microbiologists, FEMS 2011 Talk given at a conference Iron transport across the inner membrane with the siderophores pyochelin and enantio-pyochelin in Pseudomonas species 26.06.2011 Geneva, Switzerland, Switzerland Reimmann Cornelia;


Associated projects

Number Title Start Funding scheme
102174 Role and regulation of the Pseudomonas aeruginosa siderophore pyochelin and its uptake system 01.01.2004 Project funding
113955 Pyochelin-mediated signalling in Pseudomonas 01.04.2007 Project funding

Abstract

Background. The bacterial siderophore pyochelin consists of salicylate and two cysteine-derived heterocycles, the second of which is modified by reduction and N-methylation during biosynthesis. In Pseudomonas aeruginosa the first cysteine residue is converted to its D-isoform during thiazoline ring formation whereas the second cysteine remains in its L-configuration. Stereochemistry is opposite in the Pseudomonas fluorescens siderophore enantio-pyochelin, in which the first ring originates from L-cysteine and the second ring from D-cysteine. Both siderophores promote growth of the producer organism during iron limitation and induce the expression of their biosynthesis genes by activating the transcriptional AraC-type regulator PchR. However, neither siderophore is functional as an iron carrier or as a transcriptional inducer in the other species, demonstrating that both processes are highly stereospecific. Stereospecificity of pyochelin/enantio-pyochelin-mediated iron uptake is conferred by the specificity of the outer membrane receptors towards their cognate siderophore, while stereospecificity in transcriptional activation by pyochelin/enantio-pyochelin is based on the specificity of the two PchR proteins for their respective siderophore effector.Broader scientific context. The inability of the two enantiomers to function in the other species demonstrates the importance of chirality in siderophore-mediated iron acquisition, an aspect which has not been considered so far. By analogy with the vast structural variety of pyoverdine siderophores made by fluorescent pseudomonads, we speculate that stereochemical variation of siderophores represents a defence strategy to prevent siderophore piracy among bacteria occupying the same ecological niche. Research project. We propose to better characterize the following aspects of siderophore chirality in pyochelin/enantio-pyochelin-mediated iron acquisition.• Enantio-pyochelin biosynthesis. We will investigate how the different stereochemistry in pyochelin/enantio-pyochelin is generated during biosynthesis. In particular, we will investigate which enzyme is responsible for the epimerization of the second cysteine in enantio-pyochelin. This will be done by reconstituting the synthesis of enantio-pyochelin with purified proteins in vitro to identify pathway intermediates and endproducts. • Siderophore binding to PchR. We will develop an in vitro binding assay based on isothermal titration calorimetry to demonstrate and quantify binding of pyochelin and enantio-pyochelin to the N-terminal domain of their respective PchR protein. We will test whether binding is stereospecific and compare the binding affinities of iron-free and iron-loaded siderophores to see if iron is important for the interaction. • Pyochelin and enantio-pyochelin production in Pseudomonas and Burkholderia spp. We will study the natural occurrence of pyochelin and enantio-pyochelin in pseudomonads and class 2 members of the Burkholderia cepacia complex. This will be done by a combination of two biotests which are based on the stereospecificity of iron uptake and transcriptional activation, and by chiral HPLC. For a more rapid discrimination between pyochelin and enantio-pyochelin producers, a PCR-based approach will be developed and validated.Experimental design. The necessary biological material is already available in our department or will be provided by other research groups. Molecular and biochemical methods used to carry out this project are well established in our department. Where we lack expertise or a special equipment, we will collaborate with experts in the field as indicated in the text.Significance. This project will unravel the biosynthesis of the novel siderophore enantio-pyochelin, demonstrate its binding to an AraC-type regulator, and reveal its distribution in the microbial world.
-