title: In vitro-binding of the natural siderophore enantiomers pyochelin and enantiopyochelin to their AraC-type regulators PchR in Pseudomonas.
abstract: The enantiomeric siderophores pyochelin and enantiopyochelin of Pseudomonas aeruginosa and Pseudomonas protegens promote growth under iron limitation and activate transcription of their biosynthesis and uptake genes via the AraC-type regulator PchR. Here we investigated siderophore binding to PchR in vitro using fluorescence spectroscopy. A fusion of the N-terminal domain of P. aeruginosa PchR with maltose binding protein (MBP-PchR'PAO) bound iron-loaded (ferri-) pyochelin with an affinity (Kd) of 41 ± 5 μM. By contrast, no binding occurred with ferri-enantiopyochelin. Stereospecificity of a similar fusion protein of the P. protegens PchR (MBP-PchR'CHA0) was less pronounced. The Kd's of MBP-PchR'CHA0 for ferri-enantiopyochelin and ferri-pyochelin were 24 ± 5 and 40 ± 7 μM, respectively. None of the proteins interacted with the iron-free siderophore enantiomers, suggesting that transcriptional activation by PchR occurs only when the respective siderophore actively procures iron to the cell.
authors.affiliation: Département de Microbiologie Fondamentale, Université de Lausanne, Bâtiment Biophore, Quartier UNIL-Sorge, 1015, Lausanne, Switzerland.
authors.forename: Po-Chi
authors.initials: PC
authors.lastname: Lin
authors.forename: Zeb A
authors.initials: ZA
authors.lastname: Youard
authors.forename: Cornelia
authors.initials: C
authors.lastname: Reimmann
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D007501:Iron / Q000378:metabolism*; D007700:Kinetics; D058619:Maltose-Binding Proteins / Q000235:genetics / Q000378:metabolism; D010636:Phenols / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D011549:Pseudomonas / Q000235:genetics / Q000378:metabolism*; D011550:Pseudomonas aeruginosa / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017262:Siderophores / Q000737:chemistry / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013237:Stereoisomerism; D013844:Thiazoles / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D058619:Maltose-Binding Proteins; D010636:Phenols; D011993:Recombinant Fusion Proteins; D017262:Siderophores; D013844:Thiazoles; D014157:Transcription Factors; C083187:pchR protein, Pseudomonas aeruginosa; C025316:pyochelin; D007501:Iron