GNPS - Iron titration into siderophore standards - negative ionization
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Q Exactive
ORGANISM(S): Not Applicable
SUBMITTER: 
Pieter C. Dorrestein
PROVIDER: MSV000085352 | GNPS | Wed Apr 29 19:45:00 BST 2020
REPOSITORIES: GNPS
Similar Datasets
Project description:Iron was titrated into 5 commercial siderophore standards to calculate dissociation constants.
2020-03-14 | MSV000085102 | GNPS
Project description:Competition for limited iron resources is a key driver of microbial community structure in many regions of the surface ocean. The bacterial siderophores ferrioxamine and amphibactin have been identified in marine surface waters, suggesting that they may represent an important bacterial strategy for obtaining iron from a scarcely populated pool. We screened several strains of marine Vibrio for the presence of putative amphibactin biosynthesis gene homologues and amphibactin production. Whole cell proteomics, siderophore isolation, and isotopically labeled iron uptake experiments were performed. Here, we show that an amphibactin-producing marine bacterium, Vibrio cyclitrophicus str. 1F-53, harbors an independently regulated uptake pathway for ferrioxamines. Proteomic analyses identified upregulation of the amphibactin NRPS system and a putative amphibactin siderophore transporter in response to low iron concentrations. In addition, multiple other transporters were upregulated, however when desferrioxamine was present, amphibactin production decreased and the ferrioxamine receptor increased in abundance. Such cheating phenotypes, which appear widespread among marine amphibactin producers, highlight the strategies that contribute to the fitness of marine bacteria in the face of iron stress. These results demonstrate siderophore producer and cheater phenotypes and highlight the cellular restructuring which is involved due to competition for iron, that shapes the community structure of marine ecosystems.
2021-04-19 | PXD011427 | Pride
Project description:Serratia marcescens is a bacterium frequently found in the environment, but over the last several decades it has evolved into a concerning clinical pathogen, causing fatal bacteremia. To establish such infections, pathogens require specific nutrients; one very limited but essential nutrient is iron. We sought to characterize the iron acquisition systems in S. marcescens isolate UMH9, which was recovered from a clinical bloodstream infection. Using RNA sequencing (RNA-seq), we identified two predicted siderophore gene clusters (cbs and sch) that were regulated by iron. Mutants were constructed to delete each iron acquisition locus individually and in conjunction, generating both single and double mutants for the putative siderophore systems. Mutants lacking the sch gene cluster lost their iron-chelating ability as quantified by the chrome azurol S (CAS) assay, whereas the cbs mutant retained wild-type activity. Mass spectrometry-based analysis identified the chelating siderophore to be serratiochelin, a siderophore previously identified in Serratia plymuthica. Serratiochelin-producing mutants also displayed a decreased growth rate under iron-limited conditions created by dipyridyl added to LB medium. Additionally, mutants lacking serratiochelin were significantly outcompeted during cochallenge with wild-type UMH9 in the kidneys and spleen after inoculation via the tail vein in a bacteremia mouse model. This result was further confirmed by an independent challenge, suggesting that serratiochelin is required for full S. marcescens pathogenesis in the bloodstream. Nine other clinical isolates have at least 90% protein identity to the UMH9 serratiochelin system; therefore, our results are broadly applicable to emerging clinical isolates of S. marcescens causing bacteremia.
2020-05-24 | GSE145587 | GEO
Project description:Iron is an essential nutrient for bacterial growth but poorly bioavailable. To scavenge ferric iron present in their environment, bacteria synthesize and secrete siderophores, small compounds with a high affinity for iron. Pyochelin (PCH) is one of the two siderophores produced by the opportunistic pathogen Pseudomonas aeruginosa. Once having captured a ferric iron, PCH-Fe is imported back into bacteria first by the outer membrane transporter FptA and afterwards by the inner membrane permease FptX. Here using molecular biology, 55Fe uptake assays and LC-MS/MS quantification of PCH in the different bacterial cell fractions, we show that (i) PCH (probably under its PCH-Fe form) is able to rich bacterial periplasm and cytoplasm when both FptA and FptX are expressed, and (ii) that PchHI (a heterodimeric ABC transporter) plays a role in the translocation of siderophore-free iron siderophore-free iron across the inner membrane into the cytoplasm. Consequently, probably the first fraction of PCH-Fe internalized by FptA may be transported further by FptX in the bacterial cytoplasm to activate the transcriptional regulator PchR, regulating the transcription of all genes of the PCH pathway. The further fractions of PCH-Fe transported by FptA may dissociate in the bacterial periplasm by an unknown mechanism, with the siderophore-free iron being transported into the cytoplasm by PchHI.
2022-03-15 | PXD026833 | Pride
Project description:Pseudomonas and other environmental microorganisms have been proven capable of synthesizing siderophores, which are instrumental in the removal of iron and other metal ions from a variety of waste materials. We have elucidated the molecular mechanisms by which the BfmRS two-component system (TCS) mediates environmental stress signals to regulate siderophore production in Pseudomonas aeruginosa. In this study, we further confirm the pivotal role of the BfmRS system in bacterial iron metabolism, demonstrating its regulatory influence on key genes involved in siderophore synthesis. Moreover, overexpression of BfmR led to a marked increase in mRNA levels of siderophore-related genes, with a 2.4- to 6.7-fold elevation, which in turn significantly enhanced siderophore production and consequently improved iron utilization efficiency when compared to the wild type (WT). The heightened efficiency of the genetically modified strain to extract iron from coal fly ash (CFA) suggests the feasibility of engineered bacterial system in bioremediation. This work not only validate the intricate TCSs involved in siderophore regulation within P. aeruginosa, but also provides a compelling strategy in heavy metal recovery and hazardous waste detoxification.
2023-12-25 | PXD048123 | iProX
Project description:Iron was titrated into a mixture of commercial standards and culture extracts from two organisms (E coli Nissle and arthrobacter JB182)
2020-03-14 | MSV000085103 | GNPS
Project description:Iron is essential for many cellular processes and is required by bacteria for replication. To acquire iron from the host, pathogenic Gram-negative bacteria secrete siderophores, including Enterobactin (Ent). However, Ent is bound by the host protein Lipocalin 2 (Lcn2), preventing bacterial reuptake of aferric or ferric Ent. In two experiments we treated A549 (lung cancer cell line) cells with Lcn2, Ent, and iron, alone and in combination. In experiment 1, biological duplicates of 4 conditions were used: PBS control, Lcn2, Lcn2+Ent, and Lcn2+Ent+iron. In experiment 2, 4 biological replicates of 4 conditions were used: PBS control, Ent, iron, and Ent+iron. Targets made from the samples were hybridized to Affymetrix Human Gene 1.0 ST arrays to measure transcript abundances. The RMA algorithm was used to estimate transcript levels. Replicate samples were exchangeable, so we fit one-way ANOVA models to log2-transformed data separately to each experiment, and tested for pairwise differences between groups in each experiment, as well as asking if the Ent vs. PBS differences were larger or smaller than the Ent+iron vs. iron differences (Ent by iron interactions). We report results for 29096 probe-sets that were not annotated as positive or negative controls on the array. A supplementary Excel workbook is provided that contains the estimated expression level, some probe-set annotation, and simple statistical analysis for each probe-set. It may be convenient for some users, however obtaining newer probe-set annotation may be advisable. A549 (ATCC CCL-185) cells, a human type II pneumocyte cell line, were cultured in F12K (Invitrogen, Carlsbad, CA) media supplemented with 10% fetal bovine serum (Invitrogen) and 1:100 penicillin streptomycin (Invitrogen). 24-well plates were seeded with A549 cells at a concentration of 35000 cells/well. After two days, cells were weaned from serum and antibiotics overnight. Cells were then stimulated overnight with the indicated combinations of 50 uM Ferric ammonium citrate (FAC) (Sigma, St. Louis, MO), 50 um Ent (Sigma or EMC Microcollections, Tubingen, Germany), and/or 25 uM lipocalin 2 (Lcn2) in F12K media lacking serum or antibiotics. Prior to incubation with cells, siderophore-Lcn2 complexes were prepared by sequential incubation at room temperature of FAC and Ent for 30 minutes followed by addition of Lcn2 and incubation for an additional 30 minutes. In experiment 1, 2 biological replicates of each of these 4 conditions were grown: PBS, 25 uM lipocalin 2 (Lcn2), a combination of 25 uM Lcn2 with 50 uM Enterobactin (Ent), and a combination of Lcn2, Ent and 50 uM Ferric Ammonium Citrate (FAC). In experiment 2, 4 biological replicates of each of these 4 conditions were grown: PBS, 50 uM Ent, 50 uM FAC, and a combination of 50 uM Ent and 50 uM FAC. Cells were harvested and RNA was extracted using an miRNeasy Mini Kit (Qiagen). Biotinylated cDNA targets were prepared according to the Ambion WT expression kit protocol from 150 ng total RNA. There are no batches within each experiment - the samples in each group are exchangeable. Targets were hybridized to Affymetrix Human Gene 1.0 ST arrays. Arrays were scanned and quantified by usual procedures, and transcript abundances for 29096 non-control probe-sets estimated by an RMA algorithm.
2014-07-03 | E-GEOD-54962 | biostudies-arrayexpress
Project description:RNA-seq differential gene expression profiling of Corynebacterium pseudotuberculosis under iron limitation. We use two C. pseudotuberculosis strains, a ciuA Cp13 mutant and its parental wild type T1 strain to quantify the relative gene expression of these strains in cultures with low iron availability. The ciuA Cp13 mutant is deficient for an operon which encodes a iron siderophore uptake system. The Cp13 mutant also showed reduced virulence in CLA infection models. Due to the importance of iron for many pathogenic bacteria, our primary goal was to identified the expression profile of iron regulated genes.
2019-07-31 | GSE114125 | GEO
Project description:We investigated the gene expression responses of Candidatus Pelagibacter ubique cultures to iron limitation. Differential expression was observed for genes in iron acquisition and incorporation operons. SfuC in particular was 16 times higher in iron-limited cultures and encodes a periplasmic iron-binding protein. Six natural seawater cultures were amended with minimal nutrients and inoculated with P. ubique. Close to maximum cell density, all carboys were supplemented with 100 nM ferrichrome (an iron-chelating siderophore) and three carboys were additionally supplemented with 1 µM FeCl3. Each of the six carboys was sampled for microarray analyses one, two, and eleven days after the ferrichrome addition.
2010-05-05 | E-GEOD-20962 | biostudies-arrayexpress
Project description:Transcriptional profiling of low-iron stimulon and XibR influenced regulon using wild-type Xcc 8004 and xibR mutant grown under iron-replete and iron-deplete conditions. Trancriptional analysis under iron-deplete condition, mimicking in planta environment, provides greater insights into expression pattern of several virulence-associated functions under low-iron. A genetic screen sggested the involvement of XibR (Xanthomonas iron binding regulator) in iron-uptake and metabolism. Present transcriptional analysis suggested the co-regulation of virulence associated functions including siderophore biosynthesis, motility, chemotaxis and typeIII effectors by a novel transcriptional regulator of NtrC family protein XibR and iron avability.
2016-11-02 | GSE89267 | GEO