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Sample records for alcaligin siderophore biosynthesis

  1. Evolution of a novel lysine decarboxylase in siderophore biosynthesis.

    PubMed

    Burrell, Matthew; Hanfrey, Colin C; Kinch, Lisa N; Elliott, Katherine A; Michael, Anthony J

    2012-10-01

    Structural backbones of iron-scavenging siderophore molecules include polyamines 1,3-diaminopropane and 1,5-diaminopentane (cadaverine). For the cadaverine-based desferroxiamine E siderophore in Streptomyces coelicolor, the corresponding biosynthetic gene cluster contains an ORF encoded by desA that was suspected of producing the cadaverine (decarboxylated lysine) backbone. However, desA encodes an l-2,4-diaminobutyrate decarboxylase (DABA DC) homologue and not any known form of lysine decarboxylase (LDC). The only known function of DABA DC is, together with l-2,4-aminobutyrate aminotransferase (DABA AT), to synthesize 1,3-diaminopropane. We show here that S. coelicolor desA encodes a novel LDC and we hypothesized that DABA DC homologues present in siderophore biosynthetic clusters in the absence of DABA AT ORFs would be novel LDCs. We confirmed this by correctly predicting the LDC activity of a DABA DC homologue from a Yersinia pestis siderophore biosynthetic pathway. The corollary was confirmed for a DABA DC homologue, adjacent to a DABA AT ORF in a siderophore pathway in the cyanobacterium Anabaena variabilis, which was shown to be a bona fide DABA DC. These findings enable prediction of whether a siderophore pathway will utilize 1,3-diaminopropane or cadaverine, and suggest that the majority of bacteria use DABA AT and DABA DC for siderophore, rather than norspermidine/polyamine biosynthesis. PMID:22906379

  2. AcsD catalyzes enantioselective citrate desymmetrization in siderophore biosynthesis.

    PubMed

    Schmelz, Stefan; Kadi, Nadia; McMahon, Stephen A; Song, Lijiang; Oves-Costales, Daniel; Oke, Muse; Liu, Huanting; Johnson, Kenneth A; Carter, Lester G; Botting, Catherine H; White, Malcolm F; Challis, Gregory L; Naismith, James H

    2009-03-01

    Bacterial pathogens need to scavenge iron from their host for growth and proliferation during infection. They have evolved several strategies to do this, one being the biosynthesis and excretion of small, high-affinity iron chelators known as siderophores. The biosynthesis of siderophores is an important area of study, not only for potential therapeutic intervention but also to illuminate new enzyme chemistries. Two general pathways for siderophore biosynthesis exist: the well-characterized nonribosomal peptide synthetase (NRPS)-dependent pathway and the NRPS-independent siderophore (NIS) pathway, which relies on a different family of sparsely investigated synthetases. Here we report structural and biochemical studies of AcsD from Pectobacterium (formerly Erwinia) chrysanthemi, an NIS synthetase involved in achromobactin biosynthesis. The structures of ATP and citrate complexes provide a mechanistic rationale for stereospecific formation of an enzyme-bound (3R)-citryladenylate, which reacts with L-serine to form a likely achromobactin precursor. AcsD is a unique acyladenylate-forming enzyme with a new fold and chemical catalysis strategy. PMID:19182782

  3. Genetics and Assembly Line Enzymology of Siderophore Biosynthesis in Bacteria

    PubMed Central

    Crosa, Jorge H.; Walsh, Christopher T.

    2002-01-01

    The regulatory logic of siderophore biosynthetic genes in bacteria involves the universal repressor Fur, which acts together with iron as a negative regulator. However in other bacteria, in addition to the Fur-mediated mechanism of regulation, there is a concurrent positive regulation of iron transport and siderophore biosynthetic genes that occurs under conditions of iron deprivation. Despite these regulatory differences the mechanisms of siderophore biosynthesis follow the same fundamental enzymatic logic, which involves a series of elongating acyl-S-enzyme intermediates on multimodular protein assembly lines: nonribosomal peptide synthetases (NRPS). A substantial variety of siderophore structures are produced from similar NRPS assembly lines, and variation can come in the choice of the phenolic acid selected as the N-cap, the tailoring of amino acid residues during chain elongation, the mode of chain termination, and the nature of the capturing nucleophile of the siderophore acyl chain being released. Of course the specific parts that get assembled in a given bacterium may reflect a combination of the inventory of biosynthetic and tailoring gene clusters available. This modular assembly logic can account for all known siderophores. The ability to mix and match domains within modules and to swap modules themselves is likely to be an ongoing process in combinatorial biosynthesis. NRPS evolution will try out new combinations of chain initiation, elongation and tailoring, and termination steps, possibly by genetic exchange with other microorganisms and/or within the same bacterium, to create new variants of iron-chelating siderophores that can fit a particular niche for the producer bacterium. PMID:12040125

  4. Fungal siderophore biosynthesis is partially localized in peroxisomes

    PubMed Central

    Gründlinger, Mario; Yasmin, Sabiha; Lechner, Beatrix Elisabeth; Geley, Stephan; Schrettl, Markus; Hynes, Michael; Haas, Hubertus

    2013-01-01

    Siderophores play a central role in iron metabolism and virulence of most fungi. Both Aspergillus fumigatus and Aspergillus nidulans excrete the siderophore triacetylfusarinine C (TAFC) for iron acquisition. In A. fumigatus, green fluorescence protein-tagging revealed peroxisomal localization of the TAFC biosynthetic enzymes SidI (mevalonyl-CoA ligase), SidH (mevalonyl-CoA hydratase) and SidF (anhydromevalonyl-CoA transferase), while elimination of the peroxisomal targeting signal (PTS) impaired both, peroxisomal SidH-targeting and TAFC biosynthesis. The analysis of A. nidulans mutants deficient in peroxisomal biogenesis, ATP import or protein import revealed that cytosolic mislocalization of one or two but, interestingly, not all three enzymes impairs TAFC production during iron starvation. The PTS motifs are conserved in fungal orthologues of SidF, SidH and SidI. In agreement with the evolutionary conservation of the partial peroxisomal compartmentalization of fungal siderophore biosynthesis, the SidI orthologue of coprogen-type siderophore-producing Neurospora crassa was confirmed to be peroxisomal. Taken together, this study identified and characterized a novel, evolutionary conserved metabolic function of peroxisomes. PMID:23617799

  5. Reduced Virulence of a Bordetella bronchiseptica Siderophore Mutant in Neonatal Swine

    PubMed Central

    Register, Karen B.; Ducey, Thomas F.; Brockmeier, Susan L.; Dyer, David W.

    2001-01-01

    One means by which Bordetella bronchiseptica scavenges iron is through production of the siderophore alcaligin. A nonrevertible alcaligin mutant derived from the virulent strain 4609, designated DBB25, was constructed by insertion of a kanamycin resistance gene into alcA, one of the genes essential for alcaligin biosynthesis. The virulence of the alcA mutant in colostrum-deprived, caesarean-delivered piglets was compared with that of the parent strain in two experiments. At 1 week of age, piglets were inoculated with phosphate-buffered saline, 4609, or DBB25. Two piglets in each group were euthanatized on day 10 postinfection. The remainder were euthanatized at 21 days postinfection. Clinical signs, including fever, coughing, and sneezing, were present in both groups. Nasal washes performed 7, 14, and 21 days postinoculation demonstrated that strain DBB25 colonized the nasal cavity but did so at levels that were significantly less than those achieved by strain 4609. Analysis of colonization based on the number of CFU per gram of tissue recovered from the turbinate, trachea, and lung also demonstrated significant differences between DBB25 and 4609, at both day 10 and day 21 postinfection. Mild to moderate turbinate atrophy was apparent in pigs inoculated with strain 4609, while turbinates of those infected with strain DBB25 developed no or mild atrophy. We conclude from these results that siderophore production by B. bronchiseptica is not essential for colonization of swine but is required for maximal virulence. B. bronchiseptica mutants with nonrevertible defects in genes required for alcaligin synthesis may be candidates for evaluation as attenuated, live vaccine strains in conventionally reared pigs. PMID:11254568

  6. Cellular organization of siderophore biosynthesis in Pseudomonas aeruginosa: Evidence for siderosomes.

    PubMed

    Gasser, Véronique; Guillon, Laurent; Cunrath, Olivier; Schalk, Isabelle J

    2015-07-01

    Pyoverdine I (PVDI) and pyochelin (PCH) are the two major siderophores produced by Pseudomonas aeruginosa PAO1 to import iron. The biochemistry of the biosynthesis of these two siderophores has been described in detail in the literature over recent years. PVDI assembly requires the coordinated action of seven cytoplasmic enzymes and is followed by a periplasmic maturation before secretion of the siderophore into the extracellular medium by the efflux system PvdRT-OpmQ. PCH biosynthesis also involves seven cytoplasmic enzymes but no periplasmic maturation. Recent findings indicate that the cytoplasmic enzymes involved in each of these two siderophore biosynthesis pathways can form siderophore-specific multi-enzymatic complexes called siderosomes associated with the inner leaflet of the cytoplasmic membrane. This organization may optimize the transfer of the siderophore precursors between the various participating enzymes and avoid the diffusion of siderophore precursors, able to chelate metals, throughout the cytoplasm. Here, we describe these recently published findings and discuss the existence of these siderosomes in P. aeruginosa. PMID:25697961

  7. Mixing and Matching Siderophore Clusters: Structure and Biosynthesis of Serratiochelins from Serratia sp. V4

    PubMed Central

    2012-01-01

    Interrogation of the evolutionary history underlying the remarkable structures and biological activities of natural products has been complicated by not knowing the functions they have evolved to fulfill. Siderophores—soluble, low molecular weight compounds—have an easily understood and measured function: acquiring iron from the environment. Bacteria engage in a fierce competition to acquire iron, which rewards the production of siderophores that bind iron tightly and cannot be used or pirated by competitors. The structures and biosyntheses of “odd” siderophores can reveal the evolutionary strategy that led to their creation. We report a new Serratia strain that produces serratiochelin and an analog of serratiochelin. A genetic approach located the serratiochelin gene cluster, and targeted mutations in several genes implicated in serratiochelin biosynthesis were generated. Bioinformatic analyses and mutagenesis results demonstrate that genes from two well-known siderophore clusters, the Escherichia coli enterobactin cluster and the Vibrio cholera vibriobactin cluster, were shuffled to produce a new siderophore biosynthetic pathway. These results highlight how modular siderophore gene clusters can be mixed and matched during evolution to generate structural diversity in siderophores. PMID:22830960

  8. Enzymatic tailoring of ornithine in the biosynthesis of the Rhizobium cyclic trihydroxamate siderophore vicibactin.

    PubMed

    Heemstra, John R; Walsh, Christopher T; Sattely, Elizabeth S

    2009-10-28

    To acquire iron, the N(2)-fixing, symbiotic bacterium Rhizobium sp. produce the cyclic trihydroxamate siderophore vicibactin, containing a 30-membered trilactone scaffold. Herein we report the overproduction and purification of the six proteins VbsACGOLS in the bacterial host Escherichia coli and the reconstitution of the biosynthesis of vicibactin from primary metabolites. The flavoprotein VbsO acts as a pathway-initiating l-ornithine N(5)-hydroxylase, followed by VbsA, which transfers (R)-3-hydroxybutyryl- from the CoA thioester to N(5)-hydroxyornithine to yield N(5)-((R)-3-hydroxybutyryl)-N(5)-hydroxy-l-ornithine. VbsL is a PLP-dependent epimerase acting at C(2) of the 10 atom monomer unit. VbsS, a nonribosomal peptide synthetase free-standing module, then activates N(5)-((R)-3-hydroxybutyryl)-N(5)-hydroxy-d-ornithine as the AMP anhydride on the way to cyclotrimerization to the vicibactin scaffold. The last step, tris-acetylation of the C(2) amino group of desacetyl-d-vicibactin to the mature siderophore vicibactin, is catalyzed distributively by VbsC, using three molecules of acetyl-CoA. PMID:19778043

  9. Plasmid- and chromosome-encoded siderophore anguibactin systems found in marine vibrios: biosynthesis, transport and evolution

    PubMed Central

    Liu, Moqing; Actis, Luis A.; Crosa, Jorge H.

    2013-01-01

    Vibrio anguillarum is a marine pathogen that causes vibriosis, a hemorrhagic septicemia in aquatic invertebrate as well as vertebrate animals. The siderophore anguibactin system is one of the most important virulence factors of this bacterium. Most of the anguibactin biosynthesis and transport genes are located in the 65-kb pJM1 virulence plasmid although some of them are found in the chromosome of this fish pathogen. Over 30 years of research unveiled the role numerous chromosomal and pJM1 genes play in the synthesis of anguibactin and the transport of cognate ferric complexes into the bacterial cell. Furthermore, these studies showed that pJM1-carrying strains might be originated from pJM1-less strains producing the chromosome-mediated siderophore vanchrobactin. Additionally, we recently identified a chromosome-mediated anguibactin system in V. harveyi suggesting the possible evolutional origin of the V. anguillarum anguibactin system. In this review, we present our current understanding of the mechanisms and evolution hypothesis of the anguibactin system that might have occurred in these pathogenic vibrios. PMID:23660776

  10. Plasmid- and chromosome-encoded siderophore anguibactin systems found in marine vibrios: biosynthesis, transport and evolution.

    PubMed

    Naka, Hiroaki; Liu, Moqing; Actis, Luis A; Crosa, Jorge H

    2013-08-01

    Vibrio anguillarum is a marine pathogen that causes vibriosis, a hemorrhagic septicemia in aquatic invertebrate as well as vertebrate animals. The siderophore anguibactin system is one of the most important virulence factors of this bacterium. Most of the anguibactin biosynthesis and transport genes are located in the 65-kb pJM1 virulence plasmid although some of them are found in the chromosome of this fish pathogen. Over 30 years of research unveiled the role numerous chromosomal and pJM1 genes play in the synthesis of anguibactin and the transport of cognate ferric complexes into the bacterial cell. Furthermore, these studies showed that pJM1-carrying strains might be originated from pJM1-less strains producing the chromosome-mediated siderophore vanchrobactin. Additionally, we recently identified a chromosome-mediated anguibactin system in V. harveyi suggesting the possible evolutional origin of the V. anguillarum anguibactin system. In this review, we present our current understanding of the mechanisms and evolution hypothesis of the anguibactin system that might have occurred in these pathogenic vibrios. PMID:23660776

  11. Enzymatic Tailoring of Ornithine in the Biosynthesis of the Rhizobium Cyclic Trihydroxamate Siderophore Vicibactin

    PubMed Central

    Heemstra, John R.; Walsh, Christopher T.; Sattely, Elizabeth S.

    2009-01-01

    To acquire iron, the N2-fixing, symbiotic bacterium Rhizobium sp. produce the cyclic tri-hydroxamate siderophore vicibactin, containing a 30-membered tri-lactone scaffold. Herein we report the overproduction and purification of the six proteins VbsACGOLS in the bacterial host Escherichia coli and the reconstitution of the biosynthesis of vicibactin from primary metabolites. The flavoprotein VbsO acts as a pathway-initiating L-ornithine N5-hydroxylase, followed by VbsA which transfers (R)-3-hydroxybutyryl- from the CoA thioester to N5-hydroxyornithine to yield N5-((R)-3-hydroxybutyryl)-N5-hydroxy-L-ornithine. VbsL is a PLP-dependent epimerase acting at C2 of the 10 atom monomer unit. VbsS, a nonribosomal peptide synthetase free standing module, then activates N5-((R)-3-hydroxybutyryl)-N5-hydroxy-D-ornithine as the AMP anhydride on the way to cyclotrimerization to the vicibactin scaffold. The last step, tris-acetylation of the C2 amino group of desacetyl-D-vicibactin to the mature siderophore vicibactin is catalyzed distributively by VbsC, using three molecules of acetyl-CoA. PMID:19778043

  12. Cloning of a Vibrio cholerae vibriobactin gene cluster: identification of genes required for early steps in siderophore biosynthesis.

    PubMed Central

    Wyckoff, E E; Stoebner, J A; Reed, K E; Payne, S M

    1997-01-01

    Vibrio cholerae secretes the catechol siderophore vibriobactin in response to iron limitation. Vibriobactin is structurally similar to enterobactin, the siderophore produced by Escherichia coli, and both organisms produce 2,3-dihydroxybenzoic acid (DHBA) as an intermediate in siderophore biosynthesis. To isolate and characterize V. cholerae genes involved in vibriobactin biosynthesis, we constructed a genomic cosmid bank of V. cholerae DNA and isolated clones that complemented mutations in E. coli enterobactin biosynthesis genes. V. cholerae homologs of entA, entB, entC, entD, and entE were identified on overlapping cosmid clones. Our data indicate that the vibriobactin genes are clustered, like the E. coli enterobactin genes, but the organization of the genes within these clusters is different. In this paper, we present the organization and sequences of genes involved in the synthesis and activation of DHBA. In addition, a V. cholerae strain with a chromosomal mutation in vibA was constructed by marker exchange. This strain was unable to produce vibriobactin or DHBA, confirming that in V. cholerae VibA catalyzes an early step in vibriobactin biosynthesis. PMID:9371453

  13. SREB, a GATA Transcription Factor That Directs Disparate Fates in Blastomyces dermatitidis Including Morphogenesis and Siderophore Biosynthesis

    PubMed Central

    Gauthier, Gregory M.; Sullivan, Thomas D.; Gallardo, Sergio S.; Brandhorst, T. Tristan; Vanden Wymelenberg, Amber J.; Cuomo, Christina A.; Suen, Garret; Currie, Cameron R.; Klein, Bruce S.

    2010-01-01

    Blastomyces dermatitidis belongs to a group of human pathogenic fungi that exhibit thermal dimorphism. At 22°C, these fungi grow as mold that produce conidia or infectious particles, whereas at 37°C they convert to budding yeast. The ability to switch between these forms is essential for virulence in mammals and may enable these organisms to survive in the soil. To identify genes that regulate this phase transition, we used Agrobacterium tumefaciens to mutagenize B. dermatitidis conidia and screened transformants for defects in morphogenesis. We found that the GATA transcription factor SREB governs multiple fates in B. dermatitidis: phase transition from yeast to mold, cell growth at 22°C, and biosynthesis of siderophores under iron-replete conditions. Insertional and null mutants fail to convert to mold, do not accumulate significant biomass at 22°C, and are unable to suppress siderophore biosynthesis under iron-replete conditions. The defect in morphogenesis in the SREB mutant was independent of exogenous iron concentration, suggesting that SREB promotes the phase transition by altering the expression of genes that are unrelated to siderophore biosynthesis. Using bioinformatic and gene expression analyses, we identified candidate genes with upstream GATA sites whose expression is altered in the null mutant that may be direct or indirect targets of SREB and promote the phase transition. We conclude that SREB functions as a transcription factor that promotes morphogenesis and regulates siderophore biosynthesis. To our knowledge, this is the first gene identified that promotes the conversion from yeast to mold in the dimorphic fungi, and may shed light on environmental persistence of these pathogens. PMID:20368971

  14. Genetic and Functional Analysis of the Biosynthesis of a Non-Ribosomal Peptide Siderophore in Burkholderia xenovorans LB400.

    PubMed

    Vargas-Straube, María José; Cámara, Beatriz; Tello, Mario; Montero-Silva, Francisco; Cárdenas, Franco; Seeger, Michael

    2016-01-01

    B. xenovorans LB400 is a model bacterium for the study of the metabolism of aromatic compounds. The aim of this study was the genomic and functional characterization of a non-ribosomal peptide synthetase containing gene cluster that encodes a siderophore in B. xenovorans LB400. The mba gene cluster from strain LB400 encodes proteins involved in the biosynthesis and transport of a hydroxamate-type siderophore. Strain LB400 has a unique mba gene organization, although mba gene clusters have been observed in diverse Burkholderiales. Bioinformatic analysis revealed the presence of promoters in the mba gene cluster that strongly suggest regulation by the ferric uptake regulator protein (Fur) and by the alternative RNA polymerase extracytoplasmic function sigma factor MbaF. Reverse transcriptase PCR analyses showed the expression of iron-regulated transcriptional units mbaFGHIJKL, mbaN, mbaABCE, mbaO, mbaP and mbaD genes under iron limitation. Chrome azurol S (CAS) assay strongly suggests that strain LB400 synthesized a siderophore under iron limitation. Mass spectrometry ESI-MS and MALDI-TOF-MS analyses revealed that the siderophore is a non-ribosomal peptide, and forms an iron complex with a molecular mass of 676 Da. Based on bioinformatic prediction, CAS assay and MS analyses, we propose that the siderophore is L-Nδ-hydroxy-Nδ-formylOrn-D-β-hydroxyAsp-L-Ser-L-Nδ-hydroxy-Nδ-formylOrn-1,4-diaminobutane that is closely related to malleobactin-type siderophores reported in B. thailandensis. PMID:26963250

  15. Genetic and Functional Analysis of the Biosynthesis of a Non-Ribosomal Peptide Siderophore in Burkholderia xenovorans LB400

    PubMed Central

    Vargas-Straube, María José; Cámara, Beatriz; Tello, Mario; Montero-Silva, Francisco; Cárdenas, Franco; Seeger, Michael

    2016-01-01

    B. xenovorans LB400 is a model bacterium for the study of the metabolism of aromatic compounds. The aim of this study was the genomic and functional characterization of a non-ribosomal peptide synthetase containing gene cluster that encodes a siderophore in B. xenovorans LB400. The mba gene cluster from strain LB400 encodes proteins involved in the biosynthesis and transport of a hydroxamate-type siderophore. Strain LB400 has a unique mba gene organization, although mba gene clusters have been observed in diverse Burkholderiales. Bioinformatic analysis revealed the presence of promoters in the mba gene cluster that strongly suggest regulation by the ferric uptake regulator protein (Fur) and by the alternative RNA polymerase extracytoplasmic function sigma factor MbaF. Reverse transcriptase PCR analyses showed the expression of iron-regulated transcriptional units mbaFGHIJKL, mbaN, mbaABCE, mbaO, mbaP and mbaD genes under iron limitation. Chrome azurol S (CAS) assay strongly suggests that strain LB400 synthesized a siderophore under iron limitation. Mass spectrometry ESI-MS and MALDI-TOF-MS analyses revealed that the siderophore is a non-ribosomal peptide, and forms an iron complex with a molecular mass of 676 Da. Based on bioinformatic prediction, CAS assay and MS analyses, we propose that the siderophore is L-Nδ-hydroxy-Nδ-formylOrn-D-β-hydroxyAsp-L-Ser-L-Nδ-hydroxy-Nδ-formylOrn-1,4-diaminobutane that is closely related to malleobactin-type siderophores reported in B. thailandensis. PMID:26963250

  16. Siderophore biosynthesis coordinately modulated the virulence-associated interactive metabolome of uropathogenic Escherichia coli and human urine.

    PubMed

    Su, Qiao; Guan, Tianbing; Lv, Haitao

    2016-01-01

    Uropathogenic Escherichia coli (UPEC) growth in women's bladders during urinary tract infection (UTI) incurs substantial chemical exchange, termed the "interactive metabolome", which primarily accounts for the metabolic costs (utilized metabolome) and metabolic donations (excreted metabolome) between UPEC and human urine. Here, we attempted to identify the individualized interactive metabolome between UPEC and human urine. We were able to distinguish UPEC from non-UPEC by employing a combination of metabolomics and genetics. Our results revealed that the interactive metabolome between UPEC and human urine was markedly different from that between non-UPEC and human urine, and that UPEC triggered much stronger perturbations in the interactive metabolome in human urine. Furthermore, siderophore biosynthesis coordinately modulated the individualized interactive metabolome, which we found to be a critical component of UPEC virulence. The individualized virulence-associated interactive metabolome contained 31 different metabolites and 17 central metabolic pathways that were annotated to host these different metabolites, including energetic metabolism, amino acid metabolism, and gut microbe metabolism. Changes in the activities of these pathways mechanistically pinpointed the virulent capability of siderophore biosynthesis. Together, our findings provide novel insights into UPEC virulence, and we propose that siderophores are potential targets for further discovery of drugs to treat UPEC-induced UTI. PMID:27076285

  17. Siderophore biosynthesis coordinately modulated the virulence-associated interactive metabolome of uropathogenic Escherichia coli and human urine

    PubMed Central

    Su, Qiao; Guan, Tianbing; Lv, Haitao

    2016-01-01

    Uropathogenic Escherichia coli (UPEC) growth in women’s bladders during urinary tract infection (UTI) incurs substantial chemical exchange, termed the “interactive metabolome”, which primarily accounts for the metabolic costs (utilized metabolome) and metabolic donations (excreted metabolome) between UPEC and human urine. Here, we attempted to identify the individualized interactive metabolome between UPEC and human urine. We were able to distinguish UPEC from non-UPEC by employing a combination of metabolomics and genetics. Our results revealed that the interactive metabolome between UPEC and human urine was markedly different from that between non-UPEC and human urine, and that UPEC triggered much stronger perturbations in the interactive metabolome in human urine. Furthermore, siderophore biosynthesis coordinately modulated the individualized interactive metabolome, which we found to be a critical component of UPEC virulence. The individualized virulence-associated interactive metabolome contained 31 different metabolites and 17 central metabolic pathways that were annotated to host these different metabolites, including energetic metabolism, amino acid metabolism, and gut microbe metabolism. Changes in the activities of these pathways mechanistically pinpointed the virulent capability of siderophore biosynthesis. Together, our findings provide novel insights into UPEC virulence, and we propose that siderophores are potential targets for further discovery of drugs to treat UPEC-induced UTI. PMID:27076285

  18. Environmental Factors Modulating Antibiotic and Siderophore Biosynthesis by Pseudomonas fluorescens Biocontrol Strains

    PubMed Central

    Duffy, Brion K.; Défago, Geneviève

    1999-01-01

    Understanding the environmental factors that regulate the biosynthesis of antimicrobial compounds by disease-suppressive strains of Pseudomonas fluorescens is an essential step toward improving the level and reliability of their biocontrol activity. We used liquid culture assays to identify several minerals and carbon sources which had a differential influence on the production of the antibiotics 2,4-diacetylphloroglucinol (PHL), pyoluteorin (PLT), and pyrrolnitrin and the siderophores salicylic acid and pyochelin by the model strain CHA0, which was isolated from a natural disease-suppressive soil in Switzerland. Production of PHL was stimulated by Zn2+, NH4Mo2+, and glucose; the precursor compound mono-acetylphloroglucinol was stimulated by the same factors as PHL. Production of PLT was stimulated by Zn2+, Co2+, and glycerol but was repressed by glucose. Pyrrolnitrin production was increased by fructose, mannitol, and a mixture of Zn2+ and NH4Mo2+. Pyochelin production was increased by Co2+, fructose, mannitol, and glucose. Interestingly, production of its precursor salicylic acid was increased by different factors, i.e., NH4Mo2+, glycerol, and glucose. The mixture of Zn2+ and NH4Mo2+ with fructose, mannitol, or glycerol further enhanced the production of PHL and PLT compared with either the minerals or the carbon sources used alone, but it did not improve siderophore production. Extending fermentation time from 2 to 5 days increased the accumulation of PLT, pyrrolnitrin, and pyochelin but not of PHL. When findings with CHA0 were extended to an ecologically and genetically diverse collection of 41 P. fluorescens biocontrol strains, the effect of certain factors was strain dependent, while others had a general effect. Stimulation of PHL by Zn2+ and glucose was strain dependent, whereas PLT production by all strains that can produce this compound was stimulated by Zn2+ and transiently repressed by glucose. Inorganic phosphate reduced PHL production by CHA0 and seven

  19. Baulamycins A and B, broad-spectrum antibiotics identified as inhibitors of siderophore biosynthesis in Staphylococcus aureus and Bacillus anthracis.

    PubMed

    Tripathi, Ashootosh; Schofield, Michael M; Chlipala, George E; Schultz, Pamela J; Yim, Isaiah; Newmister, Sean A; Nusca, Tyler D; Scaglione, Jamie B; Hanna, Philip C; Tamayo-Castillo, Giselle; Sherman, David H

    2014-01-29

    Siderophores are high-affinity iron chelators produced by microorganisms and frequently contribute to the virulence of human pathogens. Targeted inhibition of the biosynthesis of siderophores staphyloferrin B of Staphylococcus aureus and petrobactin of Bacillus anthracis hold considerable potential as a single or combined treatment for methicillin-resistant S. aureus (MRSA) and anthrax infection, respectively. The biosynthetic pathways for both siderophores involve a nonribosomal peptide synthetase independent siderophore (NIS) synthetase, including SbnE in staphyloferrin B and AsbA in petrobactin. In this study, we developed a biochemical assay specific for NIS synthetases to screen for inhibitors of SbnE and AsbA against a library of marine microbial-derived natural product extracts (NPEs). Analysis of the NPE derived from Streptomyces tempisquensis led to the isolation of the novel antibiotics baulamycins A (BmcA, 6) and B (BmcB, 7). BmcA and BmcB displayed in vitro activity with IC50 values of 4.8 μM and 19 μM against SbnE and 180 μM and 200 μM against AsbA, respectively. Kinetic analysis showed that the compounds function as reversible competitive enzyme inhibitors. Liquid culture studies with S. aureus , B. anthracis , E. coli , and several other bacterial pathogens demonstrated the capacity of these natural products to penetrate bacterial barriers and inhibit growth of both Gram-positive and Gram-negative species. These studies provide proof-of-concept that natural product inhibitors targeting siderophore virulence factors can provide access to novel broad-spectrum antibiotics, which may serve as important leads for the development of potent anti-infective agents. PMID:24401083

  20. Analysis of the pmsCEAB Gene Cluster Involved in Biosynthesis of Salicylic Acid and the Siderophore Pseudomonine in the Biocontrol Strain Pseudomonas fluorescens WCS374

    PubMed Central

    Mercado-Blanco, Jesús; van der Drift, Koen M. G. M.; Olsson, Per E.; Thomas-Oates, Jane E.; van Loon, Leendert C.; Bakker, Peter A. H. M.

    2001-01-01

    Mutants of Pseudomonas fluorescens WCS374 defective in biosynthesis of the fluorescent siderophore pseudobactin still display siderophore activity, indicating the production of a second siderophore. A recombinant cosmid clone (pMB374-07) of a WCS374 gene library harboring loci necessary for the biosynthesis of salicylic acid (SA) and this second siderophore pseudomonine was isolated. The salicylate biosynthesis region of WCS374 was localized in a 5-kb EcoRI fragment of pMB374-07. The SA and pseudomonine biosynthesis region was identified by transfer of cosmid pMB374-07 to a pseudobactin-deficient strain of P. putida. Sequence analysis of the 5-kb subclone revealed the presence of four open reading frames (ORFs). Products of two ORFs (pmsC and pmsB) showed homologies with chorismate-utilizing enzymes; a third ORF (pmsE) encoded a protein with strong similarity with enzymes involved in the biosynthesis of siderophores in other bacterial species. The region also contained a putative histidine decarboxylase gene (pmsA). A putative promoter region and two predicted iron boxes were localized upstream of pmsC. We determined by reverse transcriptase-mediated PCR that the pmsCEAB genes are cotranscribed and that expression is iron regulated. In vivo expression of SA genes was achieved in P. putida and Escherichia coli cells. In E. coli, deletions affecting the first ORF (pmsC) diminished SA production, whereas deletion of pmsB abolished it completely. The pmsB gene induced low levels of SA production in E. coli when expressed under control of the lacZ promoter. Several lines of evidence indicate that SA and pseudomonine biosynthesis are related. Moreover, we isolated a Tn5 mutant (374-05) that is simultaneously impaired in SA and pseudomonine production. PMID:11222588

  1. Biosynthesis of amphi-enterobactin siderophores by Vibrio harveyi BAA-1116: identification of a bifunctional nonribosomal peptide synthetase condensation domain.

    PubMed

    Zane, Hannah K; Naka, Hiroaki; Rosconi, Federico; Sandy, Moriah; Haygood, Margo G; Butler, Alison

    2014-04-16

    The genome of Vibrio harveyi BAA-1116 contains a nonribosomal peptide synthetase (NRPS) gene cluster (aebA-F) resembling that for enterobactin, yet enterobactin is not produced. A gene predicted to encode a long-chain fatty acid CoA ligase (FACL), similar to enzymes involved in the biosynthesis of acyl peptides, resides 15 kb away from the putative enterobactin-like biosynthetic gene cluster (aebG). The proximity of this FACL gene to the enterobactin-like synthetase suggested that V. harveyi may produce amphiphilic enterobactin-like siderophores. Extraction of the bacterial cell pellet of V. harveyi led to the isolation and structure determination of a suite of eight amphi-enterobactin siderophores composed of the cyclic lactone of tris-2,3-dihydroxybenzoyl-L-serine and acyl-L-serine. The FACL knockout mutant, ΔaebG V. harveyi, and the NRPS knockout mutant, ΔaebF V. harveyi, do not produce amphi-enterobactins. The amphi-enterobactin biosynthetic machinery was heterologously expressed in Escherichia coli and reconstituted in vitro, demonstrating the condensation domain of AebF has unique activity, catalyzing two distinct condensation reactions. PMID:24701966

  2. The pobA gene of Burkholderia cenocepacia encodes a group I Sfp-type phosphopantetheinyltransferase required for biosynthesis of the siderophores ornibactin and pyochelin.

    PubMed

    Asghar, Atif H; Shastri, Sravanthi; Dave, Emma; Wowk, Irena; Agnoli, Kirsty; Cook, Anne M; Thomas, Mark S

    2011-02-01

    The opportunistic pathogen Burkholderia cenocepacia produces the siderophores ornibactin and pyochelin under iron-restricted conditions. Biosynthesis of both siderophores requires the involvement of non-ribosomal peptide synthetases (NRPSs). Using a transposon containing the lacZ reporter gene, two B. cenocepacia mutants were isolated which were deficient in siderophore production. Mutant IW10 was shown to produce normal amounts of ornibactin but only trace amounts of pyochelin, whereas synthesis of both siderophores was abolished in AHA27. Growth of AHA27, but not IW10, was inhibited under iron-restricted conditions. In both mutants, the transposon had integrated into the pobA gene, which encodes a polypeptide exhibiting similarity to the Sfp-type phosphopantetheinyltransferases (PPTases). These enzymes are responsible for activation of NRPSs by the covalent attachment of the 4'-phosphopantetheine (P-pant) moiety of coenzyme A. Previously characterized PPTase genes from other bacteria were shown to efficiently complement both mutants for siderophore production when provided in trans. The B. cenocepacia pobA gene was also able to efficiently complement an Escherichia coli entD mutant for production of the siderophore enterobactin. Using mutant IW10, in which the lacZ gene carried by the transposon is inserted in the same orientation as pobA, it was shown that pobA is not appreciably iron-regulated. Finally, we confirmed that Sfp-type bacterial PPTases can be subdivided into two distinct groups, and we present the amino acid signature sequences which characterize each of these groups. PMID:20966087

  3. Isolation and characterization of Bacillus subtilis genes involved in siderophore biosynthesis: relationship between B. subtilis sfpo and Escherichia coli entD genes.

    PubMed Central

    Grossman, T H; Tuckman, M; Ellestad, S; Osburne, M S

    1993-01-01

    In response to iron deprivation, Bacillus subtilis secretes a catecholic siderophore, 2,3-dihydroxybenzoyl glycine, which is similar to the precursor of the Escherichia coli siderophore enterobactin. We isolated two sets of B. subtilis DNA sequences that complemented the mutations of several E. coli siderophore-deficient (ent) mutants with defective enterobactin biosynthesis enzymes. One set contained DNA sequences that complemented only an entD mutation. The second set contained DNA sequences that complemented various combinations of entB, entE, entC, and entA mutations. The two sets of DNA sequences did not appear to overlap. AB. subtilis mutant containing an insertion in the region of the entD homolog grew much more poorly in low-iron medium and with markedly different kinetics. These data indicate that (i) at least five of the siderophore biosynthesis genes of B. subtilis can function in E. coli, (ii) the genetic organization of these siderophore genes in B. subtilis is similar to that in E. coli, and (iii) the B. subtilis entD homolog is required for efficient growth in low-iron medium. The nucleotide sequence of the B. subtilis DNA contained in plasmid pENTA22, a clone expressing the B. subtilis entD homolog, revealed the presence of at least two genes. One gene was identified as sfpo, a previously reported gene involved in the production of surfactin in B. subtilis and which is highly homologous to the E. coli entD gene. We present evidence that the E. coli entD and B. subtilis sfpo genes are interchangeable and that their products are members of a new family of proteins which function in the secretion of peptide molecules. Images PMID:8407792

  4. Synthesis of Chromone, Quinolone, and Benzoxazinone Sulfonamide Nucleosides as Conformationally Constrained Inhibitors of Adenylating Enzymes Required for Siderophore Biosynthesis

    PubMed Central

    Engelhart, Curtis A.; Aldrich, Courtney C.

    2013-01-01

    MbtA catalyzes the first committed step of mycobactin biosynthesis in Mycobacterium tuberculosis (Mtb) and is responsible for the incorporation of salicylic acid into the mycobactin siderophores. 5′-O-[N-(Salicyl)sulfamoyl]adenosine (Sal-AMS) is an extremely potent nucleoside inhibitor of MbtA that possesses excellent activity against whole-cell Mtb, but suffers from poor bioavailability. In an effort to improve the bioavailability, we have designed four conformationally constrained analogues of Sal-AMS that remove two rotatable bonds and the ionized sulfamate group based on computational and structural studies. Herein we describe the synthesis, biochemical, and microbiological evaluation of chromone-, quinolone-, and benzoxazinone-3-sulfonamide derivatives of Sal-AMS. We developed new chemistry to assemble these three heterocycles from common β-ketosulfonamide intermediates. The synthesis of the chromone- and quinolone-3-sulfonamide intermediates features formylation of a β-ketosulfonamide employing dimethylformamide dimethyl acetal to afford an enaminone that can react intramolecularly with a phenol or intermolecularly with a primary amine via addition-elimination reaction(s). The benzoxazinone-3-sulfonamide was prepared by nitrosation of a β-ketosulfonamide followed by intramolecular nucleophilic aromatic substitution. Mitsunobu coupling of these bicyclic sulfonamides with a protected adenosine derivative followed by global deprotection provides a concise synthesis of the respective inhibitors. PMID:23805993

  5. Role for Ferredoxin:NAD(P)H Oxidoreductase (FprA) in Sulfate Assimilation and Siderophore Biosynthesis in Pseudomonads

    PubMed Central

    Glassing, Angela; Harper, Justin; Franklin, Michael J.

    2013-01-01

    Pyridine-2,6-bis(thiocarboxylate) (PDTC), produced by certain pseudomonads, is a sulfur-containing siderophore that binds iron, as well as a wide range of transition metals, and it affects the net hydrolysis of the environmental contaminant carbon tetrachloride. The pathway of PDTC biosynthesis has not been defined. Here, we performed a transposon screen of Pseudomonas putida DSM 3601 to identify genes necessary for PDTC production (Pdt phenotype). Transposon insertions within genes for sulfate assimilation (cysD, cysNC, and cysG [cobA2]) dominated the collection of Pdt mutations. In addition, two insertions were within the gene for the LysR-type transcriptional activator FinR (PP1637). Phenotypic characterization indicated that finR mutants were cysteine bradytrophs. The Pdt phenotype of finR mutants could be complemented by the known target of FinR regulation, fprA (encoding ferredoxin:NADP+ oxidoreductase), or by Escherichia coli cysJI (encoding sulfite reductase). These data indicate that fprA is necessary for effective sulfate assimilation by P. putida and that the effect of finR mutation on PDTC production was due to deficient expression of fprA and sulfite reduction. fprA expression in both P. putida and P. aeruginosa was found to be regulated by FinR, but in a manner dependent upon reduced sulfur sources, implicating FinR in sulfur regulatory physiology. The genes and phenotypes identified in this study indicated a strong dependence upon intracellular reduced sulfur/cysteine for PDTC biosynthesis and that pseudomonads utilize sulfite reduction enzymology distinct from that of E. coli and possibly similar to that of chloroplasts and other proteobacteria. PMID:23794620

  6. Structure and Biosynthesis of Amychelin, an Unusual Mixed-Ligand Siderophore from Amycolatopsis sp. AA4

    PubMed Central

    2011-01-01

    Actinobacteria generate a large number of structurally diverse small molecules with potential therapeutic value. Genomic analyses of this productive group of bacteria show that their genetic potential to manufacture small molecules exceeds their observed ability by roughly an order of magnitude, and this revelation has prompted a number of studies to identify members of the unknown majority. As a potential window into this cryptic secondary metabolome, pairwise assays for developmental interactions within a set of 20 sequenced actinomycetes were carried out. These assays revealed that Amycolatopsis sp. AA4, a so-called “rare” actinomycete, produces a novel siderophore, amychelin, which alters the developmental processes of several neighboring streptomycetes. Using this phenotype as an assay, we isolated amychelin and solved its structure by NMR and MS methods coupled with an X-ray crystallographic analysis of its Fe-complex. The iron binding affinity of amychelin was determined using EDTA competition assays, and a biosynthetic cluster was identified and annotated to provide a tentative biosynthetic scheme for amychelin. PMID:21699219

  7. Selective enrichment of environmental DNA libraries for genes encoding nonribosomal peptides and polyketides by phosphopantetheine transferase-dependent complementation of siderophore biosynthesis

    PubMed Central

    Charlop-Powers, Zachary; Banik, Jacob J.; Owen, Jeremy G.; Craig, Jeffrey W.; Brady, Sean F.

    2012-01-01

    The cloning of DNA directly from environmental samples provides a means to functionally access biosynthetic gene clusters present in the genomes of the large fraction of bacteria that remains recalcitrant to growth in the laboratory. Herein we demonstrate a method by which complementation of phosphopantetheine transferase deletion mutants can be used to restore siderophore biosynthesis and to therefore selectively enrich eDNA libraries for nonribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) gene sequences to unprecedented levels. The common use of NRPS/PKS-derived siderophores across bacterial taxa makes this method generalizable and should allow for the facile selective enrichment of NRPS/PKS-containing biosynthetic gene clusters from large environmental DNA libraries using a wide variety of phylogenetically diverse bacterial hosts. PMID:23072412

  8. Antitubercular Nucleosides that Inhibit Siderophore Biosynthesis: SAR of the Glycosyl Domain

    PubMed Central

    Somu, Ravindranadh V.; Wilson, Daniel; Bennett, Eric M.; Boshoff, Helena; Celia, Laura; Beck, Brian; Barry, Clifton E.; Aldrich, Courtney C.

    2008-01-01

    Tuberculosis (TB) is the leading cause of infectious disease mortality in the world by a bacterial pathogen. We previously demonstrated that a bisubstrate inhibitor of the adenylation enzyme MbtA, which is responsible for the second step of mycobactin biosynthesis, exhibited potent antitubercular activity. Here we systematically investigate the structure activity relationships of the bisubstrate inhibitor glycosyl domain resulting in the identification of a carbocyclic analogue that possesses a KIapp value of 2.3 nM and MIC99 values of 1.56 μM against M. tuberculosis H37Rv. The SAR data suggest the intriguing possibility that the bisubstrate inhibitors utilize a transporter for entry across the mycobacterial cell-envelope. Additionally, we report improved conditions for the expression of MbtA and biochemical analysis demonstrating that MbtA follows a random sequential enzyme mechanism for the adenylation half-reaction. PMID:17181146

  9. Total Biosynthesis and Diverse Applications of the Nonribosomal Peptide-Polyketide Siderophore Yersiniabactin

    PubMed Central

    Ahmadi, Mahmoud Kamal; Fawaz, Samar; Jones, Charles H.; Zhang, Guojian

    2015-01-01

    Yersiniabactin (Ybt) is a mixed nonribosomal peptide-polyketide natural product natively produced by the pathogen Yersinia pestis. The compound enables iron scavenging capabilities upon host infection and is biosynthesized by a nonribosomal peptide synthetase featuring a polyketide synthase module. This pathway has been engineered for expression and biosynthesis using Escherichia coli as a heterologous host. In the current work, the biosynthetic process for Ybt formation was improved through the incorporation of a dedicated step to eliminate the need for exogenous salicylate provision. When this improvement was made, the compound was tested in parallel applications that highlight the metal-chelating nature of the compound. In the first application, Ybt was assessed as a rust remover, demonstrating a capacity of ∼40% compared to a commercial removal agent and ∼20% relative to total removal capacity. The second application tested Ybt in removing copper from a variety of nonbiological and biological solution mixtures. Success across a variety of media indicates potential utility in diverse scenarios that include environmental and biomedical settings. PMID:26025901

  10. Crystallization and X-ray diffraction analysis of salicylate synthase, a chorismate-utilizing enyme involved in siderophore biosynthesis

    SciTech Connect

    Parsons, James F. Shi, Katherine; Calabrese, Kelly; Ladner, Jane E.

    2006-03-01

    Salicylate synthase, which catalyzes the first step in the synthesis of the siderophore yersiniabactin, has been crystallized. Diffraction data have been collected to 2.5 Å. Bacteria have evolved elaborate schemes that help them thrive in environments where free iron is severely limited. Siderophores such as yersiniabactin are small iron-scavenging molecules that are deployed by bacteria during iron starvation. Several studies have linked siderophore production and virulence. Yersiniabactin, produced by several Enterobacteriaceae, is derived from the key metabolic intermediate chorismic acid via its conversion to salicylate by salicylate synthase. Crystals of salicylate synthase from the uropathogen Escherichia coli CFT073 have been grown by vapour diffusion using polyethylene glycol as the precipitant. The monoclinic (P2{sub 1}) crystals diffract to 2.5 Å. The unit-cell parameters are a = 57.27, b = 164.07, c = 59.04 Å, β = 108.8°. The solvent content of the crystals is 54% and there are two molecules of the 434-amino-acid protein in the asymmetric unit. It is anticipated that the structure will reveal key details about the reaction mechanism and the evolution of salicylate synthase.

  11. Microbial Siderophores

    NASA Astrophysics Data System (ADS)

    Budzikiewicz, Herbert

    Iron is of great importance for many metabolic processes since the redox potential between its two valence states Fe2+ and Fe3+ lies within the range of physiological processes. Actually, iron is not a rare element, it is fourth in abundance in the earth crust, but it is not readily available for microorganisms. In the soil ferric oxide hydrates are formed at pH values around seven and the concentration of free Fe3+ is at best 10-17 mol/dm3 while about 10-6 mol/dm3 would be needed. In living organisms iron is usually strongly bound to peptidic substances such as transferrins. To increase the supply of soluble iron microorganisms other than those living in an acidic habitat may circumvent the problem by reduction of Fe3+ to Fe2+ (182), which seems to be of major importance for marine phytoplankton (151); see also amphiphilic marine bacteria (Sect.2.8) and Fe2+ binding ligands (Sect. 7) below. An important alternative is the production of Fe3+ chelating compounds, so-called siderophores. Siderophores are secondary metabolites with masses below 2,000 Da and a high affinity to Fe3+. Small iron-siderophore complexes can enter the cell via unspecific porins, larger ones need a transport system that recognizes the ferri-siderophore at the cell surface. In the cell, iron is released mostly by reduction to the less strongly bound Fe2+ state (137), and the free siderophore is re-exported ("shuttle mechanism"); for a modified shuttle system see pyoverdins (Sect. 2.1) and amonabactins (Sect. 2.7). Rarely the siderophore is degraded in the periplasmatic space as, e.g. enterobactin (Sect. 2.7). Alternatively Fe3+ is transferred at the cell surface from the ferri-siderophore to a trans-membrane transport system ("taxi mechanism"). A probably archaic and unspecific variety of the taxi mechanism comprises the reduction of Fe3+ at the cell surface (see ferrichrome A, Sect. 2.6 (99, 105)). The terms "shuttle" and "taxi mechanism" were coined by Raymond and Carrano (296).

  12. Acyl peptidic siderophores: structures, biosyntheses and post-assembly modifications.

    PubMed

    Kem, Michelle P; Butler, Alison

    2015-06-01

    Acyl peptidic siderophores are produced by a variety of bacteria and possess unique amphiphilic properties. Amphiphilic siderophores are generally produced in a suite where the iron(III)-binding headgroup remains constant while the fatty acid appendage varies by length and functionality. Acyl peptidic siderophores are commonly synthesized by non-ribosomal peptide synthetases; however, the method of peptide acylation during biosynthesis can vary between siderophores. Following biosynthesis, acyl siderophores can be further modified enzymatically to produce a more hydrophilic compound, which retains its ferric chelating abilities as demonstrated by pyoverdine from Pseudomonas aeruginosa and the marinobactins from certain Marinobacter species. Siderophore hydrophobicity can also be altered through photolysis of the ferric complex of certain β-hydroxyaspartic acid-containing acyl peptidic siderophores. PMID:25677460

  13. Microbial siderophore-based iron assimilation and therapeutic applications.

    PubMed

    Li, Kunhua; Chen, Wei-Hung; Bruner, Steven D

    2016-06-01

    Siderophores are structurally diverse, complex natural products that bind metals with extraordinary specificity and affinity. The acquisition of iron is critical for the survival and virulence of many pathogenic microbes and diverse strategies have evolved to synthesize, import and utilize iron. There has been a substantial increase of known siderophore scaffolds isolated and characterized in the past decade and the corresponding biosynthetic gene clusters have provided insight into the varied pathways involved in siderophore biosynthesis, delivery and utilization. Additionally, therapeutic applications of siderophores and related compounds are actively being developed. The study of biosynthetic pathways to natural siderophores augments the understanding of the complex mechanisms of bacterial iron acquisition, and enables a complimentary approach to address virulence through the interruption of siderophore biosynthesis or utilization by targeting the key enzymes to the siderophore pathways. PMID:27146331

  14. The Siderophore Metabolome of Azotobacter vinelandii

    PubMed Central

    Baars, Oliver; Zhang, Xinning

    2015-01-01

    In this study, we performed a detailed characterization of the siderophore metabolome, or “chelome,” of the agriculturally important and widely studied model organism Azotobacter vinelandii. Using a new high-resolution liquid chromatography-mass spectrometry (LC-MS) approach, we found over 35 metal-binding secondary metabolites, indicative of a vast chelome in A. vinelandii. These include vibrioferrin, a siderophore previously observed only in marine bacteria. Quantitative analyses of siderophore production during diazotrophic growth with different sources and availabilities of Fe showed that, under all tested conditions, vibrioferrin was present at the highest concentration of all siderophores and suggested new roles for vibrioferrin in the soil environment. Bioinformatic searches confirmed the capacity for vibrioferrin production in Azotobacter spp. and other bacteria spanning multiple phyla, habitats, and lifestyles. Moreover, our studies revealed a large number of previously unreported derivatives of all known A. vinelandii siderophores and rationalized their origins based on genomic analyses, with implications for siderophore diversity and evolution. Together, these insights provide clues as to why A. vinelandii harbors multiple siderophore biosynthesis gene clusters. Coupled with the growing evidence for alternative functions of siderophores, the vast chelome in A. vinelandii may be explained by multiple, disparate evolutionary pressures that act on siderophore production. PMID:26452553

  15. Catecholate Siderophores Protect Bacteria from Pyochelin Toxicity

    PubMed Central

    Adler, Conrado; Corbalán, Natalia S.; Seyedsayamdost, Mohammad R.; Pomares, María Fernanda; de Cristóbal, Ricardo E.; Clardy, Jon; Kolter, Roberto; Vincent, Paula A.

    2012-01-01

    Background Bacteria produce small molecule iron chelators, known as siderophores, to facilitate the acquisition of iron from the environment. The synthesis of more than one siderophore and the production of multiple siderophore uptake systems by a single bacterial species are common place. The selective advantages conferred by the multiplicity of siderophore synthesis remains poorly understood. However, there is growing evidence suggesting that siderophores may have other physiological roles besides their involvement in iron acquisition. Methods and Principal Findings Here we provide the first report that pyochelin displays antibiotic activity against some bacterial strains. Observation of differential sensitivity to pyochelin against a panel of bacteria provided the first indications that catecholate siderophores, produced by some bacteria, may have roles other than iron acquisition. A pattern emerged where only those strains able to make catecholate-type siderophores were resistant to pyochelin. We were able to associate pyochelin resistance to catecholate production by showing that pyochelin-resistant Escherichia coli became sensitive when biosynthesis of its catecholate siderophore enterobactin was impaired. As expected, supplementation with enterobactin conferred pyochelin resistance to the entE mutant. We observed that pyochelin-induced growth inhibition was independent of iron availability and was prevented by addition of the reducing agent ascorbic acid or by anaerobic incubation. Addition of pyochelin to E. coli increased the levels of reactive oxygen species (ROS) while addition of ascorbic acid or enterobactin reduced them. In contrast, addition of the carboxylate-type siderophore, citrate, did not prevent pyochelin-induced ROS increases and their associated toxicity. Conclusions We have shown that the catecholate siderophore enterobactin protects E. coli against the toxic effects of pyochelin by reducing ROS. Thus, it appears that catecholate

  16. Fungal siderophore metabolism with a focus on Aspergillus fumigatus

    PubMed Central

    2014-01-01

    Covering: up to 2014 Siderophores are chelators synthesized by microbes to sequester iron. This article summarizes the knowledge on the fungal siderophore metabolism with a focus on Aspergillus fumigatus. In recent years, A. fumigatus became a role model for fungal biosynthesis, uptake and degradation of siderophores as well as regulation of siderophore-mediated iron handling and the elucidation of siderophore functions. Siderophore functions comprise uptake, intracellular transport and storage of iron. This proved to be crucial not only for adaptation to iron starvation conditions but also for germination, asexual and sexual propagation, antioxidative defense, mutual interaction, microbial competition as well as virulence in plant and animal hosts. Recent studies also indicate the high potential of siderophores and its biosynthetic pathway to improve diagnosis and therapy of fungal infections. PMID:25140791

  17. Siderophore Biosynthesis but Not Reductive Iron Assimilation Is Essential for the Dimorphic Fungus Nomuraea rileyi Conidiation, Dimorphism Transition, Resistance to Oxidative Stress, Pigmented Microsclerotium Formation, and Virulence.

    PubMed

    Li, Yan; Wang, Zhongkang; Liu, Xuee; Song, Zhangyong; Li, Ren; Shao, Changwen; Yin, Youping

    2016-01-01

    Iron is an indispensable factor for the dimorphic insect pathogenic Nomuraea rileyi to form persistent microsclerotia which can replace conidia or blastospores for commercial mass production. There are two high affinity iron acquisition pathways in N. rileyi, siderophore-assisted iron mobilization and reductive iron assimilation systems. Transcription of the two iron uptake pathways related genes is induced under iron-limiting conditions. Stage-specific iron uptake-related genes expression during microsclerotia development shows siderophore-mediated iron acquisition genes are rigorously upregulated specifically during the formation and mature period while reductive iron assimilation related genes just display a higher expression at the late maturation period. Abrogation of reductive iron assimilation, by the deletion of the high affinity iron permease (NrFtrA), has no visible effect on microsclerotia biogenesis in N. rileyi. In sharp contrast, N. rileyi L-ornithine-N(5)-monooxygenase (NrSidA), required for synthesis of all siderophores, is absolutely necessary for the development of pigmented microsclerotia. In agreement with the lower intracellular iron contents of microsclerotia in ΔNrSidA strains, not only the pigments, but both the number and the biomass are also noticeably reduced. Certain concentration of ROS is required for promoting microsclerotia biogenesis. Combined with expression pattern analysis of related genes and quantitative of intracellular iron or extracellular siderophore in WT and mutants, these data demonstrate the lack of adequate intracellular iron caused by the loss of the siderophore results in the deficiency of ROS detoxication. Furthermore, ΔNrSidA strains show significantly increased sensitivity to hydrogen peroxide. Besides, NrSidA, but not NrFtrA, play a crucial role in vegetative growth under iron-limiting conditions, conidiation, and dimorphic switching. Remarkably, the slower growth of the ΔNrSidA strains in vivo due to a

  18. Siderophore Biosynthesis but Not Reductive Iron Assimilation Is Essential for the Dimorphic Fungus Nomuraea rileyi Conidiation, Dimorphism Transition, Resistance to Oxidative Stress, Pigmented Microsclerotium Formation, and Virulence

    PubMed Central

    Li, Yan; Wang, Zhongkang; Liu, Xuee; Song, Zhangyong; Li, Ren; Shao, Changwen; Yin, Youping

    2016-01-01

    Iron is an indispensable factor for the dimorphic insect pathogenic Nomuraea rileyi to form persistent microsclerotia which can replace conidia or blastospores for commercial mass production. There are two high affinity iron acquisition pathways in N. rileyi, siderophore-assisted iron mobilization and reductive iron assimilation systems. Transcription of the two iron uptake pathways related genes is induced under iron-limiting conditions. Stage-specific iron uptake-related genes expression during microsclerotia development shows siderophore-mediated iron acquisition genes are rigorously upregulated specifically during the formation and mature period while reductive iron assimilation related genes just display a higher expression at the late maturation period. Abrogation of reductive iron assimilation, by the deletion of the high affinity iron permease (NrFtrA), has no visible effect on microsclerotia biogenesis in N. rileyi. In sharp contrast, N. rileyi L-ornithine-N5-monooxygenase (NrSidA), required for synthesis of all siderophores, is absolutely necessary for the development of pigmented microsclerotia. In agreement with the lower intracellular iron contents of microsclerotia in ΔNrSidA strains, not only the pigments, but both the number and the biomass are also noticeably reduced. Certain concentration of ROS is required for promoting microsclerotia biogenesis. Combined with expression pattern analysis of related genes and quantitative of intracellular iron or extracellular siderophore in WT and mutants, these data demonstrate the lack of adequate intracellular iron caused by the loss of the siderophore results in the deficiency of ROS detoxication. Furthermore, ΔNrSidA strains show significantly increased sensitivity to hydrogen peroxide. Besides, NrSidA, but not NrFtrA, play a crucial role in vegetative growth under iron-limiting conditions, conidiation, and dimorphic switching. Remarkably, the slower growth of the ΔNrSidA strains in vivo due to a reduced

  19. Iron-Regulated Protein HupB of Mycobacterium tuberculosis Positively Regulates Siderophore Biosynthesis and Is Essential for Growth in Macrophages

    PubMed Central

    Pandey, Satya Deo; Choudhury, Mitali; Yousuf, Suhail; Wheeler, Paul R.; Gordon, Stephen V.; Ranjan, Akash

    2014-01-01

    Mycobacterium tuberculosis expresses the 28-kDa protein HupB (Rv2986c) and the Fe3+-specific high-affinity siderophores mycobactin and carboxymycobactin upon iron limitation. The objective of this study was to understand the functional role of HupB in iron acquisition. A hupB mutant strain of M. tuberculosis, subjected to growth in low-iron medium (0.02 μg Fe ml−1), showed a marked reduction of both siderophores with low transcript levels of the mbt genes encoding the MB biosynthetic machinery. Complementation of the mutant strain with hupB restored siderophore production to levels comparable to that of the wild type. We demonstrated the binding of HupB to the mbtB promoter by both electrophoretic mobility shift assays and DNA footprinting. The latter revealed the HupB binding site to be a 10-bp AT-rich region. While negative regulation of the mbt machinery by IdeR is known, this is the first report of positive regulation of the mbt operon by HupB. Interestingly, the mutant strain failed to survive inside macrophages, suggesting that HupB plays an important role in vivo. PMID:24610707

  20. The crystal structure of Rv1347c, a putative antibiotic resistance protein from Mycobacterium tuberculosis, reveals a GCN5-related fold and suggests an alternative function in siderophore biosynthesis

    SciTech Connect

    Card, G L; Peterson, N A; Smith, C A; Rupp, B; Schick, B M; Baker, E N

    2005-02-15

    Mycobacterium tuberculosis, the cause of TB, is a devastating human pathogen. The emergence of multi-drug resistance in recent years has prompted a search for new drug targets and for a better understanding of mechanisms of resistance. Here we focus on the gene product of an open reading frame from M. tuberculosis, Rv1347c, which is annotated as a putative aminoglycoside N-acetyltransferase. The Rv1347c protein does not show this activity, however, and we show from its crystal structure, coupled with functional and bioinformatic data, that its most likely role is in the biosynthesis of mycobactin, the M. tuberculosis siderophore. The crystal structure of Rv1347c was determined by MAD phasing from selenomethionine-substituted protein and refined at 2.2 {angstrom} resolution (R = 0.227, R{sub free} = 0.257). The protein is monomeric, with a fold that places it in the GCN5-related N-acetyltransferase (GNAT) family of acyltransferases. Features of the structure are an acylCoA binding site that is shared with other GNAT family members, and an adjacent hydrophobic channel leading to the surface that could accommodate long-chain acyl groups. Modeling the postulated substrate, the N{sup {var_epsilon}}-hydroxylysine side chain of mycobactin, into the acceptor substrate binding groove identifies two residues at the active site, His130 and Asp168, that have putative roles in substrate binding and catalysis.

  1. 2-Aryl-8-aza-3-deazaadenosine analogues of 5'-O-[N-(salicyl)sulfamoyl]adenosine: Nucleoside antibiotics that block siderophore biosynthesis in Mycobacterium tuberculosis.

    PubMed

    Krajczyk, Anna; Zeidler, Joanna; Januszczyk, Piotr; Dawadi, Surendra; Boshoff, Helena I; Barry, Clifton E; Ostrowski, Tomasz; Aldrich, Courtney C

    2016-07-15

    A series of 5'-O-[N-(salicyl)sulfamoyl]-2-aryl-8-aza-3-deazaadenosines were designed to block mycobactin biosynthesis in Mycobacterium tuberculosis (Mtb) through inhibition of the essential adenylating enzyme MbtA. The synthesis of the 2-aryl-8-aza-3-deazaadenosine nucleosides featured sequential copper-free palladium-catalyzed Sonogashira coupling of a precursor 4-cyano-5-iodo-1,2,3-triazolonucleoside with terminal alkynes and a Minakawa-Matsuda annulation reaction. These modified nucleosides were shown to inhibit MbtA with apparent Ki values ranging from 6.1 to 25nM and to inhibit Mtb growth under iron-deficient conditions with minimum inhibitory concentrations ranging from 12.5 to >50μM. PMID:27265685

  2. Detection of photoactive siderophore biosynthetic genes in the marine environment.

    PubMed

    Gärdes, Astrid; Triana, Christopher; Amin, Shady A; Green, David H; Romano, Ariel; Trimble, Lyndsay; Carrano, Carl J

    2013-06-01

    Iron is an essential element for oceanic microbial life but its low bioavailability limits microorganisms in large areas of the oceans. To acquire this metal many marine bacteria produce organic chelates that bind and transport iron (siderophores). While it has been hypothesized that the global production of siderophores by heterotrophic bacteria and some cyanobacteria constitutes the bulk of organic ligands binding iron in the ocean because stability constants of siderophores and these organic ligands are similar, and because ligand concentrations rise sharply in response to iron fertilization events, direct evidence for this proposal is lacking. This lack is due to the difficulty in characterizing these ligands due both to their extremely low concentrations and their highly heterogeneous nature. The situation for characterizing photoactive siderophores in situ is more problematic because of their expected short lifetimes in the photic zone. An alternative approach is to make use of high sensitivity molecular technology (qPCR) to search for siderophore biosynthesis genes related to the production of photoactive siderophores. In this way one can access their "biochemical potential" and utilize this information as a proxy for the presence of these siderophores in the marine environment. Here we show, using qPCR primers designed to detect biosynthetic genes for the siderophores vibrioferrin, petrobactin and aerobactin that such genes are widespread and based on their abundance, the "biochemical potential" for photoactive siderophore production is significant. Concurrently we also briefly examine the microbial biodiversity responsible for such production as a function of depth and location across a North Atlantic transect. PMID:23700243

  3. Inhibition of Siderophore Biosynthesis in Mycobacterium tuberculosis with Nucleoside Bisubstrate Analogues: Structure–Activity Relationships of the Nucleobase Domain of 5′-O-[N-(Salicyl)sulfamoyl]adenosine

    PubMed Central

    Neres, João; Labello, Nicholas P.; Somu, Ravindranadh V.; Boshoff, Helena I.; Wilson, Daniel J.; Vannada, Jagadeshwar; Chen, Liqiang; Barry, Clifton E.; Bennett, Eric M.; Aldrich, Courtney C.

    2009-01-01

    5′-O-[N-(salicyl)sulfamoyl]adenosine (Sal-AMS) is a prototype for a new class of antitubercular agents that inhibit the aryl acid adenylating enzyme (AAAE) known as MbtA involved in biosynthesis of the mycobactins. Herein, we report the structure-based design, synthesis, biochemical, and biological evaluation of a comprehensive and systematic series of analogues, exploring the structure–activity relationship of the purine nucleobase domain of Sal-AMS. Significantly, 2-phenyl-Sal-AMS derivative 26 exhibited exceptionally potent antitubercular activity with an MIC99 under iron-deficient conditions of 0.049 µM while the N-6-cyclopropyl-Sal-AMS 16 led to improved potency and to a 64-enhancement in activity under iron-deficient conditions relative to iron-replete conditions, a phenotype concordant with the designed mechanism of action. The most potent MbtA inhibitors disclosed here display in vitro antitubercular activity superior to most current first line TB drugs, and these compounds are also expected to be useful against a wide range of pathogens that require aryl-capped siderphores for virulence. PMID:18690677

  4. Siderophore production by Pseudomonas pseudomallei.

    PubMed Central

    Yang, H M; Chaowagul, W; Sokol, P A

    1991-01-01

    Eighty-four strains of Pseudomonas pseudomallei isolated from patients with melioidosis were examined for siderophore production. All the strains were shown to produce siderophore both on chrome azurol S agar plates and in liquid medium under iron-deficient conditions. Chemical assays indicated that the siderophore belongs to the hydroxamate class. Addition of iron to the culture medium resulted in increased culture growth with markedly decreased yield of siderophore. Siderophore produced by strain U7 was purified by gel filtration chromatography, and the molecular weight was estimated to be 1,000. When this partially purified siderophore was added to culture medium, it promoted iron uptake by P. pseudomallei in the presence of EDTA and enhanced growth of the organism in the presence of transferrin. We have given this siderophore the trivial name malleobactin. PMID:1825486

  5. Precursor activation in a pyoverdine biosynthesis.

    PubMed

    Menhart, N; Viswanatha, T

    1990-03-29

    The siderophore produced by Azotobacter vinelandii strain UW belongs to a large family of peptidic siderophores collectively called pyoverdines. The biosynthesis of the peptidyl moiety of this siderophore was shown to involve activation of the constituent amino acids as their adenylates, as demonstrated by amino acid-dependent ATP-[32P]pyrophosphate exchange. The enzyme system responsible for this activation was partially purified by chromatographic techniques. PMID:2156571

  6. Siderophore-mediated iron acquisition in Bacillus anthracis and related strains.

    PubMed

    Hotta, Kinya; Kim, Chu-Young; Fox, David T; Koppisch, Andrew T

    2010-07-01

    Recent observations have shed light on some of the endogenous iron-acquisition mechanisms of members of the Bacillus cereus sensu lato group. In particular, pathogens in the B. cereus group use siderophores with both unique chemical structures and biological roles. This review will focus on recent discoveries in siderophore biosynthesis and biology in this group, which contains numerous human pathogens, most notably the causative agent of anthrax, Bacillus anthracis. PMID:20466767

  7. Bisucaberin B, a linear hydroxamate class siderophore from the marine bacterium Tenacibaculum mesophilum.

    PubMed

    Fujita, Masaki J; Nakano, Koji; Sakai, Ryuichi

    2013-01-01

    A siderophore, named bisucaberin B, was isolated from Tenacibaculum mesophilum bacteria separated from a marine sponge collected in the Republic of Palau. Using spectroscopic and chemical methods, the structure of bisucaberin B (1) was clearly determined to be a linear dimeric hydroxamate class siderophore. Although compound 1 is an open form of the known macrocyclic dimer bisucaberin (2), and was previously described as a bacterial degradation product of desferrioxamine B (4), the present report is the first description of the de novo biosynthesis of 1. To the best of our knowledge, compound 1 is the first chemically characterized siderophore isolated from a bacterium belonging to the phylum Bacteroidetes. PMID:23549298

  8. Synthesis of siderophores by plant-associated metallotolerant bacteria under exposure to Cd(2.).

    PubMed

    Złoch, Michał; Thiem, Dominika; Gadzała-Kopciuch, Renata; Hrynkiewicz, Katarzyna

    2016-08-01

    Rhizosphere and endophytic bacteria are well known producers of siderophores, organic compounds that chelate ferric iron (Fe(3+)), and therefore play an important role in plant growth promotion in metalliferous areas, thereby improving bioremediation processes. However, in addition to their primary function in iron mobilization, siderophores also have the capacity to chelate other heavy metals, such as Al(3+), Zn(2+), Cu(2+), Pb(2+) and Cd(2+), that can affect homeostasis and the heavy metal tolerance of microorganisms. The main goal of our study was to select the most efficient siderophore-producing bacterial strains isolated from the roots (endophytes) and rhizosphere of Betula pendula L. and Alnus glutinosa L. growing at two heavy metal contaminated sites in southern Poland. Siderophore biosynthesis of these strains in the presence of increasing concentrations of Cd(2+) (0, 0.5, 1, 2 and 3 mM) under iron-deficiency conditions was analysed using spectrophotometric chemical tests for hydroxamates, catecholates and phenolates, as well as the separation of bacterial siderophores by HPLC and characterization of their structure by UHPLC-QTOF/MS. We proved that (i) siderophore-producing bacterial strains seems to be more abundant in the rhizosphere (47%) than in root endophytes (18%); (ii) the strains most effective at siderophore synthesis belonged to the genus Streptomyces and were able to secrete three types of siderophores under Cd(2+) stress: hydroxamates, catecholates and phenolates; (iii) in general, the addition of Cd(2+) enhanced siderophore synthesis, particularly ferrioxamine B synthesis, which may indicate that siderophores play a significant role in tolerance to Cd(2+) in Streptomyces sp. PMID:27183333

  9. Role of the FeoB Protein and Siderophore in Promoting Virulence of Xanthomonas oryzae pv. oryzae on Rice▿

    PubMed Central

    Pandey, Alok; Sonti, Ramesh V.

    2010-01-01

    Xanthomonas oryzae pv. oryzae causes bacterial blight, a serious disease of rice. Our analysis revealed that the X. oryzae pv. oryzae genome encodes genes responsible for iron uptake through FeoB (homolog of the major bacterial ferrous iron transporter) and a siderophore. A mutation in the X. oryzae pv. oryzae feoB gene causes severe virulence deficiency, growth deficiency in iron-limiting medium, and constitutive production of a siderophore. We identified an iron regulated xss gene cluster, in which xssABCDE (Xanthomonas siderophore synthesis) and xsuA (Xanthomonas siderophore utilization) genes encode proteins involved in biosynthesis and utilization of X. oryzae pv. oryzae siderophore. Mutations in the xssA, xssB, and xssE genes cause siderophore deficiency and growth restriction under iron-limiting conditions but are virulence proficient. An xsuA mutant displayed impairment in utilization of native siderophore, suggesting that XsuA acts as a specific receptor for a ferric-siderophore complex. Histochemical and fluorimetric assays with gusA fusions indicate that, during in planta growth, the feoB gene is expressed and that the xss operon is not expressed. This study represents the first report describing a role for feoB in virulence of any plant-pathogenic bacterium and the first functional characterization of a siderophore-biosynthetic gene cluster in any xanthomonad. PMID:20382771

  10. Bacterial Growth Stimulation with Exogenous Siderophore and Synthetic N-Acyl Homoserine Lactone Autoinducers under Iron-Limited and Low-Nutrient Conditions

    PubMed Central

    Guan, Le Luo; Onuki, Hiroyuki; Kamino, Kei

    2000-01-01

    The growth of marine bacteria under iron-limited conditions was investigated. Neither siderophore production nor bacterial growth was detected for Pelagiobacter sp. strain V0110 when Fe(III) was present in the culture medium at a concentration of <1.0 μM. However, the growth of V0110 was strongly stimulated by the presence of trace amounts of exogenous siderophore from an alpha proteobacterium, V0902, and 1 nM N-acyl-octanoylhomoserine lactone (C8-HSL), which is known as a quorum-sensing chemical signal. Even though the iron-binding functionality of a hydroxamate siderophore was undetected in the supernatant of V0902, a hydroxamate siderophore was detected in the supernatant of V0110 under the above conditions. These results indicated that hydroxamate siderophore biosynthesis by V0110 began in response to the exogenous siderophore from V0902 when in the presence of C8-HSL; however, C8-HSL production by V0110 and V0902 was not detected. Direct interaction between V0902 and V0110 through siderophore from V0902 was observed in the dialyzing culture. Similar stimulated growth by exogenous siderophore and HSL was also observed in other non-siderophore-producing bacteria isolated from marine sponges and seawater. The requirement of an exogenous siderophore and an HSL for heterologous siderophore production indicated the possibility that cell-cell communication between different species was occurring. PMID:10877770

  11. Contribution of Siderophore Systems to Growth and Urinary Tract Colonization of Asymptomatic Bacteriuria Escherichia coli

    PubMed Central

    Watts, Rebecca E.; Totsika, Makrina; Challinor, Victoria L.; Mabbett, Amanda N.; Ulett, Glen C.; De Voss, James J.

    2012-01-01

    The molecular mechanisms that define asymptomatic bacteriuria (ABU) Escherichia coli colonization of the human urinary tract remain to be properly elucidated. Here, we utilize ABU E. coli strain 83972 as a model to dissect the contribution of siderophores to iron acquisition, growth, fitness, and colonization of the urinary tract. We show that E. coli 83972 produces enterobactin, salmochelin, aerobactin, and yersiniabactin and examine the role of these systems using mutants defective in siderophore biosynthesis and uptake. Enterobactin and aerobactin contributed most to total siderophore activity and growth in defined iron-deficient medium. No siderophores were detected in an 83972 quadruple mutant deficient in all four siderophore biosynthesis pathways; this mutant did not grow in defined iron-deficient medium but grew in iron-limited pooled human urine due to iron uptake via the FecA ferric citrate receptor. In a mixed 1:1 growth assay with strain 83972, there was no fitness disadvantage of the 83972 quadruple biosynthetic mutant, demonstrating its capacity to act as a “cheater” and utilize siderophores produced by the wild-type strain for iron uptake. An 83972 enterobactin/salmochelin double receptor mutant was outcompeted by 83972 in human urine and the mouse urinary tract, indicating a role for catecholate receptors in urinary tract colonization. PMID:21930757

  12. The interchangeability of siderophores in Staphylococci.

    PubMed

    Sobis-Glinkowska, M; Lisiecki, P; Mikucki, J

    1995-01-01

    The functional interchangeability of siderophores among 90 strains belonging to 26 species of genus Staphylococcus was tested. Among the species which synthesized siderophores most commonly utilized (the best donors) were S. epidermidis, S. aureus, S. cohnii, S. hominis and S. simulans. The species which utilized the widest range of exogenous siderophores from other species (the best recipient) were S. epidermidis, S. saprophyticus, S. kloosii and S. schleiferi. The donor and recipient of the widest range of siderophores was S. epidermidis. The potential ability to exchange siderophores as well as the range of their functional interchangeability in staphylococci were found as the features characterizing individual strains. PMID:8906932

  13. A Mammalian Siderophore Synthesized by an Enzyme with a Bacterial Homologue Involved in Enterobactin Production

    PubMed Central

    Devireddy, Laxminarayana R.; Hart, Daniel O.; Goetz, David; Green, Michael R.

    2010-01-01

    SUMMARY Intracellular iron homeostasis is critical for survival and proliferation. Lipocalin 24p3 is an iron trafficking protein that binds iron through association with a bacterial siderophore, such as enterobactin, or a postulated mammalian siderophore. Here we show that the iron-binding moiety of the 24p3-associated mammalian siderophore is 2,5-dihydroxybenzoic acid (2,5-DHBA), which is similar to 2,3-DHBA, the iron-binding component of enterobactin. We find that the murine enzyme responsible for 2,5-DHBA synthesis is the homologue of bacterial EntA, which catalyzes 2,3-DHBA production during enterobactin biosynthesis. RNA interference-mediated knockdown of the murine homologue of EntA results in siderophore depletion. Mammalian cells lacking the siderophore accumulate abnormally high amounts of cytoplasmic iron, resulting in elevated levels of reactive oxygen species, whereas the mitochondria are iron deficient. Siderophore-depleted mammalian cells and zebrafish embryos fail to synthesize heme, an iron-dependent mitochondrial process. Our results reveal features of intracellular iron homeostasis that are conserved from bacteria through humans. PMID:20550936

  14. Iron acquisition with the natural siderophore enantiomers pyochelin and enantio-pyochelin in Pseudomonas species.

    PubMed

    Youard, Zeb A; Wenner, Nicolas; Reimmann, Cornelia

    2011-06-01

    The bacterial siderophore pyochelin is composed 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 ensured at two levels: (i) by the outer membrane siderophore receptors and (ii) by the cytosolic PchR regulators. PMID:21188474

  15. Individual and combined roles of malonichrome, ferricrocin, and TAFC siderophores in Fusarium graminearum pathogenic and sexual development

    PubMed Central

    Oide, Shinichi; Berthiller, Franz; Wiesenberger, Gerlinde; Adam, Gerhard; Turgeon, B. Gillian

    2015-01-01

    Intra- and extracellular iron-chelating siderophores produced by fungal non-ribosomal peptide synthetases have been shown to be involved in reproductive and pathogenic developmental processes and in iron and oxidative stress management. Here we report individual and combined contributions of three of these metabolites to developmental success of the destructive cereal pathogen Fusarium graminearum. In previous work, we determined that deletion of the NPS2 gene, responsible for intracellular siderophore biosynthesis, results in inability to produce sexual spores when mutants of this homothallic ascomycete are selfed. Deletion of the NPS6 gene, required for extracellular siderophore biosynthesis, does not affect sexual reproduction but results in sensitivity to iron starvation and oxidative stress and leads to reduced virulence to the host. Building on this, we report that double mutants lacking both NPS2 and NPS6 are augmented in all collective phenotypes of single deletion strains (i.e., abnormal sexual and pathogenic development, hypersensitivity to oxidative and iron-depletion stress), which suggests overlap of function. Using comparative biochemical analysis of wild-type and mutant strains, we show that NPS1, a third gene associated with siderophore biosynthesis, is responsible for biosynthesis of a second extracellular siderophore, malonichrome. nps1 mutants fail to produce this metabolite. Phenotypic characterization reveals that, although single nps1 mutants are like wild-type with respect to sexual development, hypersensitivity to ROS and iron-depletion stress, and virulence to the host, triple nps1nps2nps6 deletion strains, lacking all three siderophores, are even more impaired in these attributes than double nps2nps6 strains. Thus, combinatorial mutants lacking key iron-associated genes uncovered malonichrome function. The intimate connection between presence/absence of siderophores and resistance/sensitivity to ROS is central to sexual and pathogenic

  16. Individual and combined roles of malonichrome, ferricrocin, and TAFC siderophores in Fusarium graminearum pathogenic and sexual development.

    PubMed

    Oide, Shinichi; Berthiller, Franz; Wiesenberger, Gerlinde; Adam, Gerhard; Turgeon, B Gillian

    2014-01-01

    Intra- and extracellular iron-chelating siderophores produced by fungal non-ribosomal peptide synthetases have been shown to be involved in reproductive and pathogenic developmental processes and in iron and oxidative stress management. Here we report individual and combined contributions of three of these metabolites to developmental success of the destructive cereal pathogen Fusarium graminearum. In previous work, we determined that deletion of the NPS2 gene, responsible for intracellular siderophore biosynthesis, results in inability to produce sexual spores when mutants of this homothallic ascomycete are selfed. Deletion of the NPS6 gene, required for extracellular siderophore biosynthesis, does not affect sexual reproduction but results in sensitivity to iron starvation and oxidative stress and leads to reduced virulence to the host. Building on this, we report that double mutants lacking both NPS2 and NPS6 are augmented in all collective phenotypes of single deletion strains (i.e., abnormal sexual and pathogenic development, hypersensitivity to oxidative and iron-depletion stress), which suggests overlap of function. Using comparative biochemical analysis of wild-type and mutant strains, we show that NPS1, a third gene associated with siderophore biosynthesis, is responsible for biosynthesis of a second extracellular siderophore, malonichrome. nps1 mutants fail to produce this metabolite. Phenotypic characterization reveals that, although single nps1 mutants are like wild-type with respect to sexual development, hypersensitivity to ROS and iron-depletion stress, and virulence to the host, triple nps1nps2nps6 deletion strains, lacking all three siderophores, are even more impaired in these attributes than double nps2nps6 strains. Thus, combinatorial mutants lacking key iron-associated genes uncovered malonichrome function. The intimate connection between presence/absence of siderophores and resistance/sensitivity to ROS is central to sexual and pathogenic

  17. Evaluation of photo-reactive siderophore producing bacteria before, during and after a bloom of the dinoflagellate Lingulodinium polyedrum.

    PubMed

    Yarimizu, Kyoko; Polido, Geraldine; Gärdes, Astrid; Carter, Melissa L; Hilbern, Mary; Carrano, Carl J

    2014-06-01

    Evidence is increasing for a mutualistic relationship between phytoplankton and heterotrophic marine bacteria. It has been proposed that bacteria producing photoactive iron binding compounds known as siderophores could play an important role in such mutualistic associations by producing bioavailable iron utilizable by phytoplankton and in exchange receive autotrophically derived DOM. In order to understand the potential role photoactive siderophores might be playing in bacterial-algal mutualism or marine biogeochemistry in general, it is important to be able to detect and quantify their presence in various environments. One approach to accomplish that end is to make use of high sensitivity genomics technology (qPCR) to search for siderophore biosynthesis genes related to the production of photoactive siderophores. In this way one can access their "biochemical potential" and utilize this information as a proxy for the presence of these siderophores in the marine environment. In this report we studied the correlation of the presence of bacteria producing one of the three photoactive siderophores relative to total bacterial and dinoflagellate numbers from surface water at the Scripps Pier before, during, and after fall bloom of the dinoflagellate Lingulodinium polyedrum. We believe that these findings will aid us in gauging the importance of photoactive siderophores in the marine environment and in harmful algal bloom dynamics in particular. PMID:24760287

  18. Characterization of microbial siderophores by mass spectrometry.

    PubMed

    Pluháček, Tomáš; Lemr, Karel; Ghosh, Dipankar; Milde, David; Novák, Jiří; Havlíček, Vladimír

    2016-01-01

    Siderophores play important roles in microbial iron piracy, and are applied as infectious disease biomarkers and novel pharmaceutical drugs. Inductively coupled plasma and molecular mass spectrometry (ICP-MS) combined with high resolution separations allow characterization of siderophores in complex samples taking advantages of mass defect data filtering, tandem mass spectrometry, and iron-containing compound quantitation. The enrichment approaches used in siderophore analysis and current ICP-MS technologies are reviewed. The recent tools for fast dereplication of secondary metabolites and their databases are reported. This review on siderophores is concluded with their recent medical, biochemical, geochemical, and agricultural applications in mass spectrometry context. PMID:25980644

  19. Pseudomonas fluorescens Pirates both Ferrioxamine and Ferricoelichelin Siderophores from Streptomyces ambofaciens

    PubMed Central

    Galet, Justine; Deveau, Aurélie; Hôtel, Laurence; Frey-Klett, Pascale; Leblond, Pierre

    2015-01-01

    Iron is essential in many biological processes. However, its bioavailability is reduced in aerobic environments, such as soil. To overcome this limitation, microorganisms have developed different strategies, such as iron chelation by siderophores. Some bacteria have even gained the ability to detect and utilize xenosiderophores, i.e., siderophores produced by other organisms. We illustrate an example of such an interaction between two soil bacteria, Pseudomonas fluorescens strain BBc6R8 and Streptomyces ambofaciens ATCC 23877, which produce the siderophores pyoverdine and enantiopyochelin and the siderophores desferrioxamines B and E and coelichelin, respectively. During pairwise cultures on iron-limiting agar medium, no induction of siderophore synthesis by P. fluorescens BBc6R8 was observed in the presence of S. ambofaciens ATCC 23877. Cocultures with a Streptomyces mutant strain that produced either coelichelin or desferrioxamines, as well as culture in a medium supplemented with desferrioxamine B, resulted in the absence of pyoverdine production; however, culture with a double mutant deficient in desferrioxamines and coelichelin production did not. This strongly suggests that P. fluorescens BBbc6R8 utilizes the ferrioxamines and ferricoelichelin produced by S. ambofaciens as xenosiderophores and therefore no longer activates the production of its own siderophores. A screening of a library of P. fluorescens BBc6R8 mutants highlighted the involvement of the TonB-dependent receptor FoxA in this process: the expression of foxA and genes involved in the regulation of its biosynthesis was induced in the presence of S. ambofaciens. In a competitive environment, such as soil, siderophore piracy could well be one of the driving forces that determine the outcome of microbial competition. PMID:25724953

  20. Pseudomonas fluorescens pirates both ferrioxamine and ferricoelichelin siderophores from Streptomyces ambofaciens.

    PubMed

    Galet, Justine; Deveau, Aurélie; Hôtel, Laurence; Frey-Klett, Pascale; Leblond, Pierre; Aigle, Bertrand

    2015-05-01

    Iron is essential in many biological processes. However, its bioavailability is reduced in aerobic environments, such as soil. To overcome this limitation, microorganisms have developed different strategies, such as iron chelation by siderophores. Some bacteria have even gained the ability to detect and utilize xenosiderophores, i.e., siderophores produced by other organisms. We illustrate an example of such an interaction between two soil bacteria, Pseudomonas fluorescens strain BBc6R8 and Streptomyces ambofaciens ATCC 23877, which produce the siderophores pyoverdine and enantiopyochelin and the siderophores desferrioxamines B and E and coelichelin, respectively. During pairwise cultures on iron-limiting agar medium, no induction of siderophore synthesis by P. fluorescens BBc6R8 was observed in the presence of S. ambofaciens ATCC 23877. Cocultures with a Streptomyces mutant strain that produced either coelichelin or desferrioxamines, as well as culture in a medium supplemented with desferrioxamine B, resulted in the absence of pyoverdine production; however, culture with a double mutant deficient in desferrioxamines and coelichelin production did not. This strongly suggests that P. fluorescens BBbc6R8 utilizes the ferrioxamines and ferricoelichelin produced by S. ambofaciens as xenosiderophores and therefore no longer activates the production of its own siderophores. A screening of a library of P. fluorescens BBc6R8 mutants highlighted the involvement of the TonB-dependent receptor FoxA in this process: the expression of foxA and genes involved in the regulation of its biosynthesis was induced in the presence of S. ambofaciens. In a competitive environment, such as soil, siderophore piracy could well be one of the driving forces that determine the outcome of microbial competition. PMID:25724953

  1. Mammalian Siderophores, Siderophore-binding Lipocalins, and the Labile Iron Pool*

    PubMed Central

    Correnti, Colin; Strong, Roland K.

    2012-01-01

    Bacteria use tight-binding, ferric-specific chelators called siderophores to acquire iron from the environment and from the host during infection; animals use proteins such as transferrin and ferritin to transport and store iron. Recently, candidate compounds that could serve endogenously as mammalian siderophore equivalents have been identified and characterized through associations with siderocalin, the only mammalian siderophore-binding protein currently known. Siderocalin, an antibacterial protein, acts by sequestering iron away from infecting bacteria as siderophore complexes. Candidate endogenous siderophores include compounds that only effectively transport iron as ternary complexes with siderocalin, explaining pleiotropic activities in normal cellular processes and specific disease states. PMID:22389496

  2. Unsuspected control of siderophore production by N-acetylglucosamine in streptomycetes.

    PubMed

    Craig, Matthias; Lambert, Stéphany; Jourdan, Samuel; Tenconi, Elodie; Colson, Séverine; Maciejewska, Marta; Ongena, Marc; Martin, Juan Francisco; van Wezel, Gilles; Rigali, Sébastien

    2012-10-01

    Iron is one of the most abundant elements on earth but is found in poorly soluble forms hardly accessible to microorganisms. To subsist, they have developed iron-chelating molecules called siderophores that capture this element in the environment and the resulting complexes are internalized by specific uptake systems. While biosynthesis of siderophores in many bacteria is regulated by iron availability and oxidative stress, we describe here a new type of regulation of siderophore production. We show that in Streptomyces coelicolor, their production is also controlled by N-acetylglucosamine (GlcNAc) via the direct transcriptional repression of the iron utilization repressor dmdR1 by DasR, the GlcNAc utilization regulator. This regulatory nutrient-metal relationship is conserved among streptomycetes, which indicates that the link between GlcNAc utilization and iron uptake repression, however unsuspected, is the consequence of a successful evolutionary process. We describe here the molecular basis of a novel inhibitory mechanism of siderophore production that is independent of iron availability. We speculate that the regulatory connection between GlcNAc and siderophores might be associated with the competition for iron between streptomycetes and their fungal soil competitors, whose cell walls are built from the GlcNAc-containing polymer chitin. Alternatively, GlcNAc could emanate from streptomycetes' own peptidoglycan that goes through intense remodelling throughout their life cycle, thereby modulating the iron supply according to specific needs at different stages of their developmental programme. PMID:23760896

  3. Monitoring iron uptake by siderophores.

    PubMed

    Hoegy, Françoise; Schalk, Isabelle J

    2014-01-01

    Iron is an important element for almost all forms of life. In order to get access to this essential nutriment, Pseudomonads produce two major siderophores, pyoverdine PVD and pyochelin (PCH). Uptake of iron in bacterial cells can be monitored accurately using (55)Fe. Bacteria cells are incubated in the presence of either PVD or PCH loaded with (55)Fe. After incubation, extracellular iron ions are separated from those accumulated in the bacteria cells by either centrifugation or filtration on glass microfiber filters, for the PCH and PVD assays, respectively. (55)Fe contained in the harvested cells on the filter or in the cell pellet is counted in scintillation cocktail. The number of moles of (55)Fe transported can be determined using the specific activity of the radionuclide. PMID:24818918

  4. Siderophore production by marine-derived fungi

    PubMed Central

    Holinsworth, Brian; Martin, Jessica D.

    2009-01-01

    Siderophore production by marine-derived fungi has not been extensively explored. Three studies have investigated the ability of marine-derived fungi to produce siderophores in response to iron limitation [(Vala et al. in Indian J Mar Sci 29:339-340, 2000; Can J Microbiol 52:603-607, 2006); Baakza et al. in J Exp Mar Biol Ecol 311:1-9, 2004]. In all, 24 of 28 marine fungal strains were found to secrete hydroxamate or carboxylate siderophores; no evidence was found for production of catecholate siderophores. These studies did not determine the structures of the iron-binding compounds. More recently, a study of the natural products secreted by a marine Penicillium bilaii revealed that this strain produced the rare catecholate siderophore pistillarin when grown under relatively high iron concentrations (Capon et al. J Nat Prod 70:1746-1752, 2007). Additionally, the production of rhizoferrin by a marine isolate of Cunninghamella elegans (ATCC36112) is reported in this manuscript. The current state of knowledge about marine fungal siderophores is reviewed in light of these promising results. PMID:19350395

  5. Production of the Catechol Type Siderophore Bacillibactin by the Honey Bee Pathogen Paenibacillus larvae

    PubMed Central

    Garcia-Gonzalez, Eva; Poppinga, Lena; Süssmuth, Roderich D.; Genersch, Elke

    2014-01-01

    The Gram-positive bacterium Paenibacillus larvae is the etiological agent of American Foulbrood. This bacterial infection of honey bee brood is a notifiable epizootic posing a serious threat to global honey bee health because not only individual larvae but also entire colonies succumb to the disease. In the recent past considerable progress has been made in elucidating molecular aspects of host pathogen interactions during pathogenesis of P. larvae infections. Especially the sequencing and annotation of the complete genome of P. larvae was a major step forward and revealed the existence of several giant gene clusters coding for non-ribosomal peptide synthetases which might act as putative virulence factors. We here present the detailed analysis of one of these clusters which we demonstrated to be responsible for the biosynthesis of bacillibactin, a P. larvae siderophore. We first established culture conditions allowing the growth of P. larvae under iron-limited conditions and triggering siderophore production by P. larvae. Using a gene disruption strategy we linked siderophore production to the expression of an uninterrupted bacillibactin gene cluster. In silico analysis predicted the structure of a trimeric trithreonyl lactone (DHB-Gly-Thr)3 similar to the structure of bacillibactin produced by several Bacillus species. Mass spectrometric analysis unambiguously confirmed that the siderophore produced by P. larvae is identical to bacillibactin. Exposure bioassays demonstrated that P. larvae bacillibactin is not required for full virulence of P. larvae in laboratory exposure bioassays. This observation is consistent with results obtained for bacillibactin in other pathogenic bacteria. PMID:25237888

  6. Production of the catechol type siderophore bacillibactin by the honey bee pathogen Paenibacillus larvae.

    PubMed

    Hertlein, Gillian; Müller, Sebastian; Garcia-Gonzalez, Eva; Poppinga, Lena; Süssmuth, Roderich D; Genersch, Elke

    2014-01-01

    The Gram-positive bacterium Paenibacillus larvae is the etiological agent of American Foulbrood. This bacterial infection of honey bee brood is a notifiable epizootic posing a serious threat to global honey bee health because not only individual larvae but also entire colonies succumb to the disease. In the recent past considerable progress has been made in elucidating molecular aspects of host pathogen interactions during pathogenesis of P. larvae infections. Especially the sequencing and annotation of the complete genome of P. larvae was a major step forward and revealed the existence of several giant gene clusters coding for non-ribosomal peptide synthetases which might act as putative virulence factors. We here present the detailed analysis of one of these clusters which we demonstrated to be responsible for the biosynthesis of bacillibactin, a P. larvae siderophore. We first established culture conditions allowing the growth of P. larvae under iron-limited conditions and triggering siderophore production by P. larvae. Using a gene disruption strategy we linked siderophore production to the expression of an uninterrupted bacillibactin gene cluster. In silico analysis predicted the structure of a trimeric trithreonyl lactone (DHB-Gly-Thr)3 similar to the structure of bacillibactin produced by several Bacillus species. Mass spectrometric analysis unambiguously confirmed that the siderophore produced by P. larvae is identical to bacillibactin. Exposure bioassays demonstrated that P. larvae bacillibactin is not required for full virulence of P. larvae in laboratory exposure bioassays. This observation is consistent with results obtained for bacillibactin in other pathogenic bacteria. PMID:25237888

  7. TonB-dependent outer-membrane proteins and siderophore utilization in Pseudomonas fluorescens Pf-5.

    PubMed

    Hartney, Sierra L; Mazurier, Sylvie; Kidarsa, Teresa A; Quecine, Maria Carolina; Lemanceau, Philippe; Loper, Joyce E

    2011-04-01

    The soil bacterium Pseudomonas fluorescens Pf-5 produces two siderophores, a pyoverdine and enantio-pyochelin, and its proteome includes 45 TonB-dependent outer-membrane proteins, which commonly function in uptake of siderophores and other substrates from the environment. The 45 proteins share the conserved β-barrel and plug domains of TonB-dependent proteins but only 18 of them have an N-terminal signaling domain characteristic of TonB-dependent transducers (TBDTs), which participate in cell-surface signaling systems. Phylogenetic analyses of the 18 TBDTs and 27 TonB-dependent receptors (TBDRs), which lack the N-terminal signaling domain, suggest a complex evolutionary history including horizontal transfer among different microbial lineages. Putative functions were assigned to certain TBDRs and TBDTs in clades including well-characterized orthologs from other Pseudomonas spp. A mutant of Pf-5 with deletions in pyoverdine and enantio-pyochelin biosynthesis genes was constructed and characterized for iron-limited growth and utilization of a spectrum of siderophores. The mutant could utilize as iron sources a large number of pyoverdines with diverse structures as well as ferric citrate, heme, and the siderophores ferrichrome, ferrioxamine B, enterobactin, and aerobactin. The diversity and complexity of the TBDTs and TBDRs with roles in iron uptake clearly indicate the importance of iron in the fitness and survival of Pf-5 in the environment. PMID:21080032

  8. Impact of siderophore production by Pseudomonas syringae pv. syringae 22d/93 on epiphytic fitness and biocontrol activity against Pseudomonas syringae pv. glycinea 1a/96.

    PubMed

    Wensing, Annette; Braun, Sascha D; Büttner, Petra; Expert, Dominique; Völksch, Beate; Ullrich, Matthias S; Weingart, Helge

    2010-05-01

    The use of naturally occurring microbial antagonists to suppress plant diseases offers a favorable alternative to classical methods of plant protection. The soybean epiphyte Pseudomonas syringae pv. syringae strain 22d/93 shows great potential for controlling P. syringae pv. glycinea, the causal agent of bacterial blight of soybean. Its activity against P. syringae pv. glycinea is highly reproducible even in field trials, and the suppression mechanisms involved are of special interest. In this work we demonstrated that P. syringae pv. syringae 22d/93 produced a significantly larger amount of siderophores than the pathogen P. syringae pv. glycinea produced. While P. syringae pv. syringae 22d/93 and P. syringae pv. glycinea produce the same siderophores, achromobactin and pyoverdin, the regulation of siderophore biosynthesis in the former organism is very different from that in the latter organism. The epiphytic fitness of P. syringae pv. syringae 22d/93 mutants defective in siderophore biosynthesis was determined following spray inoculation of soybean leaves. The population size of the siderophore-negative mutant P. syringae pv. syringae strain 22d/93DeltaSid was 2 orders of magnitude lower than that of the wild type 10 days after inoculation. The growth deficiency was compensated for when wound inoculation was used, indicating the availability of iron in the presence of small lesions on the leaves. Our results suggest that siderophore production has an indirect effect on the biocontrol activity of P. syringae pv. syringae 22d/93. Although siderophore-defective mutants of P. syringae pv. syringae 22d/93 still suppressed development of bacterial blight caused by P. syringae pv. glycinea, siderophore production enhanced the epiphytic fitness and thus the competitiveness of the antagonist. PMID:20208028

  9. Requirement of siderophore biosynthesis for plant colonization by Salmonella enterica

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Contaminated fresh produce has become the number one vector of non-typhoidal salmonellosis to humans. However, Salmonella enterica genes essential for the life cycle of this organism outside the mammalian host are for the most part unknown. Screening deletion mutants led to the discovery that an aro...

  10. Siderophores in environmental research: roles and applications

    PubMed Central

    Ahmed, E; Holmström, S J M

    2014-01-01

    Siderophores are organic compounds with low molecular masses that are produced by microorganisms and plants growing under low iron conditions. The primary function of these compounds is to chelate the ferric iron [Fe(III)] from different terrestrial and aquatic habitats and thereby make it available for microbial and plant cells. Siderophores have received much attention in recent years because of their potential roles and applications in various areas of environmental research. Their significance in these applications is because siderophores have the ability to bind a variety of metals in addition to iron, and they have a wide range of chemical structures and specific properties. For instance, siderophores function as biocontrols, biosensors, and bioremediation and chelation agents, in addition to their important role in weathering soil minerals and enhancing plant growth. The aim of this literature review is to outline and discuss the important roles and functions of siderophores in different environmental habitats and emphasize the significant roles that these small organic molecules could play in applied environmental processes. PMID:24576157

  11. Brucella abortus strain 2308 produces brucebactin, a highly efficient catecholic siderophore.

    PubMed

    González Carreró, Manuel I; Sangari, Félix J; Agüero, Jesús; García Lobo, Juan M

    2002-02-01

    Brucella abortus is known to produce 2,3-dihydroxybenzoate (2,3-DHBA) and to use this catechol as a siderophore to grow under iron-limited conditions. In this study a mutant (BAM41) is described that is deficient in siderophore production by insertion of Tn5 in the virulent B. abortus strain 2308. This mutant was unable to grow on iron-deprived medium and its growth could not be restored by addition of 2,3-DHBA. Production of catecholic compounds by both the Brucella mutant and parental strains under iron-deprivation conditions was assayed by TLC. Two catecholic substances were identified in the supernatant of the parental strain 2308. The faster migrating spot showed the same retention factor (R(f)) as that of purified 2,3-DHBA. The mutant BAM41 overproduced 2,3-DHBA, but failed to form the slower migrating catechol. This defect could only be complemented by the addition of the slow-migrating catechol from strain 2308. The genomic region containing Tn5 in BAM41 was cloned and the position of the transposon was determined by nucleotide sequencing. The sequence revealed that the insertion had occurred at a gene with homology to Escherichia coli entF, a locus involved in the late steps of the biosynthesis of the complex catecholic siderophore enterobactin. Intracellular survival and growth rates of the B. abortus wild-type and entF mutant strains in mouse-derived J774 macrophages were similar, indicating that production of this siderophore was not essential in this model of infection. It is concluded that B. abortus synthesizes a previously unknown and highly efficient catecholic siderophore, different from 2,3-DHBA, for which the name brucebactin is proposed. PMID:11832499

  12. Plant mechanisms of siderophore-iron utilization

    SciTech Connect

    Crowley, D.E.

    1986-01-01

    Mechanisms of siderophore iron-utilization by plants were examined to determine whether plants have direct mechanisms for acquiring iron from microbially-produced hydroxamate siderophores or simply take up inorganic iron in equilibrium with the chelate (shuttle mechanism). Experiments were designed to determine whether the monocot plant species, oat (Avena sativa L. cv. Victory) could acquire iron from ferrichrome under hydroponic conditions in which iron uptake was most likely to occur by direct use of the chelating agent. Ten-day-old iron-deficient seedlings, grown in aerated Hoagland's nutrient solution (minus iron) buffered at pH 7.4 with CaCO/sub 3/, were placed in fresh nutrient solution containing 10/sup -7.4/M radioactive /sup 55/FeCl/sub 3/ (23.7 mCi/mg) with the synthetic chelate, EDDHA (10..pi../sup 5/M), ferrichrome (10/sup -5/M), or with no chelate. After 6 days, shoot content of /sup 55/Fe in shoots of plants provided with ferrichrome was 100-fold greater than that in shoots of plants provided with EDDHA. Therefore iron uptake by oat under these conditions not only indicates direct use of ferrichrome, but also suggest that oat may be better able to acquire iron from siderophores than from synthetic chelates. One possible mechanism for direct use of chelating agents, may involve siderophore binding sites on the plasmalemma of root cortical cells where iron is split from the chelate by enzymatic reduction of ferric to ferrous iron. To demonstrate hypothesized siderophore binding sites on oat roots, experiments examined possible competition for presumed siderophore binding sites by an inert analog of ferrichrome constructed by irreversible chelation with chromium.

  13. Siderophore-mediated iron acquisition influences motility and is required for full virulence of the xylem-dwelling bacterial phytopathogen Pantoea stewartii subsp. stewartii.

    PubMed

    Burbank, Lindsey; Mohammadi, Mojtaba; Roper, M Caroline

    2015-01-01

    Iron is a key micronutrient for microbial growth but is often present in low concentrations or in biologically unavailable forms. Many microorganisms overcome this challenge by producing siderophores, which are ferric-iron chelating compounds that enable the solubilization and acquisition of iron in a bioactive form. Pantoea stewartii subsp. stewartii, the causal agent of Stewart's wilt of sweet corn, produces a siderophore under iron-limiting conditions. The proteins involved in the biosynthesis and export of this siderophore are encoded by the iucABCD-iutA operon, which is homologous to the aerobactin biosynthetic gene cluster found in a number of enteric pathogens. Mutations in iucA and iutA resulted in a decrease in surface-based motility that P. stewartii utilizes during the early stages of biofilm formation, indicating that active iron acquisition impacts surface motility for P. stewartii. Furthermore, bacterial movement in planta is also dependent on a functional siderophore biosynthesis and uptake pathway. Most notably, siderophore-mediated iron acquisition is required for full virulence in the sweet corn host, indicating that active iron acquisition is essential for pathogenic fitness for this important xylem-dwelling bacterial pathogen. PMID:25326304

  14. Siderophore-Mediated Iron Acquisition Influences Motility and Is Required for Full Virulence of the Xylem-Dwelling Bacterial Phytopathogen Pantoea stewartii subsp. stewartii

    PubMed Central

    Burbank, Lindsey; Mohammadi, Mojtaba

    2014-01-01

    Iron is a key micronutrient for microbial growth but is often present in low concentrations or in biologically unavailable forms. Many microorganisms overcome this challenge by producing siderophores, which are ferric-iron chelating compounds that enable the solubilization and acquisition of iron in a bioactive form. Pantoea stewartii subsp. stewartii, the causal agent of Stewart's wilt of sweet corn, produces a siderophore under iron-limiting conditions. The proteins involved in the biosynthesis and export of this siderophore are encoded by the iucABCD-iutA operon, which is homologous to the aerobactin biosynthetic gene cluster found in a number of enteric pathogens. Mutations in iucA and iutA resulted in a decrease in surface-based motility that P. stewartii utilizes during the early stages of biofilm formation, indicating that active iron acquisition impacts surface motility for P. stewartii. Furthermore, bacterial movement in planta is also dependent on a functional siderophore biosynthesis and uptake pathway. Most notably, siderophore-mediated iron acquisition is required for full virulence in the sweet corn host, indicating that active iron acquisition is essential for pathogenic fitness for this important xylem-dwelling bacterial pathogen. PMID:25326304

  15. Siderophore-Based Iron Acquisition and Pathogen Control

    PubMed Central

    Miethke, Marcus; Marahiel, Mohamed A.

    2007-01-01

    Summary: High-affinity iron acquisition is mediated by siderophore-dependent pathways in the majority of pathogenic and nonpathogenic bacteria and fungi. Considerable progress has been made in characterizing and understanding mechanisms of siderophore synthesis, secretion, iron scavenging, and siderophore-delivered iron uptake and its release. The regulation of siderophore pathways reveals multilayer networks at the transcriptional and posttranscriptional levels. Due to the key role of many siderophores during virulence, coevolution led to sophisticated strategies of siderophore neutralization by mammals and (re)utilization by bacterial pathogens. Surprisingly, hosts also developed essential siderophore-based iron delivery and cell conversion pathways, which are of interest for diagnostic and therapeutic studies. In the last decades, natural and synthetic compounds have gained attention as potential therapeutics for iron-dependent treatment of infections and further diseases. Promising results for pathogen inhibition were obtained with various siderophore-antibiotic conjugates acting as “Trojan horse” toxins and siderophore pathway inhibitors. In this article, general aspects of siderophore-mediated iron acquisition, recent findings regarding iron-related pathogen-host interactions, and current strategies for iron-dependent pathogen control will be reviewed. Further concepts including the inhibition of novel siderophore pathway targets are discussed. PMID:17804665

  16. A cell biological view of the siderophore pyochelin iron uptake pathway in Pseudomonas aeruginosa.

    PubMed

    Cunrath, Olivier; Gasser, Véronique; Hoegy, Françoise; Reimmann, Cornelia; Guillon, Laurent; Schalk, Isabelle J

    2015-01-01

    Pyochelin (PCH) is a siderophore produced and secreted by Pseudomonas aeruginosa for iron capture. Using (55) Fe uptake and binding assays, we showed that PCH-Fe uptake in P. aeruginosa involves, in addition to the highly studied outer membrane transporter FptA, the inner membrane permease FptX, which recognizes PCH-(55) Fe with an affinity of 0.6 ± 0.2 nM and transports the ferri-siderophore complex from the periplasm into the cytoplasm: fptX deletion inhibited (55) Fe accumulation in the bacterial cytoplasm. Chromosomal replacement was used to generate P. aeruginosa strains producing fluorescent fusions with FptX, PchR (an AraC regulator), PchA (the first enzyme involved in the PCH biosynthesis) and PchE (a non-ribosomic peptide-synthetase involved in a further step). Fluorescence imaging and cellular fractionation showed a uniform repartition of FptX in the inner membrane. PchA and PchE were found in the cytoplasm, associated to the inner membrane all over the bacteria and also concentrated at the bacterial poles. PchE clustering at the bacterial poles was dependent on PchA expression, but on the opposite PchA clustering and membrane association was PchE-independent. PchA and PchE cellular organization suggests the existence of a siderosome for PCH biosynthesis as previously proposed for pyoverdine biosynthesis (another siderophore produced by P. aeruginosa). PMID:24947078

  17. Deciphering Protein Dynamics of the Siderophore Pyoverdine Pathway in Pseudomonas aeruginosa

    PubMed Central

    Guillon, Laurent; Altenburger, Stephan; Graumann, Peter L.; Schalk, Isabelle J.

    2013-01-01

    Pseudomonas aeruginosa produces the siderophore, pyoverdine (PVD), to obtain iron. Siderophore pathways involve complex mechanisms, and the machineries responsible for biosynthesis, secretion and uptake of the ferri-siderophore span both membranes of Gram-negative bacteria. Most proteins involved in the PVD pathway have been identified and characterized but the way the system functions as a whole remains unknown. By generating strains expressing fluorescent fusion proteins, we show that most of the proteins are homogeneously distributed throughout the bacterial cell. We also studied the dynamics of these proteins using fluorescence recovery after photobleaching (FRAP). This led to the first diffusion coefficients ever determined in P. aeruginosa. Cytoplasmic and periplamic diffusion appeared to be slower than in Escherichia coli but membrane proteins seemed to behave similarly in the two species. The diffusion of cytoplasmic and periplasmic tagged proteins involved in the PVD pathway was dependent on the interaction network to which they belong. Importantly, the TonB protein, motor of the PVD-Fe uptake process, was mostly immobile but its mobility increased substantially in the presence of PVD-Fe. PMID:24205369

  18. Production of siderophores increases resistance to fusaric acid in Pseudomonas protegens Pf-5.

    PubMed

    Ruiz, Jimena A; Bernar, Evangelina M; Jung, Kirsten

    2015-01-01

    Fusaric acid is produced by pathogenic fungi of the genus Fusarium, and is toxic to plants and rhizobacteria. Many fluorescent pseudomonads can prevent wilt diseases caused by these fungi. This study was undertaken to evaluate the effect of fusaric acid on P. protegens Pf-5 and elucidate the mechanisms that enable the bacterium to survive in the presence of the mycotoxin. The results confirm that fusaric acid negatively affects growth and motility of P. protegens. Moreover, a notable increase in secretion of the siderophore pyoverdine was observed when P. protegens was grown in the presence of fusaric acid. Concomitantly, levels of enzymes involved in the biosynthesis of pyoverdine and enantio-pyochelin, the second siderophore encoded by P. protegens, increased markedly. Moreover, while similar levels of resistance to fusaric acid were observed for P. protegens mutants unable to synthesize either pyoverdine or enanto-pyochelin and the wild type strain, a double mutant unable to synthesize both kinds of siderophores showed a dramatically reduced resistance to this compound. This reduced resistance was not observed when this mutant was grown under conditions of iron excess. Spectrophotometric titrations revealed that fusaric acid binds not only Fe2+ and Fe3+, but also Zn2+, Mn2+ and Cu2+, with high affinity. Our results demonstrate that iron sequestration accounts at least in part for the deleterious effect of the mycotoxin on P. protegens. PMID:25569682

  19. Production of Siderophores Increases Resistance to Fusaric Acid in Pseudomonas protegens Pf-5

    PubMed Central

    Ruiz, Jimena A.; Bernar, Evangelina M.; Jung, Kirsten

    2015-01-01

    Fusaric acid is produced by pathogenic fungi of the genus Fusarium, and is toxic to plants and rhizobacteria. Many fluorescent pseudomonads can prevent wilt diseases caused by these fungi. This study was undertaken to evaluate the effect of fusaric acid on P. protegens Pf-5 and elucidate the mechanisms that enable the bacterium to survive in the presence of the mycotoxin. The results confirm that fusaric acid negatively affects growth and motility of P. protegens. Moreover, a notable increase in secretion of the siderophore pyoverdine was observed when P. protegens was grown in the presence of fusaric acid. Concomitantly, levels of enzymes involved in the biosynthesis of pyoverdine and enantio-pyochelin, the second siderophore encoded by P. protegens, increased markedly. Moreover, while similar levels of resistance to fusaric acid were observed for P. protegens mutants unable to synthesize either pyoverdine or enanto-pyochelin and the wild type strain, a double mutant unable to synthesize both kinds of siderophores showed a dramatically reduced resistance to this compound. This reduced resistance was not observed when this mutant was grown under conditions of iron excess. Spectrophotometric titrations revealed that fusaric acid binds not only Fe2+ and Fe3+, but also Zn2+, Mn2+ and Cu2+, with high affinity. Our results demonstrate that iron sequestration accounts at least in part for the deleterious effect of the mycotoxin on P. protegens. PMID:25569682

  20. Microbial siderophores and their potential applications: a review.

    PubMed

    Saha, Maumita; Sarkar, Subhasis; Sarkar, Biplab; Sharma, Bipin Kumar; Bhattacharjee, Surajit; Tribedi, Prosun

    2016-03-01

    Siderophores are small organic molecules produced by microorganisms under iron-limiting conditions which enhance the uptake of iron to the microorganisms. In environment, the ferric form of iron is insoluble and inaccessible at physiological pH (7.35-7.40). Under this condition, microorganisms synthesize siderophores which have high affinity for ferric iron. These ferric iron-siderophore complexes are then transported to cytosol. In cytosol, the ferric iron gets reduced into ferrous iron and becomes accessible to microorganism. In recent times, siderophores have drawn much attention due to its potential roles in different fields. Siderophores have application in microbial ecology to enhance the growth of several unculturable microorganisms and can alter the microbial communities. In the field of agriculture, different types of siderophores promote the growth of several plant species and increase their yield by enhancing the Fe uptake to plants. Siderophores acts as a potential biocontrol agent against harmful phyto-pathogens and holds the ability to substitute hazardous pesticides. Heavy-metal-contaminated samples can be detoxified by applying siderophores, which explicate its role in bioremediation. Siderophores can detect the iron content in different environments, exhibiting its role as a biosensor. In the medical field, siderophore uses the "Trojan horse strategy" to form complexes with antibiotics and helps in the selective delivery of antibiotics to the antibiotic-resistant bacteria. Certain iron overload diseases for example sickle cell anemia can be treated with the help of siderophores. Other medical applications of siderophores include antimalarial activity, removal of transuranic elements from the body, and anticancer activity. The aim of this review is to discuss the important roles and applications of siderophores in different sectors including ecology, agriculture, bioremediation, biosensor, and medicine. PMID:25758420

  1. Microbial Tailoring of Acyl Peptidic Siderophores

    PubMed Central

    2015-01-01

    Marine bacteria produce an abundance of suites of acylated siderophores characterized by a unique, species-dependent headgroup that binds iron(III) and one of a series of fatty acid appendages. Marinobacter sp. DS40M6 produces a suite of seven acylated marinobactins, with fatty acids ranging from saturated and unsaturated C12–C18 fatty acids. In the present study, we report that in the late log phase of growth, the fatty acids are hydrolyzed by an amide hydrolase producing the peptidic marinobactin headgroup. Halomonas aquamarina str. DS40M3, another marine bacterium isolated originally from the same sample of open ocean water as Marinobacter sp. DS40M6, produces the acyl aquachelins, also as a suite composed of a peptidic headgroup distinct from that of the marinobactins. In contrast to the acyl marinobactins, hydrolysis of the suite of acyl aquachelins is not detected, even when H. aquamarina str. DS40M3 is grown into the stationary phase. The Marinobacter cell-free extract containing the acyl amide hydrolase is active toward exogenous acyl-peptidic siderophores (e.g., aquachelin C, loihichelin C, as well as octanoyl homoserine lactone used in quorum sensing). Further, when H. aquamarina str. DS40M3 is cultured together with Marinobacter sp. DS40M6, the fatty acids of both suites of siderophores are hydrolyzed, and the aquachelin headgroup is also produced. The present study demonstrates that coculturing bacteria leads to metabolically tailored metabolites compared to growth in a single pure culture, which is interesting given the importance of siderophore-mediated iron acquisition for bacterial growth and that Marinobacter sp. DS40M6 and H. aquamarina str. DS40M3 were isolated from the same sample of seawater. PMID:24735218

  2. Microbial tailoring of acyl peptidic siderophores.

    PubMed

    Gauglitz, Julia M; Iinishi, Akira; Ito, Yusai; Butler, Alison

    2014-04-29

    Marine bacteria produce an abundance of suites of acylated siderophores characterized by a unique, species-dependent headgroup that binds iron(III) and one of a series of fatty acid appendages. Marinobacter sp. DS40M6 produces a suite of seven acylated marinobactins, with fatty acids ranging from saturated and unsaturated C12-C18 fatty acids. In the present study, we report that in the late log phase of growth, the fatty acids are hydrolyzed by an amide hydrolase producing the peptidic marinobactin headgroup. Halomonas aquamarina str. DS40M3, another marine bacterium isolated originally from the same sample of open ocean water as Marinobacter sp. DS40M6, produces the acyl aquachelins, also as a suite composed of a peptidic headgroup distinct from that of the marinobactins. In contrast to the acyl marinobactins, hydrolysis of the suite of acyl aquachelins is not detected, even when H. aquamarina str. DS40M3 is grown into the stationary phase. The Marinobacter cell-free extract containing the acyl amide hydrolase is active toward exogenous acyl-peptidic siderophores (e.g., aquachelin C, loihichelin C, as well as octanoyl homoserine lactone used in quorum sensing). Further, when H. aquamarina str. DS40M3 is cultured together with Marinobacter sp. DS40M6, the fatty acids of both suites of siderophores are hydrolyzed, and the aquachelin headgroup is also produced. The present study demonstrates that coculturing bacteria leads to metabolically tailored metabolites compared to growth in a single pure culture, which is interesting given the importance of siderophore-mediated iron acquisition for bacterial growth and that Marinobacter sp. DS40M6 and H. aquamarina str. DS40M3 were isolated from the same sample of seawater. PMID:24735218

  3. Siderophores: The special ingredient to cyanobacterial blooms

    NASA Astrophysics Data System (ADS)

    Du, Xue; Creed, Irena; Trick, Charles

    2013-04-01

    Freshwater lakes provide a number of significant ecological services including clean drinking water, habitat for aquatic biota, and economic benefits. The provision of these ecological services, as well as the health of these aquatic systems, is threatened by the excessive growth of algae, specifically, cyanobacteria. Historically, blooms have been linked to eutrophication but recent occurrences indicate that there are less dramatic changes that induce these blooms. Iron is an essential micronutrient required for specific essential metabolic pathways; however, the amount of biologically available iron in naturally occurring lake ranges from saturation to much lower than cell transport affinities. To assist in the modulation of iron availabilities, cyanobacteria in culture produce low molecular weight compounds that function in an iron binding and acquisition system; nevertheless, this has yet to be confirmed in naturally occurring lakes. This project explored the relationship of P, N and in particular, Fe, in the promotion of cyanobacteria harmful algal blooms in 30 natural freshwater lakes located in and around the Elk Island National Park, Alberta. It is hypothesized that cyanobacteria produce and utilize iron chelators called siderophores in low Fe and nitrogen (N) conditions, creating a competitive advantage over other algae in freshwater lakes. Lakes were selected to represent a range of iron availability to explore the nutrient composition of lakes that propagated cyanobacteria harmful algal blooms (cHABs) compared to lakes that did not. Lake water was analyzed for nutrients, microbial composition, siderophore concentration, and toxin concentration. Modifications were made to optimize the Czaky and Arnow tests for hydroxamate- and catecholate-type siderophores, respectively, for field conditions. Preliminary results indicate the presence of iron-binding ligands (0.11-2.34 mg/L) in freshwater lakes characterized by widely ranging Fe regimes (0.04-2.74 mg

  4. XAFS Determination of Pb and Cd Speciation with Siderophores and the Metal/Siderophore/Kaolinite System

    SciTech Connect

    Mishra, Bhoopesh; Haack, Elizabeth A.; Vasconcelos, Igor F.; Maurice, Patricia A.; Bunker, Bruce A.

    2008-06-16

    We provide evidence for hexadentate complexes of Pb{sup 2+} and Cd{sup 2+} with the trihydroxamate siderophore desferrioxamine B (DFO-B) at pH 7.5, and 9.0, respectively. Analysis of the species of Pb{sup 2+} and Cd{sup 2+} adsorbed at the surface of kaolinite clay under the same pH conditions and in the presence of DFO-B indicate that Pb{sup 2+} is sorbed as a metal-siderophore complex while Cd{sup 2+} is not.

  5. Structure and biosynthetic assembly of cupriachelin, a photoreactive siderophore from the bioplastic producer Cupriavidus necator H16.

    PubMed

    Kreutzer, Martin F; Kage, Hirokazu; Nett, Markus

    2012-03-21

    The bacterium Cupriavidus necator H16 produces a family of linear lipopeptides when grown under low iron conditions. The structural composition of these molecules, exemplified by the main metabolite cupriachelin, is reminiscent of siderophores that are excreted by marine bacteria. Comparable to marine siderophores, the ferric form of cupriachelin exhibits photoreactive properties. Exposure to UV light induces an oxidation of its peptidic backbone and a concomitant reduction of the coordinated Fe(III). Here, we report the genomics-inspired isolation and structural characterization of cupriachelin as well as its encoding gene cluster, which was identified by insertional mutagenesis. Based upon the functional characterization of adenylation domain specificity, a model for cupriachelin biosynthesis is proposed. PMID:22381697

  6. Siderophore-promoted dissolution of smectite by fluorescent Pseudomonas.

    PubMed

    Ferret, Claire; Sterckeman, Thibault; Cornu, Jean-Yves; Gangloff, Sophie; Schalk, Isabelle J; Geoffroy, Valérie A

    2014-10-01

    Siderophores are organic chelators produced by microorganisms to fulfil their iron requirements. Siderophore-promoted dissolution of iron-bearing minerals has been clearly documented for some siderophores, but few studies have addressed metabolizing siderophore-producing bacteria. We investigated iron acquisition from clays by fluorescent Pseudomonads, bacteria that are ubiquitous in the environment. We focused on the interactions between smectite and Pseudomonas aeruginosa, a bacterium producing two structurally different siderophores: pyoverdine and pyochelin. The presence of smectite in iron-limited growth media promoted planktonic growth of P. aeruginosa and biofilm surrounding the smectite aggregates. Chemical analysis of the culture media indicated increases in the dissolved silicon, iron and aluminium concentrations following smectite supplementation. The use of P. aeruginosa mutants unable to produce either one or both of the two siderophores indicated that pyoverdine, the siderophore with the higher affinity for iron, was involved in iron and aluminium solubilization by the wild-type strain. However, in the absence of pyoverdine, pyochelin was also able to solubilize iron but with a twofold lower efficiency. In conclusion, pyoverdine and pyochelin, two structurally different siderophores, can solubilize structural iron from smectite and thereby make it available for bacterial growth. PMID:25646536

  7. Uranium extraction by complexation with siderophores

    NASA Astrophysics Data System (ADS)

    Bahamonde Castro, Cristina

    One of the major concerns of energy production is the environmental impact associated with the extraction of natural resources. Nuclear energy fuel is obtained from uranium, an abundant and naturally occurring element in the environment, but the currently used techniques for uranium extraction leave either a significant fingerprint (open pit mines) or a chemical residue that alters the pH of the environment (acid or alkali leaching). It is therefore clear that a new and greener approach to uranium extraction is needed. Bioleaching is one potential alternative. In bioleaching, complexants naturally produced from fungi or bacteria may be used to extract the uranium. In the following research, the siderophore enterobactin, which is naturally produced by bacteria to extract and solubilize iron from the environment, is evaluated to determine its potential for complexing with uranium. To determine whether enterobactin could be used for uranium extraction, its acid dissociation and its binding strength with the metal of interest must be determined. Due to the complexity of working with radioactive materials, lanthanides were used as analogs for uranium. In addition, polyprotic acids were used as structural and chemical analogs for the siderophore during method development. To evaluate the acid dissociation of enterobactin and the subsequent binding constants with lanthanides, three different analytical techniques were studied including: potentiometric titration, UltraViolet Visible (UV-Vis) spectrophotometry and Isothermal Titration Calorimetry (ITC). After evaluation of three techniques, a combination of ITC and potentiometric titrations was deemed to be the most viable way for studying the siderophore of interest. The results obtained from these studies corroborate the ideal pH range for enterobactin complexation to the lanthanide of interest and pave the way for determining the strength of complexation relative to other naturally occurring metals. Ultimately, this

  8. Chemistry and biology of pyoverdines, Pseudomonas primary siderophores.

    PubMed

    Cézard, C; Farvacques, N; Sonnet, P

    2015-01-01

    as signal molecules for the production of acute virulence factors and are involved in biofilm formation as well. The ongoing expanding pathogenicity of P. aeruginosa has become a major public health problem, and finding alternative strategies to classical antibiotics is urgently needed. Pyoverdines along with the iron pathway recently gained interest among academical researchers as potential new approaches to "fight" the bacteria. This review describes the classification of the nearly 60 pyoverdines identified so far, their structural and chemical properties and their (bio)synthesis. The different mechanisms underlying the steps of a pyoverdine's life in Pseudomonas are detailed as well: the affinity by which a pyoverdine chelates iron(III), the description of the interactions inducing the siderophore-receptor recognition, the specific transport of the pyoverdine-Fe(III) complex. As pyoverdine production and severe infections are linked, we will also report on situations where pyoverdines are considered as being P. aeruginosa Achilles heel: the propensity of FpvA to transport exo-pyoverdines, organic synthesis of pyoverdines and analogs, grafting of antibiotics on pyoverdines in a Trojan Horse strategy. PMID:25312210

  9. Microbial production of four biodegradable siderophores under submerged fermentation.

    PubMed

    Fazary, Ahmed E; Al-Shihri, Ayed S; Alfaifi, Mohammad Y; Saleh, Kamel A; Alshehri, Mohammed A; Elbehairi, Serag Eldin I; Ju, Yi-Hsu

    2016-07-01

    Four siderophore analogues were isolated and purified from Escherichia coli, Bacillus spp. ST13, and Streptomyces pilosus microorganisms under some specific submerged fermentation conditions. In order to evaluate the highest production of this siderophore analogues through the growth, a rapid spectrophotometric screening semi-quantitative method was used, in which interestingly the analogues were isolated in its own form not its iron chelate. After chromatographic separation, the chemical structures of the isolated and purified siderophores were illustrated using detailed spectroscopic techniques. The biodegradation studies were done on that four novel isolated and purified siderophores following OECD protocols. In addition, the bioactivities of these siderophores and their iron complexes were examined and evaluated. PMID:27091230

  10. Beyond iron: non-classical biological functions of bacterial siderophores.

    PubMed

    Johnstone, Timothy C; Nolan, Elizabeth M

    2015-04-14

    Bacteria secrete small molecules known as siderophores to acquire iron from their surroundings. For over 60 years, investigations into the bioinorganic chemistry of these molecules, including fundamental coordination chemistry studies, have provided insight into the crucial role that siderophores play in bacterial iron homeostasis. The importance of understanding the fundamental chemistry underlying bacterial life has been highlighted evermore in recent years because of the emergence of antibiotic-resistant bacteria and the need to prevent the global rise of these superbugs. Increasing reports of siderophores functioning in capacities other than iron transport have appeared recently, but reports of "non-classical" siderophore functions have long paralleled those of iron transport. One particular non-classical function of these iron chelators, namely antibiotic activity, was documented before the role of siderophores in iron transport was established. In this Perspective, we present an exposition of past and current work into non-classical functions of siderophores and highlight the directions in which we anticipate that this research is headed. Examples include the ability of siderophores to function as zincophores, chalkophores, and metallophores for a variety of other metals, sequester heavy metal toxins, transport boron, act as signalling molecules, regulate oxidative stress, and provide antibacterial activity. PMID:25764171

  11. Beyond Iron: Non-Classical Biological Functions of Bacterial Siderophores

    PubMed Central

    Johnstone, Timothy C.; Nolan, Elizabeth M.

    2015-01-01

    Bacteria secrete small molecules known as siderophores to acquire iron from their surroundings. For over 60 years, investigations into the bioinorganic chemistry of these molecules, including fundamental coordination chemistry studies, have provided insight into the crucial role that siderophores play in bacterial iron homeostasis. The importance of understanding the fundamental chemistry underlying bacterial life has been highlighted evermore in recent years because of the emergence of antibiotic-resistant bacteria and the need to prevent the global rise of these superbugs. Increasing reports of siderophores functioning in capacities other than iron transport have appeared recently, but reports of “non-classical” siderophore functions have long paralleled those of iron transport. One particular non-classical function of these iron chelators, namely antibiotic activity, was even documented before the role of siderophores in iron transport was established. In this Perspective, we present an exposition of past and current work into non-classical functions of siderophores and highlight the directions in which we anticipate that this research is headed. Examples include the ability of siderophores to function as zincophores, chalkophores, and metallophores for a variety of other metals, sequester heavy metal toxins, transport boron, act as signalling molecules, regulate oxidative stress, and provide antibacterial activity. PMID:25764171

  12. Six Siderophore-Producing Microorganisms Identified in Biological Soil Crusts

    NASA Astrophysics Data System (ADS)

    Noonan, K.; Anbar, A. D.; Garcia-Pichel, F.; Poret-peterson, A. T.; Hartnett, H. E.

    2011-12-01

    Biological soil crusts (BSCs) are diverse microbial communities that colonize soils in arid and semi-arid environments. Cyanobacteria in BSCs are pioneer organisms that increase ecosystem habitability by providing fixed carbon (C) and nitrogen (N) as well as by reducing water run-off and increasing infiltration. Photosynthesis and N fixation, in particular, require a variety of metals in large quantities, and yet, metals are predominantly insoluble in the environments where BSCs thrive. Therefore, BSC organisms must have efficient strategies for extracting metals from soil minerals. We hypothesized that BSC microbes, particularly the cyanobacteria, produce siderophores to serve their metal-acquisition needs. Siderophores are small organic compounds that bind Fe with high affinity and are produced by a variety of microorganisms, including cyanobacteria. Most siderophores bind Fe, primarily; however, some can also bind Mo, V, and Cu. Soil siderophores are released by microbes to increase the solubility of metals from minerals and to facilitate microbial uptake. Thus, siderophores serve as chemical weathering agents and provide a direct link between soil microbes and minerals. Studying siderophore production in BSCs provides insight into how BSCs tackle the challenge of acquiring insoluble metals, and may help conservationists determine useful fertilizers for BSC growth by facilitating metal acquisition. Biological soil crusts were collected near Moab, UT. Soil slurries were prepared in deionized water and transferred to modified BG-11 agar plates. The O-CAS agar plate assay was used to screen organisms for siderophore production. Siderophore producing microbes were isolated and identified by16S rRNA gene sequencing. Cultures were then grown in 3 L batch cultures under metal limitation, and siderophore presence was monitored using the traditional liquid CAS assay. After siderophore detection, cells were removed by centrifugation, organic compounds were separated using

  13. Siderophores from Neighboring Organisms Promote the Growth of Uncultured Bacteria

    PubMed Central

    D’Onofrio, Anthony; Crawford, Jason M.; Stewart, Eric J.; Witt, Kathrin; Gavrish, Ekaterina; Epstein, Slava; Clardy, Jon; Lewis, Kim

    2010-01-01

    Summary The majority of bacterial species do not grow on synthetic media. Many non-growers require growth factors from other bacteria, but the nature of these compounds is largely unknown. We show here that previously uncultured isolates from marine sediment biofilm grow on a Petri dish in the presence of cultured organisms from the same environment. The growth factors produced by one cultured helper strain were identified as new acyl-desferrioxamine siderophores. A panel of previously uncultured isolates exhibited a range of siderophore promiscuity for growth promotion. This siderophore-based approach has enabled the culturing of organisms only distantly related to previously cultured microbes. The lack of growth in the lab for many strains from this habitat stems from an inability to autonomously produce siderophores, and the resulting chemical dependence on other microorganisms regulates community establishment in the environment. PMID:20338517

  14. Identification of the hydroxamate siderophore ferricrocin in Cladosporium cladosporioides.

    PubMed

    Pourhassan, Nina; Gagnon, René; Wichard, Thomas; Bellenger, Jean-Philippe

    2014-04-01

    The hydroxamate siderophore ferricrocin was identified in Cladosporium cladosporioides growth medium by solid phase extraction and ultra high pressure liquid chromatography coupled to a time of flight mass spectrometer (UHPLC/QTOF-MS). Both desferricrocin and ferricrocin were detected in the extracellular medium assisted by high resolution mass spectrometry. This is the first identification of a hydroxamate siderophore in Cladosporium cladosporioides. This finding emphasizes the common meaning of ferricrocin in fungi. PMID:24868878

  15. Regulation of mammalian siderophore 2,5-DHBA in the innate immune response to infection

    PubMed Central

    Liu, Zhuoming; Reba, Scott; Chen, Wei-Dong; Porwal, Suheel Kumar; Boom, W. Henry; Petersen, Robert B.; Rojas, Roxana; Viswanathan, Rajesh

    2014-01-01

    Competition for iron influences host–pathogen interactions. Pathogens secrete small iron-binding moieties, siderophores, to acquire host iron. In response, the host secretes siderophore-binding proteins, such as lipocalin 24p3, which limit siderophore-mediated iron import into bacteria. Mammals produce 2,5-dihydroxy benzoic acid, a compound that resembles a bacterial siderophore. Our data suggest that bacteria use both mammalian and bacterial siderophores. In support of this idea, supplementation with mammalian siderophore enhances bacterial growth in vitro. In addition, mice lacking the mammalian siderophore resist E. coli infection. Finally, we show that the host responds to infection by suppressing siderophore synthesis while up-regulating lipocalin 24p3 expression via TLR signaling. Thus, reciprocal regulation of 24p3 and mammalian siderophore is a protective mechanism limiting microbial access to iron. PMID:24863067

  16. In vitro-binding of the natural siderophore enantiomers pyochelin and enantiopyochelin to their AraC-type regulators PchR in Pseudomonas.

    PubMed

    Lin, Po-Chi; Youard, Zeb A; Reimmann, Cornelia

    2013-12-01

    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. PMID:24037597

  17. Siderophore production in high iron environments

    NASA Astrophysics Data System (ADS)

    Bennett, S. A.; Hoffman, C. L.; Moffett, J. W.; Edwards, K. J.

    2010-12-01

    Up until recently, the geochemical cycling of Fe in deep sea hydrothermal plumes has assumed to be inorganically dominated, resulting in quantitative precipitation of all hydrothermally sourced Fe to the seafloor. Recent detection of organic Fe binding ligands within both the dissolved and particulate phase (Bennett et al., 2008; Toner et al., 2009), suggests that hydrothermally sourced Fe may be important on a global scale (Tagliabue et al., 2010). The source of these organic ligands is currently unknown; hypotheses include the possible entrainment of organic carbon from the biologically rich diffuse flow areas, or in-situ production from microbial processes. However, the microbial production of organic ligands is only expected when Fe is a limited micronutrient, which is not the case in the hydrothermal environment. The importance of Fe cycling microorganisms within hydrothermal systems was previously overlooked due to the poor energetics with regards to Fe oxidation and reduction. But their recent detection within the hydrothermal system, both around low temperature Fe rich mineral deposits and within hydrothermal plumes (Edwards et al., 2004; Sylvan et al., In prep) suggests that they may have an important role in the hydrothermal Fe cycle, potentially resulting in an interplay between Fe and organic carbon. Within the laboratory, we have carried out experiments to investigate an Fe oxidizing bacteria in a variety of high Fe environments. We have detected both the production of siderophores and an increase in reduced Fe when the Fe oxidizing bacteria is exposed to both Fe(III) and Fe(II) rich minerals. The role of these microbes in the mineral dissolution of Fe sulfides along the seafloor and within the hydrothermal plume, may have important implications on the speciation of Fe and the role of siderophores in the marine environment. Bennett, S.A. et al. 2008. EPSL, 270: 157-167. Edwards, K.J. et al. 2004. Geomicrobiology Journal, 21: 393-404. Sylvan, J.B. et al

  18. Non-nucleoside Inhibitors of BasE, An Adenylating Enzyme in the Siderophore Biosynthetic Pathway of the Opportunistic Pathogen Acinetobacter baumannii

    PubMed Central

    Neres, João; Engelhart, Curtis A.; Drake, Eric J.; Wilson, Daniel J.; Fu, Peng; Boshoff, Helena I.; Barry, Clifton E.; Gulick, Andrew M.; Aldrich, Courtney C.

    2013-01-01

    Siderophores are small-molecule iron chelators produced by bacteria and other microorganisms for survival under iron limiting conditions, such as found in a mammalian host. Siderophore biosynthesis is essential for the virulence of many important Gram-negative pathogens including Acinetobacter baumannii, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Escherichia coli. We performed high-throughput screening of against BasE, which is involved in siderophore biosynthesis in A. baumannii and identified 6-phenyl-1-(pyridin-4-ylmethyl)-1H-pyrazolo[3,4-b]pyridine-4-carboxylic acid 15. Herein we report the synthesis, biochemical, and microbiological evaluation of a systematic series of analogues of the HTS hit 15. Analogue 67 is the most potent analogue with a KD of 2 nM against BasE. Structural characterization of the inhibitors with BasE reveal they bind in a unique orientation in the active site occupying all three substrate binding sites, and thus can be considered multisubstrate inhibitors. These results provide a foundation for future studies aimed at both increasing enzyme potency and antibacterial activity. PMID:23437866

  19. Intracellular siderophore but not extracellular siderophore is required for full virulence in Metarhizium robertsii.

    PubMed

    Giuliano Garisto Donzelli, Bruno; Gibson, Donna M; Krasnoff, Stuart B

    2015-09-01

    Efficient iron acquisition mechanisms are fundamental for microbial survival in the environment and for pathogen virulence within their hosts. M. robertsii produces two known iron-binding natural products: metachelins, which are used to scavenge extracellular iron, and ferricrocin, which is strictly intracellular. To study the contribution of siderophore-mediated iron uptake and storage to M. robertsii fitness, we generated null mutants for each siderophore synthase gene (mrsidD and mrsidC, respectively), as well as for the iron uptake transcriptional repressor mrsreA. All of these mutants showed impaired germination speed, differential sensitivity to hydrogen peroxide, and differential ability to overcome iron chelation on growth-limiting iron concentrations. RT-qPCR data supported regulation of mrsreA, mrsidC, and mrsidD by supplied iron in vitro and during growth within the insect host, Spodoptera exigua. We also observed strong upregulation of the insect iron-binding proteins, transferrins, during infection. Insect bioassays revealed that ferricrocin is required for full virulence against S. exigua; neither the loss of metachelin production nor the deletion of the transcription factor mrsreA significantly affected M. robertsii virulence. PMID:26135511

  20. Pyoverdine, the Major Siderophore in Pseudomonas aeruginosa, Evades NGAL Recognition

    PubMed Central

    Peek, Mary E.; Bhatnagar, Abhinav; McCarty, Nael A.; Zughaier, Susu M.

    2012-01-01

    Pseudomonas aeruginosa is the most common pathogen that persists in the cystic fibrosis lungs. Bacteria such as P. aeruginosa secrete siderophores (iron-chelating molecules) and the host limits bacterial growth by producing neutrophil-gelatinase-associated lipocalin (NGAL) that specifically scavenges bacterial siderophores, therefore preventing bacteria from establishing infection. P. aeruginosa produces a major siderophore known as pyoverdine, found to be important for bacterial virulence and biofilm development. We report that pyoverdine did not bind to NGAL, as measured by tryptophan fluorescence quenching, while enterobactin bound to NGAL effectively causing a strong response. The experimental data indicate that pyoverdine evades NGAL recognition. We then employed a molecular modeling approach to simulate the binding of pyoverdine to human NGAL using NGAL's published crystal structures. The docking of pyoverdine to NGAL predicted nine different docking positions; however, neither apo- nor ferric forms of pyoverdine docked into the ligand-binding site in the calyx of NGAL where siderophores are known to bind. The molecular modeling results offer structural support that pyoverdine does not bind to NGAL, confirming the results obtained in the tryptophan quenching assay. The data suggest that pyoverdine is a stealth siderophore that evades NGAL recognition allowing P. aeruginosa to establish chronic infections in CF lungs. PMID:22973307

  1. Pyoverdine, the Major Siderophore in Pseudomonas aeruginosa, Evades NGAL Recognition.

    PubMed

    Peek, Mary E; Bhatnagar, Abhinav; McCarty, Nael A; Zughaier, Susu M

    2012-01-01

    Pseudomonas aeruginosa is the most common pathogen that persists in the cystic fibrosis lungs. Bacteria such as P. aeruginosa secrete siderophores (iron-chelating molecules) and the host limits bacterial growth by producing neutrophil-gelatinase-associated lipocalin (NGAL) that specifically scavenges bacterial siderophores, therefore preventing bacteria from establishing infection. P. aeruginosa produces a major siderophore known as pyoverdine, found to be important for bacterial virulence and biofilm development. We report that pyoverdine did not bind to NGAL, as measured by tryptophan fluorescence quenching, while enterobactin bound to NGAL effectively causing a strong response. The experimental data indicate that pyoverdine evades NGAL recognition. We then employed a molecular modeling approach to simulate the binding of pyoverdine to human NGAL using NGAL's published crystal structures. The docking of pyoverdine to NGAL predicted nine different docking positions; however, neither apo- nor ferric forms of pyoverdine docked into the ligand-binding site in the calyx of NGAL where siderophores are known to bind. The molecular modeling results offer structural support that pyoverdine does not bind to NGAL, confirming the results obtained in the tryptophan quenching assay. The data suggest that pyoverdine is a stealth siderophore that evades NGAL recognition allowing P. aeruginosa to establish chronic infections in CF lungs. PMID:22973307

  2. Siderophore production by pathogenic mucorales and uptake of deferoxamine B.

    PubMed

    Larcher, Gérald; Dias, Marylène; Razafimandimby, Bienvenue; Bomal, Danielle; Bouchara, Jean-Philippe

    2013-12-01

    Clinical reports have established that mucormycosis, mainly caused by Rhizopus spp., frequently occurs in patients treated with deferoxamine B (DFO, Desferal(®)) which is misappropriated by these fungi. Siderophore production by twenty mucoralean isolates was therefore investigated using a commercial iron-depleted culture medium. Siderophore production was detected for most of the isolates. Our experiments confirmed that feroxamine B (iron chelate of DFO) promoted in vitro growth of Rhizopus arrhizus. Electrophoretic analysis of somatic extracts revealed iron-regulated proteins of 60 and 32 kDa which were lacking in iron-depleted culture conditions. Using a fluorescent derivative of deferoxamine B, we showed by fluorescence microscopy the entry of the siderophore within the fungal cells, thus suggesting a shuttle mechanism encompassing the uptake of the entire siderophore-ion complex into the cell. This useful tool renders possible a better understanding of iron metabolism in Mucorales which could lead to the development of new diagnostic method or new antifungal therapy using siderophores as imaging contrast agents or active drug vectors. PMID:23982284

  3. Siderophore production and utilization by milk spoilage Pseudomonas species.

    PubMed

    Brown, A G; Luke, R K J

    2010-04-01

    Many bacteria respond to potentially growth-limiting availability of iron by producing low-molecular-weight iron chelators (siderophores). The aim of this work was to examine the siderophores synthesized and utilized by Pseudomonas spp. implicated in milk spoilage. Twenty isolates of Pseudomonas spp. previously shown to have significant milk spoilage potential were tested for the ability to produce siderophores. Of these, 14 produced pyoverdin and 2 of these also produced pyochelin; 1 produced only pyochelin; 1 produced only salicylate; 2 produced non-pyoverdin, hydroxamate-containing siderophore; and 2 produced chrome azurol sulfonate reactive material that was neither pyoverdin nor pyochelin. There was considerable diversity among the pyoverdins produced. All isolates were shown to utilize iron complexed with exogenous pyoverdin, but usage of particular exogenous pyoverdins differed among isolates. Interference with the iron-uptake systems of the Pseudomonas spp. may be a means by which food spoilage can be slowed, and the pyoverdin system would appear to be a potential target. However, given the diversity of pyoverdins produced and utilized, and the presence of other siderophores, successful interference with bacterial iron acquisition in this context may be challenging. PMID:20338412

  4. The role of siderophores in the transport of radionuclides

    SciTech Connect

    Hersman, L.E.; Palmer, P.D.; Hobart, D.E.

    1993-12-31

    Iron exists in aerobic soil and water environments most commonly as insoluble Fe(III). Siderophores are powerful, microbially produced chelating agents that are used to mobilize the insoluble Fe(III) cation. Over 80 siderophores have been isolated and characterized, with some reportedly having iron-binding constants as high as 10{sup 52}. Fe(III) and Pu(IV) are similar in their charge/ionic radius ratio (4.6 and 4.2, respectively); therefore, Pu(IV) may serve as analog to Fe(III). It is possible that some radioactive wastes could be chelated by naturally occurring siderophores, thereby altering the transport rates of those elements through the subsurface environment. This investigation was initiated to investigate that possibility. The binding of {sup 239}(IV) by four chelating agents is reported in this paper: a siderophore isolated and purified from a Pseudomonas sp.; desferal, a ferrioxamine siderophore commonly used for deferration therapy; EDTA, ethylenediaminetetraacetic acid; and, citrate, trisodium salt.

  5. Self-Assembling Amphiphilic Siderophores from Marine Bacteria

    NASA Astrophysics Data System (ADS)

    Martinez, J. S.; Zhang, G. P.; Holt, P. D.; Jung, H.-T.; Carrano, C. J.; Haygood, M. G.; Butler, Alison

    2000-02-01

    Most aerobic bacteria secrete siderophores to facilitate iron acquisition. Two families of siderophores were isolated from strains belonging to two different genera of marine bacteria. The aquachelins, from Halomonas aquamarina strain DS40M3, and the marinobactins, from Marinobacter sp. strains DS40M6 and DS40M8, each contain a unique peptidic head group that coordinates iron(III) and an appendage of one of a series of fatty acid moieties. These siderophores have low critical micelle concentrations (CMCs). In the absence of iron, the marinobactins are present as micelles at concentrations exceeding their CMC; upon addition of iron(III), the micelles undergo a spontaneous phase change to form vesicles. These observations suggest that unique iron acquisition mechanisms may have evolved in marine bacteria.

  6. Genetic diversity of siderophore-producing bacteria of tobacco rhizosphere

    PubMed Central

    Tian, Fang; Ding, Yanqin; Zhu, Hui; Yao, Liangtong; Du, Binghai

    2009-01-01

    The genetic diversity of siderophore-producing bacteria of tobacco rhizosphere was studied by amplified ribosomal DNA restriction analysis (ARDRA), 16S rRNA sequence homology and phylogenetics analysis methods. Studies demonstrated that 85% of the total 354 isolates produced siderophores in iron limited liquid medium. A total of 28 ARDRA patterns were identified among the 299 siderophore-producing bacterial isolates. The 28 ARDRA patterns represented bacteria of 14 different genera belonging to six bacterial divisions, namely β-, γ-, α-Proteobacteria, Sphingobacteria, Bacilli, and Actinobacteria. Especially, γ-Proteobacteria consisting of Pseudomonas, Enterobacter, Serratia, Pantoea, Erwinia and Stenotrophomonas genus encountered 18 different ARDRA groups. Results also showed a greater siderophore-producing bacterial diversity than previous researches. For example, Sphingobacterium (isolates G-2-21-1 and G-2-27-2), Pseudomonas poae (isolate G-2-1-1), Enterobacter endosymbiont (isolates G-2-10-2 and N-5-10), Delftia acidovorans (isolate G-1-15), and Achromobacter xylosoxidans (isolates N-46-11HH and N-5-20) were reported to be able to produce siderophores under low-iron conditions for the first time. Gram-negative isolates were more frequently encountered, with more than 95% total frequency. For Gram-positive bacteria, the Bacillus and Rhodococcus were the only two genera, with 1.7% total frequency. Furthermore, the Pseudomonas and Enterobacter were dominant in this environment, with 44.5% and 24.7% total frequency, respectively. It was also found that 75 percent of the isolates that had the high percentages of siderophore units (% between 40 and 60) belonged to Pseudomonas. Pseudomonas sp. G-229-21 screened out in this study may have potential to apply to low-iron soil to prevent plant soil-borne fungal pathogen diseases. PMID:24031358

  7. Characterization of iron uptake from hydroxamate siderophores by Chlorella vulgaris

    SciTech Connect

    Allnutt, F.C.T.

    1985-01-01

    Iron uptake by Chlorella vulgaris from ferric-hydroxamate siderophores and the possible production of siderophores by these algae was investigated. No production of siderophores or organic acids was observed. Iron from the two hydroxamate siderophores tested, ferrioximine B (Fe/sup 3 +/-DFOB) and ferric-rhodotorulate (Fe/sup 3 +/-RA), was taken up at the same rate as iron chelated by citrate or caffeate. Two synthetic chelates, Fe/sup 3 +/-EDTA and Fe/sup 3 +/-EDDHA, provided iron at a slower rate. Iron uptake was inhibited by 50 ..mu..M CCCP or 1 mM vanadate. Cyanide (100 ..mu..M KCN) or 25 ..mu..M antimycin A failed to demonstrate a link between uptake and respiration. Labeled iron (/sup 55/Fe) was taken up while labeled ligands ((/sup 14/C) citrate or RA) were not accumulated. Cation competition from Ni/sup 2 +/ and Co/sup 2 +/ observed using Fe/sup 3 +/-DFOB and Fe/sup 3 +/-RA while iron uptake from Fe/sup 3 +/-citrate was stimulated. Iron-stress induced iron uptake from the hydroxamate siderophores. Ferric reduction from the ferric-siderophores was investigated with electron paramagnetic resonance (EPR) and bathophenathroline disulfonate (BPDS). Ferric reduction was induced by iron-stress and inhibited by CCCP. A close correlation between iron uptake and ferric reduction was measured by the EPR method. Ferric reduction measured by the BPDS method was greater than that measure by EPR. BPDS reduction was interpreted to indicate a potential for reduction while EPR measures the physiological rate of reduction. BPDS inhibition of iron uptake and ferricyanide interference with reduction indicate that reduction and uptake occur exposed to the external medium. Presumptive evidence using a binding dose response curve for Fe/sup 3 +/-DFOB indicated that a receptor may be involved in this mechanism.

  8. Role of bacterial siderophores in dissolution of hornblende

    NASA Astrophysics Data System (ADS)

    Liermann, Laura J.; Kalinowski, Birgitta E.; Brantley, Susan L.; Ferry, James G.

    2000-02-01

    Hornblende, a common mineral in granitic soils, may act as a source for a variety of metals needed by bacterial species for enzyme function (e.g., Fe, Zn, Mn, Cu, Co, Mo, V, Ni). A species of the bacterial genus Streptomyces was cultured from an Adirondack soil and isolated because of its ability to grow robustly in low Fe medium with hornblende present. Studies with unbuffered culture medium, to discover whether Streptomyces sp. cultures affected solution pH, showed a decrease of 2.0 pH units in 21 d, then an increase of 3.0 pH units at 56 d. Cells that adhered to the hornblende surface at 56 days were difficult to remove, presumably because of mycelial growth deep into pits and cracks. Decreases and increases in pH may have been due to production of organic acids and ammonia respectively. Increases in pH could also have been related to release of components during death of organisms. In a buffered medium, Streptomyces sp. increased the initial Fe release rate from hornblende approximately fivefold over that of an abiotic control. A catechol derivative, produced by the Streptomyces sp. and characterized by chromatography and mass spectrometry, is presumed to cause this Fe release enhancement. Hornblende dissolution was also analyzed in the presence of a commercially available hydroxamate siderophore, desferrioxamine mesylate (DFAM). DFAM is the methane sulfonate form of one of many siderophores known to be a product of streptomycetes. The rate of Fe release obtained when incubating the hornblende with 24 μm of DFAM was similar to the rate observed in the presence of the Streptomyces sp. isolate. Higher concentrations of DFAM increased the dissolution rate nonlinearly, described by the rate equation R = (7.6 × 10 -13)C 0.47, where R is the release rate of Fe (mol/m 2s), and C is the concentration (mol/l) of DFAM. The DFAM also increased release of Al and Si from hornblende into solution; however, these release rates were not increased by addition of the

  9. Disruption of transporters affiliated with enantio-pyochelin biosynthesis gene cluster of Pseudomonas protegens Pf-5 has pleiotropic effects

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Pseudomonas protegens Pf-5 (formerly Pseudomonas fluorescens) is a biocontrol bacterium that produces the siderophore enantio-pyochelin under conditions of iron starvation in a process that is often accompanied by the secretion of its biosynthesis intermediates, salicylic acid and dihydroaeruginoic ...

  10. Role of Siderophores in Dissimilatory Iron Reduction in Arctic Soils : Effect of Direct Amendment of Siderophores to Arctic Soil

    NASA Astrophysics Data System (ADS)

    Srinivas, A. J.; Dinsdale, E. A.; Lipson, D.

    2014-12-01

    Dissimilatory iron reduction (DIR), where ferric iron (Fe3+) is reduced to ferrous iron (Fe2+) anaerobically, is an important respiratory pathway used by soil bacteria. DIR contributes to carbon dioxide (CO2) efflux from the wet sedge tundra biome in the Arctic Coastal Plain (ACP) in Alaska, and could competitively inhibit the production of methane, a stronger greenhouse gas than CO2, from arctic soils. The occurrence of DIR as a dominant anaerobic process depends on the availability of substantial levels of Fe3+ in soils. Siderophores are metabolites made by microbes to dissolve Fe3+ from soil minerals in iron deficient systems, making Fe3+ soluble for micronutrient uptake. However, as the ACP is not iron deficient, siderophores in arctic soils may play a vital role in anaerobic respiration by dissolving Fe3+ for DIR. We studied the effects of direct siderophore addition to arctic soils through a field study conducted in Barrow, Alaska, and a laboratory incubation study conducted at San Diego State University. In the field experiment, 50μM deferroxamine mesylate (a siderophore), 50μM trisodium nitrilotriacetate (an organic chelator) or an equal volume of water was added to isolated experimental plots, replicated in clusters across the landscape. Fe2+ concentrations were measured in soil pore water samples collected periodically to measure DIR over time in each. In the laboratory experiment, frozen soil samples obtained from drained thaw lake basins in the ACP, were cut into cores and treated with the above-mentioned compounds to the same final concentrations. Along with measuring Fe2+ concentrations, CO2 output was also measured to monitor DIR over time in each core. Experimental addition of siderophores to soils in both the field and laboratory resulted in increased concentrations of soluble Fe3+ and a sustained increase in Fe2+concentrations over time, along with increased respiration rates in siderophore-amended cores. These results show increased DIR in

  11. Molecular Dynamics and Electron Density Studies of Siderophores and Peptides.

    NASA Astrophysics Data System (ADS)

    Fidelis, Krzysztof Andrzej

    1990-08-01

    The dissertation comprises three separate studies of siderophores and peptides. In the first of these studies the relative potential energies for a series of diastereomers of a siderophore neocoprogen I are evaluated with molecular mechanics force field methods. Charges on the hydroxamate moiety are determined with a synthetic model siderophore compound using valence population refinements, and alternatively, with the theoretical ab initio/ESP calculations. The single diastereomer found in the crystal structure is among four characterized by the low potential energy, while prevalence of Delta vs. Lambda configuration about the iron is found to be a property of the entire series. In the second study the crystal structure of a ferrichrome siderophore ferrirhodin is reported. The crystal structure conformation of the molecular backbone as well as the iron coordination geometry compare well with other ferrichrome structures. The differences between the acyl groups of ferrirubin and ferrirhodin are explored using the methods of molecular mechanics. The third study a 300 ps, 300 K, in vacuo molecular dynamics simulation of didemnin A and B yields distinct molecular conformers, which are different from the one found in the crystal structure or modeled in solution, using the Nuclear Overhauser Effect data. Evaluations of the relative potential energy are performed with short 10 ps simulations in solution. Didemnins are natural depsipeptides isolated from a Caribbean tunicate and characterized by particularly potent antiproliferative and immunomodulatory activity. Conformationally rigid and flexible regions of the molecule are described. A short review of the molecular mechanics methodology is given in the introduction.

  12. Synthesis of fluorescent probes based on the pyochelin siderophore scaffold.

    PubMed

    Noël, Sabrina; Guillon, Laurent; Schalk, Isabelle J; Mislin, Gaëtan L A

    2011-03-01

    Pyochelin is a siderophore common to several pathogenic bacterial strains. Two conjugates, 1 and 2, between the NBD (4-nitro-benzo[1,2,5]oxadiazole) fluorophore and an N3''-functionalized pyochelin were synthesized. These fluorescent probes unexpectedly increased their fluorescence in an aqueous medium in the presence of iron(III) and were transported into bacterial cells. PMID:21294578

  13. Rational design of a transition state analogue with picomolar affinity for Pseudomonas aeruginosa PvdQ, a siderophore biosynthetic enzyme.

    PubMed

    Clevenger, Kenneth D; Wu, Rui; Er, Joyce A V; Liu, Dali; Fast, Walter

    2013-10-18

    The Pseudomonas aeruginosa enzyme PvdQ can process different substrates involved in quorum-sensing or in siderophore biosynthesis. Substrate selectivity was evaluated using steady-state kinetic constants for hydrolysis of N-acyl-homoserine lactones (HSLs) and p-nitrophenyl fatty acid esters. PvdQ prefers substrates with alkyl chains between 12 and 14 carbons long that do not bear a 3-oxo substitution and is revealed here to have a relatively high specificity constant for selected N-acyl-HSLs (kcat/KM = 10(5) to 10(6) M(-1) s(-1)). However, endogenous P. aeruginosa N-acyl-HSLs are ≥100-fold disfavored, supporting the conclusion that PvdQ was not primarily evolved to regulate endogenous quorum-sensing. PvdQ plays an essential biosynthetic role for the siderophore pyoverdine, on which P. aeruginosa depends for growth in iron-limited environments. A series of alkylboronate inhibitors was found to be reversible, competitive, and extremely potent (Ki ≥ 190 pM). A 1.8 Å X-ray structure shows that 1-tridecylboronic acid forms a monocovalent bond with the N-terminal β-chain Ser residue in the PvdQ heterodimer, mimicking a reaction transition state. This boronic acid inhibits growth of P. aeruginosa in iron-limited media, reproducing the phenotype of a genetic pvdQ disruption, although co-administration of an efflux pump inhibitor is required to maintain growth inhibition. These findings support the strategy of designing boron-based inhibitors of siderophore biosynthetic enzymes to control P. aeruginosa infections. PMID:23883096

  14. Siderophore-promoted dissolution of cobalt from hydroxide minerals

    NASA Astrophysics Data System (ADS)

    Bi, Yuqiang; Hesterberg, Dean L.; Duckworth, Owen W.

    2010-05-01

    Recent research has revealed that siderophores, a class of biogenic ligands with high affinities for Fe(III), can also strongly complex Co(III), an element essential to the normal metabolic function of microbes and animals. This study was conducted to quantify the rates and identify the products and mechanisms of the siderophore-promoted dissolution of Co from synthetic Co-bearing minerals. The dissolution reactions of heterogenite (CoOOH) and four Co-substituted goethites (Co-FeOOH) containing different Co concentrations were investigated in the presence of a trihydroxamate siderophore, desferrioxamine B (DFOB), using batch and flow-through experiments. Results showed that DFOB-promoted dissolution of Co from Co-bearing minerals may occur via pH-dependent ligand-promoted or reductive dissolution mechanisms. For heterogenite, ligand-promoted dissolution was the dominant pathway at neutral to alkaline pH, while production of dissolved Co(II) for pH <6. It was not possible from our data to decouple the separate contributions of homogenous and heterogeneous reduction reactions to the aqueous Co(II) pool. Cobalt substitution in Co-substituted goethite, possibly caused by distortion of goethite structure and increased lattice strain, resulted in enhanced total dissolution rates of both Co and Fe. The DFOB-promoted dissolution rates of Co-bearing minerals, coupled with the high affinity of Co(III) for DFOB, suggest that siderophores may be effective for increasing Co solubility, and thus possibly Co bioavailability. The results also suggest that siderophores may contribute to the mobilization of radioactive 60Co from Co-bearing mineral phases through mineral weathering and dissolution processes.

  15. MmpL transporter-mediated export of cell-wall associated lipids and siderophores in mycobacteria.

    PubMed

    Chalut, Christian

    2016-09-01

    Mycobacteria produce a large variety of surface-exposed lipids with unusual structures. Some of these compounds are ubiquitously present in mycobacteria and play an important role in the structural organization of the cell envelope, while others are species-specific. The biosynthesis of most of these lipids requires modular polyketide synthases (PKS) or non-ribosomal peptide synthetases (NRPS) that are intracellular, suggesting that the assembly of these compounds takes place in the cytosolic compartment or near the inner leaflet of the plasma membrane. The molecular mechanisms that mediate the export of these lipid components across the cell envelope remain poorly understood. Mycobacterial membrane protein Large (MmpL) transporters, a subclass of Resistance-Nodulation-Cell Division (RND) transporters, appear to play a major role in this process, acting as scaffold proteins that couple lipid synthesis and transport. Recent studies have shown that this family of transporters also contributes to siderophore secretion in Mycobacterium tuberculosis. The goal of this review is to provide the most recent advances in our understanding of the molecular mechanisms involved in lipid and siderophore transport mediated by MmpL transporters. PMID:27553408

  16. Identification of inhibitors of PvdQ, an enzyme involved in the synthesis of the siderophore pyoverdine.

    PubMed

    Wurst, Jacqueline M; Drake, Eric J; Theriault, Jimmy R; Jewett, Ivan T; VerPlank, Lynn; Perez, Jose R; Dandapani, Sivaraman; Palmer, Michelle; Moskowitz, Samuel M; Schreiber, Stuart L; Munoz, Benito; Gulick, Andrew M

    2014-07-18

    Pseudomonas aeruginosa produces the peptide siderophore pyoverdine, which is used to acquire essential Fe(3+) ions from the environment. PvdQ, an Ntn hydrolase, is required for the biosynthesis of pyoverdine. PvdQ knockout strains are not infectious in model systems, suggesting that disruption of siderophore production via PvdQ inhibition could be exploited as a target for novel antibacterial agents, by preventing cells from acquiring iron in the low iron environments of most biological settings. We have previously described a high-throughput screen to identify inhibitors of PvdQ that identified inhibitors with IC50 values of ∼100 μM. Here, we describe the discovery of ML318, a biaryl nitrile inhibitor of PvdQ acylase. ML318 inhibits PvdQ in vitro (IC50 = 20 nM) by binding in the acyl-binding site, as confirmed by the X-ray crystal structure of PvdQ bound to ML318. Additionally, the PvdQ inhibitor is active in a whole cell assay, preventing pyoverdine production and limiting the growth of P. aeruginosa under iron-limiting conditions. PMID:24824984

  17. Identification of Specific In Vivo-Induced (ivi) Genes in Yersinia ruckeri and Analysis of Ruckerbactin, a Catecholate Siderophore Iron Acquisition System

    PubMed Central

    Fernández, L.; Márquez, I.; Guijarro, J. A.

    2004-01-01

    This work reports the utilization of an in vivo expression technology system to identify in vivo-induced (ivi) genes in Yersinia ruckeri after determination of the conditions needed for its selection in fish. Fourteen clones were selected, and the cloned DNA fragments were analyzed after partial sequencing. In addition to sequences with no significant similarity, homology with genes encoding proteins putatively involved in two-component and type IV secretion systems, adherence, specific metabolic functions, and others were found. Among these sequences, four were involved in iron acquisition through a catechol siderophore (ruckerbactin). Thus, unlike other pathogenic yersiniae producing yersiniabactin, Y. ruckeri might be able to produce and utilize only this phenolate. The genetic organization of the ruckerbactin biosynthetic and uptake loci was similar to that of the Escherichia coli enterobactin gene cluster. Genes rucC and rupG, putative counterparts of E. coli entC and fepG, respectively, involved in the biosynthesis and transport of the iron siderophore complex, respectively, were analyzed further. Thus, regulation of expression by iron and temperature and their presence in other Y. ruckeri siderophore-producing strains were confirmed for these two loci. Moreover, 50% lethal dose values 100-fold higher than those of the wild-type strain were obtained with the rucC isogenic mutant, showing the importance of ruckerbactin in the pathogenesis caused by this microorganism. PMID:15345400

  18. NPS6, Encoding a Nonribosomal Peptide Synthetase Involved in Siderophore-Mediated Iron Metabolism, Is a Conserved Virulence Determinant of Plant Pathogenic Ascomycetes[W

    PubMed Central

    Oide, Shinichi; Moeder, Wolfgang; Krasnoff, Stuart; Gibson, Donna; Haas, Hubertus; Yoshioka, Keiko; Turgeon, B. Gillian

    2006-01-01

    NPS6, encoding a nonribosomal peptide synthetase, is a virulence determinant in the maize (Zea mays) pathogen Cochliobolus heterostrophus and is involved in tolerance to H2O2. Deletion of NPS6 orthologs in the rice (Oryza sativa) pathogen, Cochliobolus miyabeanus, the wheat (Triticum aestivum) pathogen, Fusarium graminearum, and the Arabidopsis thaliana pathogen, Alternaria brassicicola, resulted in reduced virulence and hypersensitivity to H2O2. Introduction of the NPS6 ortholog from the saprobe Neurospora crassa to the Δnps6 strain of C. heterostrophus restored wild-type virulence to maize and tolerance to H2O2, demonstrating functional conservation in filamentous ascomycete phytopathogens and saprobes. Increased sensitivity to iron depletion was identified as a conserved phenotype of Δnps6 strains. Exogenous application of iron enhanced the virulence of Δnps6 strains of C. heterostrophus, C. miyabeanus, F. graminearum, and A. brassicicola to each host. NPS6 is responsible for the biosynthesis of extracellular siderophores by C. heterostrophus, F. graminearum, and A. brassicicola. Application of the extracellular siderophore of A. brassicicola restored wild-type virulence of the ΔAbnps6 strain to Arabidopsis. It is proposed that the role of extracellular siderophores in fungal virulence to plants is to supply an essential nutrient, iron, to their producers in planta and not to act as phytotoxins, depriving their hosts of iron. PMID:17056706

  19. Auxin biosynthesis.

    PubMed

    Zhao, Yunde

    2014-01-01

    lndole-3-acetic acid (IAA), the most important natural auxin in plants, is mainly synthesized from the amino acid tryptophan (Trp). Recent genetic and biochemical studies in Arabidopsis have unambiguously established the first complete Trp-dependent auxin biosynthesis pathway. The first chemical step of auxin biosynthesis is the removal of the amino group from Trp by the TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS (TAA) family of transaminases to generate indole-3-pyruvate (IPA). IPA then undergoes oxidative decarboxylation catalyzed by the YUCCA (YUC) family of flavin monooxygenases to produce IAA. This two-step auxin biosynthesis pathway is highly conserved throughout the plant kingdom and is essential for almost all of the major developmental processes. The successful elucidation of a complete auxin biosynthesis pathway provides the necessary tools for effectively modulating auxin concentrations in plants with temporal and spatial precision. The progress in auxin biosynthesis also lays a foundation for understanding polar auxin transport and for dissecting auxin signaling mechanisms during plant development. PMID:24955076

  20. Auxin Biosynthesis

    PubMed Central

    Zhao, Yunde

    2014-01-01

    lndole-3-acetic acid (IAA), the most important natural auxin in plants, is mainly synthesized from the amino acid tryptophan (Trp). Recent genetic and biochemical studies in Arabidopsis have unambiguously established the first complete Trp-dependent auxin biosynthesis pathway. The first chemical step of auxin biosynthesis is the removal of the amino group from Trp by the TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS (TAA) family of transaminases to generate indole-3-pyruvate (IPA). IPA then undergoes oxidative decarboxylation catalyzed by the YUCCA (YUC) family of flavin monooxygenases to produce IAA. This two-step auxin biosynthesis pathway is highly conserved throughout the plant kingdom and is essential for almost all of the major developmental processes. The successful elucidation of a complete auxin biosynthesis pathway provides the necessary tools for effectively modulating auxin concentrations in plants with temporal and spatial precision. The progress in auxin biosynthesis also lays a foundation for understanding polar auxin transport and for dissecting auxin signaling mechanisms during plant development. PMID:24955076

  1. Analysis of achromobactin biosynthesis by Pseudomonas syringae pv. syringae B728a.

    PubMed

    Berti, Andrew D; Thomas, Michael G

    2009-07-01

    Pseudomonas syringae pv. syringae B728a is known to produce the siderophore pyoverdine under iron-limited conditions. It has also been proposed that this pathovar has the ability to produce a second siderophore, achromobactin. Here we present genetic and biochemical evidence supporting the hypothesis that P. syringae pv. syringae B728a produces both of these siderophores. We show that strains unable to synthesize either pyoverdine or achromobactin are unable to grow under iron-limiting conditions, which is consistent with these two molecules being the only siderophores synthesized by P. syringae pv. syringae B728a. Enzymes associated with achromobactin biosynthesis were purified and analyzed for substrate recognition. We showed that AcsD, AcsA, and AcsC together are able to condense citrate, ethanolamine, 2,4-diaminobutyrate, and alpha-ketoglutarate into achromobactin. Replacement of ethanolamine with ethylene diamine or 1,3-diaminopropane in these reactions resulted in the formation of achromobactin analogs that were biologically active. This work provides insights into the biosynthetic steps in the formation of achromobactin and is the first in vitro reconstitution of achromobactin biosynthesis. PMID:19482931

  2. Oxytetracycline Biosynthesis*

    PubMed Central

    Pickens, Lauren B.; Tang, Yi

    2010-01-01

    Oxytetracycline (OTC) is a broad-spectrum antibiotic that acts by inhibiting protein synthesis in bacteria. It is an important member of the bacterial aromatic polyketide family, which is a structurally diverse class of natural products. OTC is synthesized by a type II polyketide synthase that generates the poly-β-ketone backbone through successive decarboxylative condensation of malonyl-CoA extender units, followed by modifications by cyclases, oxygenases, transferases, and additional tailoring enzymes. Genetic and biochemical studies have illuminated most of the steps involved in the biosynthesis of OTC, which is detailed here as a representative case study in type II polyketide biosynthesis. PMID:20522541

  3. Amphiphilic siderophore production by oil-associating microbes.

    PubMed

    Kem, Michelle P; Zane, Hannah K; Springer, Stephen D; Gauglitz, Julia M; Butler, Alison

    2014-06-01

    The Deepwater Horizon oil spill in 2010 released an unprecedented amount of oil into the ocean waters of the Gulf of Mexico. As a consequence, bioremediation by oil-degrading microbes has been a topic of increased focus. One factor limiting the rate of hydrocarbon degradation by microbial communities is the availability of necessary nutrients, including iron. The siderophores produced from two Vibrio spp. isolated from the Gulf of Mexico following the Deepwater Horizon oil spill, along with the well-studied oil-degrading microbe, Alcanivorax borkumensis SK2, are studied under iron-limiting conditions. Here we report the amphiphilic amphibactin siderophores produced by the oil-associated bacteria, Vibrio sp. S1B, Vibrio sp. S2A and Alcanivorax borkumensis SK2. These findings provide insight into oil-associating microbial iron acquisition. PMID:24663669

  4. Azospirillum brasilense siderophores with antifungal activity against Colletotrichum acutatum.

    PubMed

    Tortora, María L; Díaz-Ricci, Juan C; Pedraza, Raúl O

    2011-04-01

    Anthracnose, caused by the fungus Colletotrichum acutatum is one of the most important diseases in strawberry crop. Due to environmental pollution and resistance produced by chemical fungicides, nowadays biological control is considered a good alternative for crop protection. Among biocontrol agents, there are plant growth-promoting bacteria, such as members of the genus Azospirillum. In this work, we demonstrate that under iron limiting conditions different strains of A. brasilense produce siderophores, exhibiting different yields and rates of production according to their origin. Chemical assays revealed that strains REC2 and REC3 secrete catechol type siderophores, including salicylic acid, detected by thin layer chromatography coupled with fluorescence spectroscopy and gas chromatography-mass spectrometry analysis. Siderophores produced by them showed in vitro antifungal activity against C. acutatum M11. Furthermore, this latter coincided with results obtained from phytopathological tests performed in planta, where a reduction of anthracnose symptoms on strawberry plants previously inoculated with A. brasilense was observed. These outcomes suggest that some strains of A. brasilense could act as biocontrol agent preventing anthracnose disease in strawberry. PMID:21234749

  5. Germination-defective mutant of Neurospora crassa that responds to siderophores

    NASA Technical Reports Server (NTRS)

    Charlang, G.; Williams, N. P.

    1977-01-01

    A conditionally germination-defective mutant of Neurospora crassa has been found to be partially curable by ferricrocin and other siderophores. The mutant conidia rapidly lose their membrane-bound siderophores when suspended in buffer or growth media. Germination is consequently delayed unless large numbers of conidia are present (positive population effect). This indicates that the mutant has a membrane defect involving the siderophore attachment site.

  6. Consecutive enzymatic modification of ornithine generates the hydroxamate moieties of the siderophore erythrochelin.

    PubMed

    Robbel, Lars; Helmetag, Verena; Knappe, Thomas A; Marahiel, Mohamed A

    2011-07-12

    Biosynthesis of the hydroxamate-type siderophore erythrochelin requires the generation of δ-N-acetyl-δ-N-hydroxy-L-ornithine (L-haOrn), which is incorporated into the tetrapeptide at positions 1 and 4. Bioinformatic analysis revealed the FAD-dependent monooxygenase EtcB and the bifunctional malonyl-CoA decarboxylase/acetyltransferase Mcd to be putatively involved in the generation of L-haOrn. To investigate if EtcB and Mcd constitute a two-enzyme pathway for the biosynthesis of L-haOrn, they were produced in a recombinant manner and subjected to biochemical studies in vitro. Hydroxylation assays employing recombinant EtcB gave rise to δ-N-hydroxy-L-ornithine (L-hOrn) and confirmed the enzyme to be involved in building block assembly. Acetylation assays were carried out by incubating L-hOrn with recombinant Mcd and malonyl-CoA as the acetyl group donor. Substrate turnover was increased by substituting malonyl-CoA with acetyl-CoA, bypassing the decarboxylation reaction which represents the rate-limiting step. Consecutive enzymatic synthesis of L-haOrn was accomplished in coupled assays employing both the L-ornithine hydroxylase and Mcd. In summary, a biosynthetic route for the generation of δ-N-acetyl-δ-N-hydroxy-L-ornithine starting from L-ornithine has been established in vitro by tandem action of the FAD-dependent monooxygenase EtcB and the bifunctional malonyl-CoA decarboxylase/acetyltransferase Mcd. PMID:21650455

  7. Microbial Copper-binding Siderophores at the Host-Pathogen Interface*

    PubMed Central

    Koh, Eun-Ik; Henderson, Jeffrey P.

    2015-01-01

    Numerous pathogenic microorganisms secrete small molecule chelators called siderophores defined by their ability to bind extracellular ferric iron, making it bioavailable to microbes. Recently, a siderophore produced by uropathogenic Escherichia coli, yersiniabactin, was found to also bind copper ions during human infections. The ability of yersiniabactin to protect E. coli from copper toxicity and redox-based phagocyte defenses distinguishes it from other E. coli siderophores. Here we compare yersiniabactin to other extracellular copper-binding molecules and review how copper-binding siderophores may confer virulence-associated gains of function during infection pathogenesis. PMID:26055720

  8. Proteobactin and a yersiniabactin-related siderophore mediate iron acquisition in Proteus mirabilis

    PubMed Central

    Himpsl, Stephanie D.; Pearson, Melanie M.; Arewång, Carl J.; Nusca, Tyler D.; Sherman, David H.; Mobley, Harry L. T.

    2010-01-01

    Proteus mirabilis causes complicated urinary tract infections (UTI). While the urinary tract is an iron-limiting environment, iron acquisition remains poorly characterized for this uropathogen. Microarray analysis of P. mirabilis HI4320 cultured under iron limitation identified 45 significantly up-regulated genes (P ≤ 0.05) that represent 21 putative iron-regulated systems. Two gene clusters, PMI0229-0239 and PMI2596–2605, encode putative siderophore systems. PMI0229-0239 encodes a nonribosomal peptide synthetase (NRPS)-independent siderophore (NIS) system for producing a novel siderophore, proteobactin. PMI2596-2605 are contained within the high-pathogenicity island, originally described in Yersinia pestis, and encodes proteins with apparent homology and organization to those involved in yersiniabactin production and uptake. Cross-feeding and biochemical analysis shows that P. mirabilis is unable to utilize or produce yersiniabactin, suggesting that this yersiniabactin-related locus is functionally distinct. Only disruption of both systems resulted in an in vitro iron-chelating defect; demonstrating production and iron-chelating activity for both siderophores. These findings clearly show that proteobactin and the yersiniabactin-related siderophore function as iron acquisition systems. Despite the activity of both siderophores, only mutants lacking the yersiniabactin-related siderophore reduce fitness in vivo. The fitness requirement for the yersiniabactin-related siderophore during UTI shows, for the first time, the importance of siderophore production in vivo for P. mirabilis. PMID:20923418

  9. Burkholderia cepacia XXVI siderophore with biocontrol capacity against Colletotrichum gloeosporioides.

    PubMed

    de Los Santos-Villalobos, Sergio; Barrera-Galicia, Guadalupe Coyolxauhqui; Miranda-Salcedo, Mario Alberto; Peña-Cabriales, Juan José

    2012-08-01

    Colletotrichum gloeosporioides is the causal agent of anthracnose in mango. Burkholderia cepacia XXVI, isolated from mango rhizosphere and identified by 16S rDNA sequencing as a member of B. cepacia complex, was more effective than 6 other mango rhizosphere bacteria in inhibiting the model mango pathogen, C. gloeosporioides ATCC MYA 456. Biocontrol of this pathogen was demonstrated on Petri-dishes containing PDA by > 90 % reduction of surface colonization. The nature of the biocontrol metabolite(s) was characterized via a variety of tests. The inhibition was almost exclusively due to production of agar-diffusible, not volatile, metabolite(s). The diffusible metabolite(s) underwent thermal degradation at 70 and 121 °C (1 atm). Tests for indole acetic acid production and lytic enzyme activities (cellulase, glucanase and chitinase) by B. cepacia XXVI were negative, indicating that these metabolites were not involved in the biocontrol effect. Based on halo formation and growth inhibition of the pathogen on the diagnostic medium, CAS-agar, as well as colorimetric tests we surmised that strain XXVI produced a hydroxamate siderophore involved in the biocontrol effect observed. The minimal inhibitory concentration test showed that 0.64 μg ml(-1) of siderophore (Deferoxamine mesylate salt-equivalent) was sufficient to achieve 91.1 % inhibition of the pathogen growth on Petri-dishes containing PDA. The biocontrol capacity against C. gloeosporioides ATCC MYA 456 correlated directly with the siderophore production by B. cepacia XXVI: the highest concentration of siderophore production in PDB on day 7, 1.7 μg ml(-1) (Deferoxamine mesylate salt-equivalent), promoted a pathogen growth inhibition of 94.9 %. The growth of 5 additional strains of C. gloeosporioides (isolated from mango "Ataulfo" orchards located in the municipality of Chahuites, State of Oaxaca in Mexico) was also inhibited when confronted with B. cepacia XXVI. Results indicate that B. cepacia XXVI or its

  10. Synthesis and Pharmacokinetic Evaluation of Siderophore Biosynthesis Inhibitors for Mycobacterium tuberculosis

    PubMed Central

    Nelson, Kathryn M.; Viswanathan, Kishore; Dawadi, Surendra; Duckworth, Benjamin P.; Boshoff, Helena I.; Barry, Clifton E.; Aldrich, Courtney C.

    2015-01-01

    MbtA catalyzes the first committed biosynthetic step of the mycobactins, which are important virulence factors associated with iron acquisition in Mycobacterium tuberculosis. MbtA is a validated therapeutic target for antitubercular drug development. 5′-O-[N-(salicyl)sulfamoyl]adenosine (1) is a bisubstrate inhibitor of MbtA and exhibits exceptionally potent biochemical and antitubercular activity. However, 1 suffers from sub-optimal drug disposition properties resulting in a short half-life (t1/2), low exposure (AUC), and low bioavailability (F). Four strategies were pursued to address these liabilities including the synthesis of prodrugs, increasing the pKa of the acyl-sulfonyl moiety, modulation of the lipophilicity, and strategic introduction of fluorine into 1. Complete pharmacokinetic (PK) analysis of all compounds was performed. The most successful modifications involved fluorination of the nucleoside that provided substantial improvements in t1/2 and AUC. Increasing the pKa of the acyl-sulfonyl linker yielded incremental enhancements while modulation of the lipophilicity and prodrug approaches led to substantially poorer PK parameters. PMID:26110337

  11. Synthesis and pharmacological evaluation of nucleoside prodrugs designed to target siderophore biosynthesis in Mycobacterium tuberculosis.

    PubMed

    Dawadi, Surendra; Kawamura, Shuhei; Rubenstein, Anja; Remmel, Rory; Aldrich, Courtney C

    2016-03-15

    The nucleoside antibiotic, 5'-O-[N-(salicyl)sulfamoyl]adenosine (1), possesses potent whole-cell activity against Mycobacterium tuberculosis (Mtb), the etiological agent of tuberculosis (TB). This compound is also active in vivo, but suffers from poor drug disposition properties that result in poor bioavailability and rapid clearance. The synthesis and evaluation of a systematic series of lipophilic ester prodrugs containing linear and α-branched alkanoyl groups from two to twelve carbons at the 3'-position of a 2'-fluorinated analog of 1 is reported with the goal to improve oral bioavailability. The prodrugs were stable in simulated gastric fluid (pH 1.2) and under physiological conditions (pH 7.4). The prodrugs were also remarkably stable in mouse, rat, and human serum (relative serum stability: human∼rat≫mouse) displaying a parabolic trend in the SAR with hydrolysis rates increasing with chain length up to eight carbons (t1/2=1.6 h for octanoyl prodrug 7 in mouse serum) and then decreasing again with higher chain lengths. The permeability of the prodrugs was also assessed in a Caco-2 cell transwell model. All of the prodrugs were found to have reduced permeation in the apical-to-basolateral direction and enhanced permeation in the basolateral-to-apical direction relative to the parent compound 2, resulting in efflux ratios 5-28 times greater than 2. Additionally, Caco-2 cells were found to hydrolyze the prodrugs with SAR mirroring the serum stability results and a preference for hydrolysis on the apical side. Taken together, these results suggest that the described prodrug strategy will lead to lower than expected oral bioavailability of 2 and highlight the contribution of intestinal esterases for prodrug hydrolysis. PMID:26875934

  12. Multiple modes of iron uptake by the filamentous, siderophore-producing cyanobacterium, Anabaena sp. PCC 7120.

    PubMed

    Rudolf, Mareike; Kranzler, Chana; Lis, Hagar; Margulis, Ketty; Stevanovic, Mara; Keren, Nir; Schleiff, Enrico

    2015-08-01

    Iron is a member of a small group of nutrients that limits aquatic primary production. Mechanisms for utilizing iron have to be efficient and adapted according to the ecological niche. In respect to iron acquisition cyanobacteria, prokaryotic oxygen evolving photosynthetic organisms can be divided into siderophore- and non-siderophore-producing strains. The results presented in this paper suggest that the situation is far more complex. To understand the bioavailability of different iron substrates and the advantages of various uptake strategies, we examined iron uptake mechanisms in the siderophore-producing cyanobacterium Anabaena sp. PCC 7120. Comparison of the uptake of iron complexed with exogenous (desferrioxamine B, DFB) or to self-secreted (schizokinen) siderophores by Anabaena sp. revealed that uptake of the endogenous produced siderophore complexed to iron is more efficient. In addition, Anabaena sp. is able to take up dissolved, ferric iron hydroxide species (Fe') via a reductive mechanism. Thus, Anabaena sp. exhibits both, siderophore- and non-siderophore-mediated iron uptake. While assimilation of Fe' and FeDFB are not induced by iron starvation, FeSchizokinen uptake rates increase with increasing iron starvation. Consequently, we suggest that Fe' reduction and uptake is advantageous for low-density cultures, while at higher densities siderophore uptake is preferred. PMID:25943160

  13. An overview of siderophores for iron acquisition in microorganisms living in the extreme.

    PubMed

    De Serrano, Luis O; Camper, Anne K; Richards, Abigail M

    2016-08-01

    Siderophores are iron-chelating molecules produced by microbes when intracellular iron concentrations are low. Low iron triggers a cascade of gene activation, allowing the cell to survive due to the synthesis of important proteins involved in siderophore synthesis and transport. Generally, siderophores are classified by their functional groups as catecholates, hydroxamates and hydroxycarboxylates. Although other chemical structural modifications and functional groups can be found. The functional groups participate in the iron-chelating process when the ferri-siderophore complex is formed. Classified as acidophiles, alkaliphiles, halophiles, thermophiles, psychrophiles, piezophiles, extremophiles have particular iron requirements depending on the environmental conditions in where they grow. Most of the work done in siderophore production by extremophiles is based in siderophore concentration and/or genomic studies determining the presence of siderophore synthesis and transport genes. Siderophores produced by extremophiles are not well known and more work needs to be done to elucidate chemical structures and their role in microorganism survival and metal cycling in extreme environments. PMID:27457587

  14. TonB-Dependent outer-membrane proteins and siderophore utilization in Pseudomonas fluorescens Pf-5

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The soil bacterium Pseudomonas fluorescens Pf-5 produces two siderophores, a pyoverdine and enantio-pyochelin, and its proteome includes 45 TonB-dependent outer-membrane proteins, which commonly function in uptake of siderophores and other substrates from the environment. The 45 proteins share the ...

  15. Metachelins, mannosylated and N-oxidized coprogen-type siderophores from Metarhizium robertsii

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Under iron-depleted culture conditions, the entomopathogenic fungus Metarhizium robertsii (Bischoff, Humber, and Rehner) (= M. anisopliae) produces a complex of extracellular siderophores including novel O-glycosylated and/or N-oxidized coprogen-type compounds as well as the known fungal siderophore...

  16. The Influence of Siderophores Produced by Alkaliphilic Microorganisms on Iron and Metal Contaminant Speciation and Solubility

    NASA Astrophysics Data System (ADS)

    Aiken, A. M.; Peyton, B. M.; Petersen, J. N.; Apel, W. A.; Camper, A. K.

    2003-12-01

    Halomonas campisalis strain 4A has been identified as capable of producing siderophores under halo-alkaliphilic growth conditions. Because of the scarcity of iron under the alkaline conditions in which Halomonas campisalis thrives, we hypothesize that the siderophores secreted by Halomonas campisalis and other alkaliphilic bacteria will have a stronger affinity for binding and solubilizing ferrous iron than siderophores produced by mesophilic bacteria. Siderophore production by Halomonas campisalis was confirmed through the use of the chrome azural S (CAS) agar plate method which showed a red orange halo around the bacterial colonies indicative of siderophore production. The siderophores were found to be produced under conditions of both high salinity and pH with a salt concentrations ranging from 0.4 - 1.8 M NaCl and pH ranging from 8 - 11. The siderophores produced have been determined to be of the hydroxamate class via the Csaky method. A negative response to the Arnow assay indicated that the siderophore produced does not contain any catechol moieties in its chemical structure. It was found that maximum siderophore production was equivalent to approximately 400 mM desferrioxamine and occurred during mid stationary phase. Similar results were found at pH 8, 10 and 11. A purification scheme was developed that involved an initial extraction of the siderophore from the growth medium into benzyl alcohol followed by precipitation with diethyl ether. Additional purification was achieved via ion exchange chromatography and size exclusion chromatography. Final purification was achieved via HPLC. The structure of the purified siderophore was analyzed via LC/MS/MS equipped with an ESI source. To date, few studies have included the siderophores produced by microorganisms capable of tolerating highly saline and alkaline environments. In addition to unique structure and high affinity for iron, it is further hypothesized that siderophores from alkaliphilic bacteria will also

  17. Digital image quantification of siderophores on agar plates

    PubMed Central

    Andrews, Megan Y.; Santelli, Cara M.; Duckworth, Owen W.

    2016-01-01

    This article presents visual image data and detailed methodology for the use of a new method for quantifying the exudation of siderophores during fungal growth. The data include images showing time series for calibration, fungal exudation, and negative controls, as well as replication accuracy information. In addition, we provide detailed protocols for making CAS assay layer plates, the digital analysis protocol for determining area of color change, and discuss growth media that do and do not work with the layer plate method. The results of these data, their interpretation, and further discussion can be found in Andrews et al., 2016 [1]. PMID:26937467

  18. Digital image quantification of siderophores on agar plates.

    PubMed

    Andrews, Megan Y; Santelli, Cara M; Duckworth, Owen W

    2016-03-01

    This article presents visual image data and detailed methodology for the use of a new method for quantifying the exudation of siderophores during fungal growth. The data include images showing time series for calibration, fungal exudation, and negative controls, as well as replication accuracy information. In addition, we provide detailed protocols for making CAS assay layer plates, the digital analysis protocol for determining area of color change, and discuss growth media that do and do not work with the layer plate method. The results of these data, their interpretation, and further discussion can be found in Andrews et al., 2016 [1]. PMID:26937467

  19. Acinetobactin Isomerization Enables Adaptive Iron Acquisition in Acinetobacter baumannii through pH-Triggered Siderophore Swapping.

    PubMed

    Shapiro, Justin A; Wencewicz, Timothy A

    2016-02-12

    Pathogenic strains of Acinetobacter baumannii excrete multiple siderophores that enhance iron scavenging from host sources. The oxazoline siderophore pre-acinetobactin undergoes an unusual non-enzymatic isomerization, producing the isoxazolidinone acinetobactin. In this study, we explored the kinetics, mechanism, and biological consequence of this siderophore swapping. Pre-acinetobactin is excreted to the extracellular space where the isomerization to acinetobactin occurs with a pH-rate profile consistent with 5-exo-tet cyclization at C5' with clean stereochemical inversion. Pre-acinetobactin persists at pH <6, and acinetobactin is rapidly formed at pH >7, matching each siderophore's pH preference for iron(III) chelation and A. baumannii growth promotion. Acinetobactin isomerization provides two siderophores for the price of one, enabling A. baumannii to sequester iron over a broad pH range likely to be encountered during the course of an infection. PMID:27624967

  20. Growth of Actinobacillus pleuropneumoniae is promoted by exogenous hydroxamate and catechol siderophores.

    PubMed Central

    Diarra, M S; Dolence, J A; Dolence, E K; Darwish, I; Miller, M J; Malouin, F; Jacques, M

    1996-01-01

    Siderophores bind ferric ions and are involved in receptor-specific iron transport into bacteria. Six types of siderophores were tested against strains representing the 12 different serotypes of Actinobacillus pleuropneumoniae. Ferrichrome and bis-catechol-based siderophores showed strong growth-promoting activities for A. pleuropneumoniae in a disk diffusion assay. Most strains of A. pleuropneumoniae tested were able to use ferrichrome (21 of 22 or 95%), ferrichrome A (20 of 22 or 90%), and lysine-based bis-catechol (20 of 22 or 90%), while growth of 36% (8 of 22) was promoted by a synthetic hydroxamate, N5-acetyl-N5-hydroxy-L-ornithine tripeptide. A. pleuropneumoniae serotype 1 (strain FMV 87-682) and serotype 5 (strain 2245) exhibited a distinct yellow halo around colonies on Chrome Azurol S agar plates, suggesting that both strains can produce an iron chelator (siderophore) in response to iron stress. The siderophore was found to be neither a phenolate nor a hydroxamate by the chemical tests of Arnow and Csaky, respectively. This is the first report demonstrating the production of an iron chelator and the use of exogenous siderophores by A. pleuropneumoniae. A spermidine-based bis-catechol siderophore conjugated to a carbacephalosporin was shown to inhibit growth of A. pleuropneumoniae. A siderophore-antibiotic-resistant strain was isolated and shown to have lost the ability to use ferrichrome, synthetic hydroxamate, or catechol-based siderophores when grown under conditions of iron restriction. This observation indicated that a common iron uptake pathway, or a common intermediate, for hydroxamate- and catechol-based siderophores may exist in A. pleuropneumoniae. PMID:8975614

  1. Catechol siderophores repress the pyochelin pathway and activate the enterobactin pathway in Pseudomonas aeruginosa: an opportunity for siderophore-antibiotic conjugates development.

    PubMed

    Gasser, Véronique; Baco, Etienne; Cunrath, Olivier; August, Pamela Saint; Perraud, Quentin; Zill, Nicolas; Schleberger, Christian; Schmidt, Alexander; Paulen, Aurélie; Bumann, Dirk; Mislin, Gaëtan L A; Schalk, Isabelle J

    2016-03-01

    Previous studies have suggested that antibiotic vectorization by siderophores (iron chelators produced by bacteria) considerably increases the efficacy of such drugs. The siderophore serves as a vector: when the pathogen tries to take up iron via the siderophore, it also takes up the antibiotic. Catecholates are among the most common iron-chelating compounds used in synthetic siderophore-antibiotic conjugates. Using reverse transcription polymerase chain reaction and proteomic approaches, we showed that the presence of catecholate compounds in the medium of Pseudomonas aeruginosa led to strong activation of the transcription and expression of the outer membrane transporter PfeA, the ferri-enterobactin importer. Iron-55 uptake assays on bacteria with and without PfeA expression confirmed that catechol compounds imported iron into P. aeruginosa cells via PfeA. Uptake rates were between 0.3 × 10(3) and 2 × 10(3) Fe atoms/bacterium/min according to the used catechol siderophore in iron-restricted medium, and remained as high as 0.8 × 10(3) Fe atoms/bacterium/min for enterobactin, even in iron-rich medium. Reverse transcription polymerase chain reaction and proteomic approaches showed that in parallel to this switching on of PfeA expression, a repression of the expression of pyochelin (PCH) pathway genes (PCH being one of the two siderophores produced by P. aeruginosa for iron acquisition) was observed. PMID:26718479

  2. The Vibrio parahaemolyticus Small RNA RyhB Promotes Production of the Siderophore Vibrioferrin by Stabilizing the Polycistronic mRNA

    PubMed Central

    Funahashi, Tatsuya; Nakao, Hiroshi; Maki, Jun; Yamamoto, Shigeo

    2013-01-01

    High-affinity iron acquisition in Vibrio parahaemolyticus is mediated by the cognate siderophore vibrioferrin. We have previously reported that the vibrioferrin biosynthesis operon (pvsOp) is regulated at the transcriptional level by the iron-responsive repressor Fur (T. Tanabe, T. Funahashi, H. Nakao, S. Miyoshi, S. Shinoda, and S. Yamamoto, J. Bacteriol. 185:6938–6949, 2003). In this study, we identified the Fur-regulated small RNA RyhB and the RNA chaperone Hfq protein as additional regulatory proteins of vibrioferrin biosynthesis. We found that vibrioferrin production was greatly impaired in both the ryhB and hfq deletion mutants, and a TargetRNA search (http://snowwhite.wellesley.edu/targetRNA/index2.html) revealed that the 5′-untranslated region of pvsOp mRNA (pvsOp 5′-UTR) contains a potential base-pairing region required for the formation of the RyhB-pvsOp 5′-UTR duplex. An electrophoresis mobility shift assay indicated that RyhB can directly bind to the pvsOp 5′-UTR with the aid of Hfq. Rifampin chase experiments indicated that the half-life of pvsOp mRNA in the ryhB and hfq mutants was approximately 3-fold shorter than that in the parental strain, suggesting that both RyhB and Hfq are engaged in the stabilization of pvsOp mRNA. Chrome azurol S assays followed by electrophoresis mobility shift assays and rifampin chase experiments carried out for mutant strains indicated that base pairing between RyhB and the pvsOp 5′-UTR results in an increase in the stability of pvsOp mRNA, thereby leading to the promotion of vibrioferrin production. It is unprecedented that RyhB confers increased stability on a polycistronic mRNA involved in siderophore biosynthesis as a direct target. PMID:23772063

  3. Fatty acid hydrolysis of acyl marinobactin siderophores by Marinobacter acylases.

    PubMed

    Kem, Michelle P; Naka, Hiroaki; Iinishi, Akira; Haygood, Margo G; Butler, Alison

    2015-01-27

    The marine bacteria Marinobacter sp. DS40M6 and Marinobacter nanhaiticus D15-8W produce a suite of acyl peptidic marinobactin siderophores to acquire iron under iron-limiting conditions. During late-log phase growth, the marinobactins are hydrolyzed to form the marinobactin headgroup with release of the corresponding fatty acid tail. The bntA gene, a homologue of the Pseudomonas aeruginosa pyoverdine acylase gene, pvdQ, was identified from Marinobacter sp. DS40M6. A bntA knockout mutant of Marinobacter sp. DS40M6 produced the suite of acyl marinobactins A-E, without the usual formation of the marinobactin headgroup. Another marinobactin-producing species, M. nanhaiticus D15-8W, is predicted to have two pvdQ homologues, mhtA and mhtB. MhtA and MhtB have 67% identical amino acid sequences. MhtA catalyzes hydrolysis of the apo-marinobactin siderophores as well as the quorum sensing signaling molecule, dodecanoyl-homoserine lactone. In contrast to hydrolysis of the suite of apo-marinobactins by MhtA, hydrolysis of the iron(III)-bound marinobactins was not observed. PMID:25588131

  4. Siderophore-mediated cooperation and virulence in Pseudomonas aeruginosa.

    PubMed

    Buckling, Angus; Harrison, Freya; Vos, Michiel; Brockhurst, Michael A; Gardner, Andy; West, Stuart A; Griffin, Ashleigh

    2007-11-01

    Why should organisms cooperate with each other? Helping close relatives that are likely to share the same genes (kin selection) is one important explanation that is likely to apply across taxa. The production of metabolically costly extracellular iron-scavenging molecules (siderophores) by microorganisms is a cooperative behaviour because it benefits nearby conspecifics. We review experiments focusing on the production of the primary siderophore (pyoverdin) of the opportunistic bacterial pathogen, Pseudomonas aeruginosa, which test kin selection theories that seek to explain the evolution of cooperation. First, cooperation is indeed favoured when individuals interact with their close relatives and when there is competition between groups of cooperators and noncooperators, such that the benefit of cooperation can be realized. Second, the relative success of cheats and cooperators is a function of their frequencies within populations. Third, elevated mutation rates can confer a selective disadvantage under conditions when cooperation is beneficial, because high mutation rates reduce how closely bacteria are related to each other. Fourth, cooperative pyoverdin production is also shown to be favoured by kin selection in vivo (caterpillars), and results in more virulent infections. Finally, we briefly outline ongoing and future work using this experimental system. PMID:17919300

  5. An investigation of siderophore production by oceanic Synechococcus

    NASA Astrophysics Data System (ADS)

    Wisniewski, R. J.; Webb, E. A.; Moffett, J. W.

    2003-04-01

    Cyanobacteria are significant contributors to global primary production. They can be found in warm high-nutrient, low-chlorophyll regions where low concentrations of iron are thought to limit primary productivity. Determining how these organisms obtain iron is critical to understanding the biogeochemical cycle of iron and its role as a determinant of marine primary production. Siderophore production has been observed in halotolerant freshwater cyanobacteria (see C.G. Trick and co-authors) and marine heterotrophic bacteria (see A. Butler, M.G. Haygood and co-authors), but to date, siderophore production in truly marine cyanobacteria has not been demonstrated. We examined the response of two marine Synechococcus species (WH7803 and WH8102) to iron stress. Axenic cultures of both Synechococcus species were grown under iron-stressed and iron-replete conditions. The supernatants of these cultures were examined using competitive ligand exchange-cathodic stripping voltammetry (CLE-CSV), a sensitive method of quantitative ligand detection. Observing ligand accumulation in culture is an analytical challenge due to the low cell densities and reduced growth rates of iron stressed marine cyanobacteria. Preliminary results suggest the presence of an iron-binding ligand in the iron-stressed cultures which was not present under iron-replete conditions. The amount of ligand produced by Synechococcus was approximately 1 × 10-18 mol/cell, comparable with the amount produced by marine heterotrophic bacteria (K. Barbeau, pers. comm.).

  6. Pseudomonas siderophores in the sputum of patients with cystic fibrosis.

    PubMed

    Martin, Lois W; Reid, David W; Sharples, Katrina J; Lamont, Iain L

    2011-12-01

    The lungs of patients with cystic fibrosis become chronically infected with the bacterium Pseudomonas aeruginosa, which heralds progressive lung damage and a decline in health. Iron is a crucial micronutrient for bacteria and its acquisition is a key factor in infection. P. aeruginosa can acquire this element by secreting pyoverdine and pyochelin, iron-chelating compounds (siderophores) that scavenge iron and deliver it to the bacteria. Siderophore-mediated iron uptake is generally considered a key factor in the ability of P. aeruginosa to cause infection. We have investigated the amounts of pyoverdine in 148 sputum samples from 36 cystic fibrosis patients (30 infected with P. aeruginosa and 6 as negative controls). Pyoverdine was present in 93 samples in concentrations between 0.30 and 51 μM (median 4.6 μM) and there was a strong association between the amount of pyoverdine and the number of P. aeruginosa present. However, pyoverdine was not present, or below the limits of detection (~0.3 μM), in 21 sputum samples that contained P. aeruginosa. Pyochelin was also absent, or below the limits of detection (~1 μM), in samples from P. aeruginosa-infected patients with little or no detectable pyoverdine. Our data show that pyoverdine is an important iron-scavenging molecule for P. aeruginosa in many cystic fibrosis patients, but other P. aeruginosa iron-uptake systems must be active in some patients to satisfy the bacterial need for iron. PMID:21643731

  7. Burkholderia genome mining for nonribosomal peptide synthetases reveals a great potential for novel siderophores and lipopeptides synthesis.

    PubMed

    Esmaeel, Qassim; Pupin, Maude; Kieu, Nam Phuong; Chataigné, Gabrielle; Béchet, Max; Deravel, Jovana; Krier, François; Höfte, Monica; Jacques, Philippe; Leclère, Valérie

    2016-06-01

    Burkholderia is an important genus encompassing a variety of species, including pathogenic strains as well as strains that promote plant growth. We have carried out a global strategy, which combined two complementary approaches. The first one is genome guided with deep analysis of genome sequences and the second one is assay guided with experiments to support the predictions obtained in silico. This efficient screening for new secondary metabolites, performed on 48 gapless genomes of Burkholderia species, revealed a total of 161 clusters containing nonribosomal peptide synthetases (NRPSs), with the potential to synthesize at least 11 novel products. Most of them are siderophores or lipopeptides, two classes of products with potential application in biocontrol. The strategy led to the identification, for the first time, of the cluster for cepaciachelin biosynthesis in the genome of Burkholderia ambifaria AMMD and a cluster corresponding to a new malleobactin-like siderophore, called phymabactin, was identified in Burkholderia phymatum STM815 genome. In both cases, the siderophore was produced when the strain was grown in iron-limited conditions. Elsewhere, the cluster for the antifungal burkholdin was detected in the genome of B. ambifaria AMMD and also Burkholderia sp. KJ006. Burkholderia pseudomallei strains harbor the genetic potential to produce a novel lipopeptide called burkhomycin, containing a peptidyl moiety of 12 monomers. A mixture of lipopeptides produced by Burkholderia rhizoxinica lowered the surface tension of the supernatant from 70 to 27 mN·m(-1) . The production of nonribosomal secondary metabolites seems related to the three phylogenetic groups obtained from 16S rRNA sequences. Moreover, the genome-mining approach gave new insights into the nonribosomal synthesis exemplified by the identification of dual C/E domains in lipopeptide NRPSs, up to now essentially found in Pseudomonas strains. PMID:27060604

  8. FptA, the Fe(III)-pyochelin receptor of Pseudomonas aeruginosa: a phenolate siderophore receptor homologous to hydroxamate siderophore receptors.

    PubMed Central

    Ankenbauer, R G; Quan, H N

    1994-01-01

    The Pseudomonas aeruginosa siderophore pyochelin is structurally unique among siderophores and possesses neither hydroxamate- nor catecholate-chelating groups. The structural gene encoding the 75-kDa outer membrane Fe(III)-pyochelin receptor FptA has been isolated by plasmid rescue techniques and sequenced. The N-terminal amino acid sequence of the isolated FptA protein corresponded to that deduced from the nucleotide sequence of the fptA structural gene. The mature FptA protein has 682 amino acids and a molecular mass of 75,993 Da and has considerable overall homology with the hydroxamate siderophore receptors FpvA of P. aeruginosa, PupA and PupB of Pseudomonas putida, and FhuE of Escherichia coli. This observation indicates that homologies between siderophore receptors are an unreliable predictor of siderophore ligand class recognition by a given receptor. The fptA gene was strongly regulated by iron; fptA transcription was totally repressed by 30 microM FeCl3, as determined by Northern (RNA) blotting. The promoter of the fptA gene contained the sequence 5'-ATAATGATAAGCATTATC-3', which matches the consensus E. coli Fur-binding site at 17 of 18 positions. The -10 promoter region and transcriptional start site of the fptA gene reside within this Fur-binding site. Images PMID:8288523

  9. Chemical Determinants of antimalarial activity of reversed siderophores.

    PubMed Central

    Tsafack, A; Libman, J; Shanzer, A; Cabantchik, Z I

    1996-01-01

    Reversed siderophores (RSFs) are artificial hydroxamate-based iron chelators designed after the natural siderophore ferrichrome. The modular molecular design of RSF derivatives allowed the synthesis of various congeners with controlled iron-binding capacities and partition coefficients. These two physicochemical properties were assessed by a novel fluorescent method and were found to be the major determinants of RSF permeation across erythrocyte membranes and scavenging of compartmentalized iron. The partition coefficient apparently conferred upon RSFs two major features: (i) the ability to rapidly access iron pools of in vitro-grown Plasmodium falciparum at all developmental stages and to mobilize intracellular iron and transfer it to the medium and (ii) the ability to suppress parasite growth at all developmental stages. These features of RSFs were assessed by quantitative determination of the structure-activity relationships of the biological activities and partition coefficients spanning a wide range of values. The most effective RSF containing the aromatic group of phenylalanine (RSFm2phe) showed 50% inhibitory concentration of 0.60 +/- 0.03 nmol/ml in a 48-h test and a 2-h onset of inhibition of ring development at 5 nmol/ml. The lipophilic compound RSFm2phe and the lipophilic and esterase-cleavable compound RSFm2pee inhibited parasite growth at all developmental stages whether inhibition was assessed in a continuous mode or after discontinuing drug administration. The antimalarial effects of RSFm2phe and cleavable RSFm2pee were potentiated in the presence of desferrioxamine (DFO) at concentrations at which DFO alone had no effect on parasite growth. These studies provide experimental evidence indicating that the effective and persistent antimalarial actions of RSFs are associated with drug access to infected cells and scavenging of iron from intracellular parasites. Moreover, the optimal antimalarial actions of RSFs are apparently also determined by improved

  10. Iron acquisition in the marine actinomycete genus Salinispora is controlled by the desferrioxamine family of siderophores

    PubMed Central

    Roberts, Alexandra A.; Schultz, Andrew W.; Kersten, Roland D.; Dorrestein, Pieter C.; Moore, Bradley S.

    2012-01-01

    Many bacteria produce siderophores for sequestration of growth-essential iron. Analysis of the Salinispora genomes suggests that these marine actinomycetes support multiple hydroxamate- and phenolate-type siderophore pathways. We isolated and characterized desferrioxamines (DFOs) B and E from all three recognized Salinispora species and linked their biosyntheses in S. tropica CNB-440 and S. arenicola CNS-205 to the des locus through PCR-directed mutagenesis. Gene inactivation of the predicted iron-chelator biosynthetic loci sid2-4 did not abolish siderophore chemistry. Additionally, these pathways could not restore the native growth characteristics of the des mutants in iron-limited media, although differential iron-dependent regulation was observed for the yersiniabactin-like sid2 pathway. Consequently, this study indicates that DFOs are the primary siderophores in laboratory cultures of Salinispora. PMID:22812504

  11. Synthesis of siderophores by strains of Staphylococcus cohnii isolated from various environments.

    PubMed

    Szarapińska-Kwaszewska, Jadwiga; Farkas, Lukasz I

    2003-01-01

    Siderophore activity as the feature of microorganisms enabling colonization of human body and the survival in inanimate environment was investigated in 108 strains of Staphylococcus cohnii; S. cohnii ssp. cohnii (50 strains) and S. cohnii ssp. urealyticus (58 strains). Strains were isolated from people, hospital and non-hospital environment. Highest siderophore activity was noted in strains S. cohnii ssp. urealyticus particularly from the inanimate environments origin. In 86% analyzed strains siderophores of hydroxamate class were detected. Larger amounts of these compounds were synthesized in strains S. cohnii ssp. urealyticus. Strains belonging to both subspecies from human origin showed lower activity of siderophores (total pool) and did not produce hydroxamate class chelators or produced very small amounts of these compounds. PMID:14743978

  12. Development and application of an assay for uranyl complexation by fungal metabolites, including siderophores.

    PubMed

    Renshaw, Joanna C; Halliday, Verity; Robson, Geoffrey D; Trinci, Anthony P J; Wiebe, Marilyn G; Livens, Francis R; Collison, David; Taylor, Robin J

    2003-06-01

    An assay to detect UO(2)(2+) complexation was developed based on the chrome azurol S (CAS) assay for siderophores (B. Schwyn and J. B. Neilands, Anal. Biochem. 160:47-56, 1987) and was used to investigate the ability of fungal metabolites to complex actinides. In this assay the discoloration of two dyed agars (one containing a CAS-Fe(3+) dye and the other containing a CAS-UO(2)(2+) dye) caused by ligands was quantified. The assay was tested by using the siderophore desferrioxamine B (DFO), and the results showed that there was a regular, reproducible relationship between discoloration and the amount of siderophore added. The ratio of the discoloration on the CAS-UO(2)(2+) agar to the discoloration on the CAS-Fe(3+) agar was independent of the amount of siderophore added. A total of 113 fungi and yeasts were isolated from three soil samples taken from the Peak District National Park. The fungi were screened for the production of UO(2)(2+) chelators by using the CAS-based assay and were also tested specifically for hydroxamate siderophore production by using the hydroxamate siderophore auxotroph Aureobacterium flavescens JG-9. This organism is highly sensitive to the presence of hydroxamate siderophores. However, the CAS-based assay was found to be less sensitive than the A. flavescens JG-9 assay. No significant difference between the results for each site for the two tests was found. Three isolates were selected for further study and were identified as two Pencillium species and a Mucor species. Our results show that the new assay can be effectively used to screen fungi for the production of UO(2)(2+) chelating ligands. We suggest that hydroxamate siderophores can be produced by mucoraceous fungi. PMID:12788768

  13. The role of siderophores in metal homeostasis of members of the genus Burkholderia.

    PubMed

    Mathew, Anugraha; Jenul, Christian; Carlier, Aurelien L; Eberl, Leo

    2016-02-01

    Although members of the genus Burkholderia can utilize a high-affinity iron uptake system to sustain growth under iron-limiting conditions, many strains also produce siderophores, suggesting that they may serve alternative functions. Here we demonstrate that the two Burkholderia siderophores pyochelin and ornibactin can protect the cells from metal toxicity and thus play an alternative role in metal homeostasis. We also demonstrate that metals such as copper and zinc induce the production of ornibactin. PMID:26621188

  14. Siderophore production by actinomycetes isolates from two soil sites in Western Australia.

    PubMed

    Lee, Joanna; Postmaster, Armin; Soon, Hooi Peng; Keast, David; Carson, Kerry C

    2012-04-01

    The actinomycetes are metabolically flexible soil micro-organisms capable of producing a range of compounds of interest, including siderophores. Siderophore production by actinomycetes sampled from two distinct and separate geographical sites in Western Australia were investigated and found to be generally similar in the total percentage of siderophore producers found. The only notable difference was the proportion of isolates producing catechol siderophores with only 3% found in site 1 (from the north-west of Western Australia and reportedly containing 40% magnetite) and 17% in site 2 (a commercial stone fruit orchard in the hills east of Perth with a soil base ranging from sandy loam to laterite). Further detailed characterization of isolates of interest identified a Streptomyces that produced extracellularly excreted enterobactin, the characteristic Enterobacteriaceae siderophore, and also revealed some of the conditions required for enterobactin production. Carriage of the entF gene, which codes for the synthetase responsible for the final assembly of the tri-cyclic structure of enterobactin, was confirmed by PCR in this isolate. Another separate Streptomyces produced a compound that matched the UV/VIS spectra of heterobactin, a siderophore previously only described in Rhodococcus and Nocardia. PMID:22038645

  15. The fate of siderophores: antagonistic environmental interactions in exudate-mediated micronutrient uptake.

    PubMed

    Harrington, James M; Duckworth, Owen W; Haselwandter, Kurt

    2015-06-01

    Organisms acquire metals from the environment by releasing small molecules that solubilize and promote their specific uptake. The best known example of this nutrient uptake strategy is the exudation of siderophores, which are a structurally-diverse class of molecules that are traditionally viewed as being integral to iron uptake. Siderophores have been proposed to act through a variety of processes, but their effectiveness can be mitigated by a variety of chemical and physical processes of both biotic and abiotic origin. Processes that occur at the surface of minerals can degrade or sequester siderophores, preventing them from fulfilling their function of returning metals to the organism. In addition, biotic processes including enzymatic degradation of the siderophore and piracy of the metal or of the siderophore complex also disrupt iron uptake. Some organisms have adapted their nutrient acquisition strategies to address these potential pitfalls, producing multiple siderophores and other exudates that take advantage of varying kinetic and thermodynamic factors to allow the continued uptake of metals. A complete understanding of the factors that contribute to metal uptake in nature will require a concerted effort to study processes identified in laboratory systems in the context of more complicated environmental systems. PMID:25619589

  16. Analysis of the Corynebacterium diphtheriae DtxR regulon: identification of a putative siderophore synthesis and transport system that is similar to the Yersinia high-pathogenicity island-encoded yersiniabactin synthesis and uptake system.

    PubMed

    Kunkle, Carey A; Schmitt, Michael P

    2003-12-01

    The diphtheria toxin repressor, DtxR, is a global iron-dependent regulatory protein in Corynebacterium diphtheriae that controls gene expression by binding to 19-bp operator sequences. To further define the DtxR regulon in C. diphtheriae, a DtxR repressor titration assay (DRTA) was developed and used to identify 10 previously unknown DtxR binding sites. Open reading frames downstream from seven of the newly identified DtxR binding sites are predicted to encode proteins associated with iron or heme transport. Electrophoretic mobility shift assays indicated that DtxR was able to bind to DNA fragments carrying the 19-bp operator regions, and transcriptional analysis of putative promoter elements adjacent to the binding site sequences revealed that most of these regions displayed iron- and DtxR-regulated activity. A putative siderophore biosynthesis and transport operon located downstream from one of the DtxR binding sites, designated sid, is similar to the yersiniabactin synthesis and uptake genes encoded on the Yersinia pestis high pathogenicity island. The siderophore biosynthetic genes in the sid operon contained a large deletion in the C. diphtheriae C7 strain, but the sid genes were unaffected in four clinical isolates that are representative of the dominant strains from the recent diphtheria epidemic in the former Soviet Union. Mutations in the siderophore biosynthetic genes in a clinical strain had no effect on siderophore synthesis or growth in low-iron conditions; however, a mutation in one of the putative transport proteins, cdtP, resulted in reduced growth in iron-depleted media, which suggests that this system may have a role in iron uptake. The findings from this study indicate that C. diphtheriae contains at least 18 DtxR binding sites and that DtxR may affect the expression of as many as 40 genes. PMID:14617647

  17. Potentiation of antibacterial activity of the MB-1 siderophore-monobactam conjugate using an efflux pump inhibitor.

    PubMed

    Tomaras, Andrew P; Crandon, Jared L; McPherson, Craig J; Nicolau, David P

    2015-04-01

    Preliminary enthusiasm over the encouraging spectrum and in vitro activities of siderophore conjugates, such as MB-1, was recently tempered by unexpected variability in in vivo efficacy. The need for these conjugates to compete for iron with endogenously produced siderophores has exposed a significant liability for this novel antibacterial strategy. Here, we have exploited dependence on efflux for siderophore secretion in Pseudomonas aeruginosa and provide evidence that efflux inhibition may circumvent this in vivo-relevant resistance liability. PMID:25605364

  18. Potentiation of Antibacterial Activity of the MB-1 Siderophore-Monobactam Conjugate Using an Efflux Pump Inhibitor

    PubMed Central

    Crandon, Jared L.; McPherson, Craig J.; Nicolau, David P.

    2015-01-01

    Preliminary enthusiasm over the encouraging spectrum and in vitro activities of siderophore conjugates, such as MB-1, was recently tempered by unexpected variability in in vivo efficacy. The need for these conjugates to compete for iron with endogenously produced siderophores has exposed a significant liability for this novel antibacterial strategy. Here, we have exploited dependence on efflux for siderophore secretion in Pseudomonas aeruginosa and provide evidence that efflux inhibition may circumvent this in vivo-relevant resistance liability. PMID:25605364

  19. Pumping iron to keep fit: modulation of siderophore secretion helps efficient aromatic utilization in Pseudomonas putida KT2440.

    PubMed

    Joshi, Hiren; Dave, Rachna; Venugopalan, V P

    2014-07-01

    Studies of biotechnology applications of Pseudomonas putida KT2440 have been predominantly focused on regulation and expression of the toluene degradation (TOL) pathway. Unfortunately, there is limited information on the role of other physiological factors influencing aromatic utilization. In this report, we demonstrate that P. putida KT2440 increases its siderophore secretion in response to the availability of benzyl alcohol, a model aromatic substrate. It is argued that accelerated siderophore secretion in response to aromatic substrates provides an iron 'boost' which is required for the effective functioning of the iron-dependent oxygenases responsible for ring opening. Direct evidence for the cardinal role of siderophores in aromatic utilization is provided by evaluation of per capita siderophore secretion and comparative growth assessments of wild-type and siderophore-negative mutant strains grown on an alternative carbon source. Accelerated siderophore secretion can be viewed as a compensatory mechanism in P. putida in the context of its inability to secrete more than one type of siderophore (pyoverdine) or to utilize heterologous siderophores. Stimulated siderophore secretion might be a key factor in successful integration and proliferation of this organism as a bio-augmentation agent for aromatic degradation. It not only facilitates efficient aromatic utilization, but also provides better opportunities for iron assimilation amongst diverse microbial communities, thereby ensuring better survival and proliferation. PMID:24742959

  20. Siderophore-Producing Bacteria from a Sand Dune Ecosystem and the Effect of Sodium Benzoate on Siderophore Production by a Potential Isolate

    PubMed Central

    Gaonkar, Teja; Nayak, Pramoda Kumar; Garg, Sandeep; Bhosle, Saroj

    2012-01-01

    Bioremediation in natural ecosystems is dependent upon the availability of micronutrients and cofactors, of which iron is one of the essential elements. Under aerobic and alkaline conditions, iron oxidizes to Fe+3 creating iron deficiency. To acquire this essential growth-limiting nutrient, bacteria produce low-molecular-weight, high-affinity iron chelators termed siderophores. In this study, siderophore-producing bacteria from rhizosphere and nonrhizosphere areas of coastal sand dunes were isolated using a culture-dependent approach and were assigned to 8 different genera with the predominance of Bacillus sp. Studies on the ability of these isolates to grow on sodium benzoate revealed that a pigmented bacterial culture TMR2.13 identified as Pseudomonas aeruginosa showed growth on mineral salts medium (MSM) with 2% of sodium benzoate and produced a yellowish fluorescent siderophore identified as pyoverdine. This was inhibited above 54 μM of added iron in MSM with glucose without affecting growth, while, in presence of sodium benzoate, siderophore was produced even up to the presence of 108 μM of added iron. Increase in the requirement of iron for metabolism of aromatic compounds in ecosystems where the nutrient deficiencies occur naturally would be one of the regulating factors for the bioremediation process. PMID:22629215

  1. Unusual non-fluorescent broad spectrum siderophore activity (SID EGYII) by Pseudomonas aeruginosa strain EGYII DSM 101801 and a new insight towards simple siderophore bioassay.

    PubMed

    Embaby, Amira M; Heshmat, Yasmin; Hussein, Ahmed

    2016-03-01

    Present study highlights an unusual non-fluorescent hydroxamate broad spectrum siderophore (SID EGYII) activity from Pseudomonas aeruginosa strain EGYII DSM 101801, a soil bacterial isolate, along with simple low cost effective siderophore bioassay. Detection of SID EGYII activity qualitatively was proved by masking this activity against Erwinia amylovora strain EGY1 DSM 101800, an indicator strain, in well-cut diffusion assay containing 100 µM FeCl3. SID EGYII activity was expressed quantitatively as arbitrary units [Siderophore arbitrary units (SAU)] 380 SAU/mL against E. amylovora strain EGY1 DSM 101800. Maximal SID EGYII activity was achieved upon growing P. aeruginosa strain EGYII DSM 101801 in PYB broth at 180 rpm for 24 h. SID EGYII displayed a broad spectrum antimicrobial activity against some human pathogens (i.e., Gram-positive bacteria, Gram-negative bacteria and yeasts) and a fireblight plant pathogen. Interestingly, transformants of Escherichia coli JM109 (DE3)pSID/EGYII harboring P. aeruginosa strain EGYII DSM 101801 plasmid demonstrated a perceivable antimicrobial activity against E. amylovora strain EGY1 DSM 101800. The broad spectrum antimicrobial activity of the unusual non-fluorescent SID EGYII would underpin its high potential in targeting bacterial pathogens posing probable threats to human health and agricultural economy. The present simple low cost effective bioassay is a new insight towards an alternative to the expensive cumbersome siderophore Chrome Azurol S assay. PMID:27015845

  2. Bacterial siderophores efficiently provide iron to iron-starved tomato plants in hydroponics culture.

    PubMed

    Radzki, W; Gutierrez Mañero, F J; Algar, E; Lucas García, J A; García-Villaraco, A; Ramos Solano, B

    2013-09-01

    Iron is one of the essential elements for a proper plant development. Providing plants with an accessible form of iron is crucial when it is scant or unavailable in soils. Chemical chelates are the only current alternative and are highly stable in soils, therefore, posing a threat to drinking water. The aim of this investigation was to quantify siderophores produced by two bacterial strains and to determine if these bacterial siderophores would palliate chlorotic symptoms of iron-starved tomato plants. For this purpose, siderophore production in MM9 medium by two selected bacterial strains was quantified, and the best was used for biological assay. Bacterial culture media free of bacteria (S) and with bacterial cells (BS), both supplemented with Fe were delivered to 12-week-old plants grown under iron starvation in hydroponic conditions; controls with full Hoagland solution, iron-free Hoagland solution and water were also conducted. Treatments were applied twice along the experiment, with a week in between. At harvest, plant yield, chlorophyll content and nutritional status in leaves were measured. Both the bacterial siderophore treatments significantly increased plant yield, chlorophyll and iron content over the positive controls with full Hoagland solution, indicating that siderophores are effective in providing Fe to the plant, either with or without the presence of bacteria. In summary, siderophores from strain Chryseobacterium C138 are effective in supplying Fe to iron-starved tomato plants by the roots, either with or without the presence of bacteria. Based on the amount of siderophores produced, an effective and economically feasible organic Fe chelator could be developed. PMID:23812968

  3. Selective ciprofloxacin antibiotic detection by fluorescent siderophore pyoverdin.

    PubMed

    Pawar, Madhuri K; Tayade, Kundan C; Sahoo, Suban K; Mahulikar, Pramod P; Kuwar, Anil S; Chaudhari, Bhushan L

    2016-07-15

    Fluorescent siderophore pyoverdin (PVD) was produced from a soil isolate Pseudomonas monteilii strain MKP 213. The PVD was purified near to homogeneity and applied for the fluorescent chemosensing of various antibiotics in aqueous solution (pH=7.0). Upon addition of ciprofloxacin, PVD showed new UV-vis absorption bands at 252 and 321nm due to an internal charge transfer mechanism. Also, the addition of ciprofloxacin induced a highly selective fluorescence enhancement of PVD with a 13nm blue shift from 458 to 445nm. The combination of a long peptide chain along with the chromophore unit of PVD generates a converging cleft for ciprofloxacin recognition with LOD and LOQ of 7.13μM and 21.6μM, respectively without interference from other studied antibiotics. The association constant (Ka) of PVD with ciprofloxacin was calculated to be as low as 1.40×10(5)M(-1) using Benesi-Hildebrand plot depicting its significance in detection. The pharmaceutical tablet analysis measures the sensing with negligible matrix effect and quantitative recovery. PMID:26971273

  4. Chemical synthesis and biological evaluation of gallidermin-siderophore conjugates.

    PubMed

    Yoganathan, Sabesan; Sit, Clarissa S; Vederas, John C

    2011-04-01

    The lantibiotic gallidermin was modified at lysine residues by regioselective attachment of derivatives of pyochelin, agrobactin and desferrioxamine B with the objective of having siderophore receptors of Gram-negative bacteria transport the antibiotic-iron chelator conjugate through the outer membrane. All of the conjugates retained activity against the Gram-positive indicator strain, Lactococcus lactis subsp. cremoris HP. However, testing of the conjugates against several Gram-negative strains yielded unexpected results. Bacteria treated with 100 μM of the conjugates complexed with Fe(3+) grew better than bacteria grown in iron-free media but worse than bacteria grown in the same media supplemented with 10 μM FeCl(3). Although these findings indicate that the conjugates are unable to inhibit the growth of Gram-negative bacteria, they indicate penetration of the outer membrane and provide structure-activity information for design of other lantibiotic conjugates. The synthetic strategy is applicable for linking biomarkers or fluorescence probes to gallidermin for studies on its localization and mode of action. As there are many lantibiotics that operate with unknown mechanisms of action, this chemical approach provides a means to modify such peptides with biomarkers for biological investigations. PMID:21290068

  5. Complexation of Pu(IV) with the natural siderophore desferrioxamine B and the redox properties of Pu(IV)(siderophore) complexes.

    PubMed

    Boukhalfa, Hakim; Reilly, Sean D; Neu, Mary P

    2007-02-01

    The bioavailability and mobility of Pu species can be profoundly affected by siderophores and other oxygen-rich organic ligands. Pu(IV)(siderophore) complexes are generally soluble and may constitute with other soluble organo-Pu(IV) complexes the main fraction of soluble Pu(IV) in the environment. In order to understand the impact of siderophores on the behavior of Pu species, it is important to characterize the formation and redox behavior of Pu(siderophore) complexes. In this work, desferrioxamine B (DFO-B) was investigated for its capacity to bind Pu(IV) as a model siderophore and the properties of the complexes formed were characterized by optical spectroscopy measurements. In a 1:1 Pu(IV)/DFO-B ratio, the complexes Pu(IV)(H2DFO-B)4+, Pu(IV)(H1DFO-B)3+, Pu(IV)(DFO-B)2+, and Pu(IV)(DFO-B)(OH)+ form with corresponding thermodynamic stability constants log beta1,1,2 = 35.48, log beta1,1,1 = 34.87, log beta1,1,0 = 33.98, and log beta1,1,-1 = 27.33, respectively. In the presence of excess DFO-B, the complex Pu(IV)H2(DFO-B)22+ forms with the formation constant log beta2,1,2 = 62.30. The redox potential of the complex Pu(IV)H2(DFO-B)22+ was determined by cyclic voltammetry to be E1/2 = -0.509 V, and the redox potential of the complex Pu(IV)(DFO-B)2+ was estimated to be E1/2 = -0.269 V. The redox properties of Pu(IV)(DFO-B)2+ complexes indicate that Pu(III)(siderophore) complexes are more than 20 orders of magnitude less stable than their Pu(IV) analogues. This indicates that under reducing conditions, stable Pu(siderophore) complexes are unlikely to persist. PMID:17257046

  6. Characterization of Fluorescent and Nonfluorescent Peptide Siderophores Produced by Pseudomonas syringae Strains and Their Potential Use in Strain Identification

    PubMed Central

    Bultreys, Alain; Gheysen, Isabelle; Maraite, Henri; de Hoffmann, Edmond

    2001-01-01

    Nonfluorescent highly virulent strains of Pseudomonas syringae pv. aptata isolated in different European countries and in Uruguay produce a nonfluorescent peptide siderophore, the production of which is iron repressed and specific to these strains. The amino acid composition of this siderophore is identical to that of the dominant fluorescent peptide siderophore produced by fluorescent P. syringae strains, and the molecular masses of the respective Fe(III) chelates are 1,177 and 1,175 atomic mass units. The unchelated nonfluorescent siderophore is converted into the fluorescent siderophore at pH 10, and colors and spectral characteristics of the unchelated siderophores and of the Fe(III)-chelates in acidic conditions are similar to those of dihydropyoverdins and pyoverdins, respectively. The nonfluorescent siderophore is used by fluorescent and nonfluorescent P. syringae strains. These results and additional mass spectrometry data strongly suggest the presence of a pyoverdin chromophore in the fluorescent siderophore and a dihydropyoverdin chromophore in the nonfluorescent siderophore, which are both ligated to a succinamide residue. When chelated, the siderophores behave differently from typical pyoverdins and dihydropyoverdins in neutral and alkaline conditions, apparently because of the ionization occurring around pH 4.5 of carboxylic acids present in β-hydroxyaspartic acid residues of the peptide chains. These differences can be detected visually by pH-dependent changes of the chelate colors and spectrophotochemically. These characteristics and the electrophoretic behavior of the unchelated and chelated siderophores offer new tools to discriminate between saprophytic fluorescent Pseudomonas species and fluorescent P. syringae and P. viridiflava strains and to distinguish between the two siderovars in P. syringae pv. aptata. PMID:11282626

  7. The heterologous siderophores ferrioxamine B and ferrichrome activate signaling pathways in Pseudomonas aeruginosa.

    PubMed

    Llamas, María A; Sparrius, Marion; Kloet, Roy; Jiménez, Connie R; Vandenbroucke-Grauls, Christina; Bitter, Wilbert

    2006-03-01

    Pseudomonas aeruginosa secretes two siderophores, pyoverdine and pyochelin, under iron-limiting conditions. These siderophores are recognized at the cell surface by specific outer membrane receptors, also known as TonB-dependent receptors. In addition, this bacterium is also able to incorporate many heterologous siderophores of bacterial or fungal origin, which is reflected by the presence of 32 additional genes encoding putative TonB-dependent receptors. In this work, we have used a proteomic approach to identify the inducing conditions for P. aeruginosa TonB-dependent receptors. In total, 11 of these receptors could be discerned under various conditions. Two of them are only produced in the presence of the hydroxamate siderophores ferrioxamine B and ferrichrome. Regulation of their synthesis is affected by both iron and the presence of a cognate siderophore. Analysis of the P. aeruginosa genome showed that both receptor genes are located next to a regulatory locus encoding an extracytoplasmic function sigma factor and a transmembrane sensor. The involvement of this putative regulatory locus in the specific induction of the ferrioxamine B and ferrichrome receptors has been demonstrated. These results show that P. aeruginosa has evolved multiple specific regulatory systems to allow the regulation of TonB-dependent receptors. PMID:16484199

  8. Scavenging iron: a novel mechanism of plant immunity activation by microbial siderophores.

    PubMed

    Aznar, Aude; Chen, Nicolas W G; Rigault, Martine; Riache, Nassima; Joseph, Delphine; Desmaële, Didier; Mouille, Grégory; Boutet, Stéphanie; Soubigou-Taconnat, Ludivine; Renou, Jean-Pierre; Thomine, Sébastien; Expert, Dominique; Dellagi, Alia

    2014-04-01

    Siderophores are specific ferric iron chelators synthesized by virtually all microorganisms in response to iron deficiency. We have previously shown that they promote infection by the phytopathogenic enterobacteria Dickeya dadantii and Erwinia amylovora. Siderophores also have the ability to activate plant immunity. We have used complete Arabidopsis transcriptome microarrays to investigate the global transcriptional modifications in roots and leaves of Arabidopsis (Arabidopsis thaliana) plants after leaf treatment with the siderophore deferrioxamine (DFO). Physiological relevance of these transcriptional modifications was validated experimentally. Immunity and heavy-metal homeostasis were the major processes affected by DFO. These two physiological responses could be activated by a synthetic iron chelator ethylenediamine-di(o-hydroxyphenylacetic) acid, indicating that siderophores eliciting activities rely on their strong iron-chelating capacity. DFO was able to protect Arabidopsis against the pathogenic bacterium Pseudomonas syringae pv tomato DC3000. Siderophore treatment caused local modifications of iron distribution in leaf cells visible by ferrocyanide and diaminobenzidine-H₂O₂ staining. Metal quantifications showed that DFO causes a transient iron and zinc uptake at the root level, which is presumably mediated by the metal transporter iron regulated transporter1 (IRT1). Defense gene expression and callose deposition in response to DFO were compromised in an irt1 mutant. Consistently, plant susceptibility to D. dadantii was increased in the irt1 mutant. Our work shows that iron scavenging is a unique mechanism of immunity activation in plants. It highlights the strong relationship between heavy-metal homeostasis and immunity. PMID:24501001

  9. Scavenging Iron: A Novel Mechanism of Plant Immunity Activation by Microbial Siderophores1[C][W

    PubMed Central

    Aznar, Aude; Chen, Nicolas W.G.; Rigault, Martine; Riache, Nassima; Joseph, Delphine; Desmaële, Didier; Mouille, Grégory; Boutet, Stéphanie; Soubigou-Taconnat, Ludivine; Renou, Jean-Pierre; Thomine, Sébastien; Expert, Dominique; Dellagi, Alia

    2014-01-01

    Siderophores are specific ferric iron chelators synthesized by virtually all microorganisms in response to iron deficiency. We have previously shown that they promote infection by the phytopathogenic enterobacteria Dickeya dadantii and Erwinia amylovora. Siderophores also have the ability to activate plant immunity. We have used complete Arabidopsis transcriptome microarrays to investigate the global transcriptional modifications in roots and leaves of Arabidopsis (Arabidopsis thaliana) plants after leaf treatment with the siderophore deferrioxamine (DFO). Physiological relevance of these transcriptional modifications was validated experimentally. Immunity and heavy-metal homeostasis were the major processes affected by DFO. These two physiological responses could be activated by a synthetic iron chelator ethylenediamine-di(o-hydroxyphenylacetic) acid, indicating that siderophores eliciting activities rely on their strong iron-chelating capacity. DFO was able to protect Arabidopsis against the pathogenic bacterium Pseudomonas syringae pv tomato DC3000. Siderophore treatment caused local modifications of iron distribution in leaf cells visible by ferrocyanide and diaminobenzidine-H2O2 staining. Metal quantifications showed that DFO causes a transient iron and zinc uptake at the root level, which is presumably mediated by the metal transporter iron regulated transporter1 (IRT1). Defense gene expression and callose deposition in response to DFO were compromised in an irt1 mutant. Consistently, plant susceptibility to D. dadantii was increased in the irt1 mutant. Our work shows that iron scavenging is a unique mechanism of immunity activation in plants. It highlights the strong relationship between heavy-metal homeostasis and immunity. PMID:24501001

  10. Siderophore as a potential plant growth-promoting agent produced by Pseudomonas aeruginosa JAS-25.

    PubMed

    Sulochana, M B; Jayachandra, S Y; Kumar, S Anil; Parameshwar, A B; Reddy, K Mohan; Dayanand, A

    2014-09-01

    Siderophores scavenges Fe(+3) from the vicinity of the roots of plants, and thus limit the amount of iron required for the growth of pathogens such as Fusarium oxysporum, Pythium ultimum, and Fusarium udum, which cause wilt and root rot disease in crops. The ability of Pseudomonas to grow and to produce siderophore depends upon the iron content, pH, and temperature. Maximum yield of siderophore of 130 μM was observed at pH 7.0 ± 0.2 and temperature of 30 °C at 30 h. The threshold level of iron was 50 μM, which increases up to 150 μM, favoring growth but drastically affecting the production of siderophore by Pseudomonas aeruginosa JAS-25. The seeds of agricultural crops like Cicer arietinum (chick pea), Cajanus cajan (pigeon pea), and Arachis hypogaea (ground nut) were treated with P. aeruginosa JAS-25, which enhanced the seed germination, root length, shoot length, and dry weight of chick pea, pigeon pea, and ground nut plants under pot studies. The efficient growth of the plants was not only due to the biocontrol activity of the siderophore produced by P. aeruginosa JAS-25 but also may be by the production of indole acetic acid (IAA), which influences the growth of the plants as phytohormones. PMID:25062779

  11. Derepression of the Azotobacter vinelandii siderophore system, using iron-containing minerals to limit iron repletion.

    PubMed Central

    Page, W J; Huyer, M

    1984-01-01

    Azotobacter vinelandii solubilized iron from certain minerals using only dihydroxybenzoic acid, which appeared to be produced constitutively. Solubilization of iron from other minerals required dihydroxybenzoic acid and the siderophore N,N'-bis-(2,3- dihydroxybenzoyl )-L-lysine ( azotochelin ) or these chelators plus the yellow-green fluorescent siderophore azotobactin . In addition to this sequential production of siderophores, cells also demonstrated partial to hyperproduction relative to the iron-limited control. The iron sources which caused partial derepression of the siderophores caused derepression of all the high-molecular-weight iron-repressible outer membrane proteins except a 77,000-molecular-weight protein, which appeared to be coordinated with azotobactin production. Increased siderophore production correlated with increased production of outer membrane proteins with molecular weights of 93,000, 85,000, and 77,000, but an 81,000-molecular-weight iron-repressible protein appeared at a constant level despite the degree of derepression. When iron was readily available, it appeared to complex with a 60,000-molecular-weight protein believed to form a surface layer on the A. vinelandii cell. Images PMID:6233258

  12. Layer plate CAS assay for the quantitation of siderophore production and determination of exudation patterns for fungi.

    PubMed

    Andrews, Megan Y; Santelli, Cara M; Duckworth, Owen W

    2016-02-01

    The chrome azurol S (CAS) assay measures the chelating activity of siderophores, but its application (especially to fungi) is limited by toxicity issues. In this note, we describe a modified version of the CAS assay that is suitable for quantifying siderophore exudation for microorganisms, including fungi. PMID:26712125

  13. Vanchrobactin and Anguibactin Siderophores Produced by Vibrio sp. DS40M4

    PubMed Central

    Sandy, Moriah; Han, Andrew; Blunt, John; Munro, Murray; Haygood, Margo; Butler, Alison

    2011-01-01

    The marine bacterium Vibrio sp. DS40M4 has been found to produce a new triscatechol amide siderophore, trivanchrobactin (1), a related new biscatecholamide compound, divanchrobactin (2), as well as the previously reported siderophores, vanchrobactin (3) and anguibactin (4). Vanchrobactin is comprised of l-serine, d-arginine- and 2,3-dihydroxybenzoic acid, while trivanchrobactin is a linear trimer of vanchrobactin joined by two serine ester linkages. The cyclic trivanchrobactin product was not detected. In addition to siderophore production, extracts of Vibrio sp. DS40M4 were screened for biologically active molecules; anguibactin was found to be cytotoxic against the P388 murine leukemia cell line (IC50 < 15 μM). PMID:20521785

  14. Structural characterization of amphiphilic siderophores produced by a soda lake isolate, Halomonas sp. SL01, reveals cysteine-, phenylalanine- and proline-containing head groups.

    PubMed

    Figueroa, Luis O'mar Serrano; Schwarz, Benjamin; Richards, Abigail M

    2015-11-01

    Soap Lake, located in Washington State, is a naturally occurring saline and alkaline lake. Several organisms inhabiting this lake have been identified as producers of siderophores that are unique in structure. Bacterial isolates, enriched from Soap Lake sediment and water samples, were screened for siderophore production using both the chrome azurol S (CAS) agar plate and liquid methods. Bacterial isolate Halomonas sp. SL01 was found to produce relatively high concentrations of siderophores in liquid medium (up to 40 µM). Siderophores from the isolate were separated from the culture supernatant using solid phase extraction and purified by high-performance liquid chromatography (HPLC). Siderophore structure was determined using LC/MS/MS (liquid chromatography/mass spectrometry/mass spectrometry) and fatty acid methyl ester (FAME) GC. Two distinct new families of amphiphilic siderophores were produced by isolate SL01. All siderophores ranged in size from 989 to 1096 atomic mass units and consisted of a conserved peptidic head group (per family), which coordinates iron, coupled to fatty acid moieties. The fatty acyl moieties were C10-C14 in length and some with hydroxyl substitutions at the third α position. These siderophores resembled amphiphilic aquachelin siderophores produced by Halomonas aquamarina strain DS40M3, a marine bacterium as well as siderophores from isolate Halomonas sp. SL28 that was found to produce amphiphilic siderophores. Bacteria thriving under saline and alkaline conditions are capable of producing unique siderophores resembling those produced by microbes inhabiting marine environments. PMID:26439615

  15. Iron acquisition from Pseudomonas aeruginosa siderophores by human phagocytes: an additional mechanism of host defense through iron sequestration?

    PubMed

    Britigan, B E; Rasmussen, G T; Olakanmi, O; Cox, C D

    2000-03-01

    Chelation of iron to iron-binding proteins is a strategy of host defense. Some pathogens counter this via the secretion of low-molecular-weight iron-chelating agents (siderophores). Human phagocytes possess a high-capacity mechanism for iron acquisition from low-molecular-weight iron chelates. Efficient acquisition and sequestration of iron bound to bacterial siderophores by host phagocytes could provide a secondary mechanism to limit microbial access to iron. In the present work we report that human neutrophils, macrophages, and myeloid cell lines can acquire iron from the two Pseudomonas aeruginosa siderophores. Analogous to iron acquisition from other low-molecular-weight chelates, iron acquisition from the siderophores is ATP independent, induced by multivalent cationic metals, and unaffected by inhibitors of endocytosis and pinocytosis. In vivo, this process could serve as an additional mechanism of host defense to limit iron availability to invading siderophore-producing microbes. PMID:10678937

  16. Iron Acquisition from Pseudomonas aeruginosa Siderophores by Human Phagocytes: an Additional Mechanism of Host Defense through Iron Sequestration?

    PubMed Central

    Britigan, Bradley E.; Rasmussen, George T.; Olakanmi, Oyebode; Cox, Charles D.

    2000-01-01

    Chelation of iron to iron-binding proteins is a strategy of host defense. Some pathogens counter this via the secretion of low-molecular-weight iron-chelating agents (siderophores). Human phagocytes possess a high-capacity mechanism for iron acquisition from low-molecular-weight iron chelates. Efficient acquisition and sequestration of iron bound to bacterial siderophores by host phagocytes could provide a secondary mechanism to limit microbial access to iron. In the present work we report that human neutrophils, macrophages, and myeloid cell lines can acquire iron from the two Pseudomonas aeruginosa siderophores. Analogous to iron acquisition from other low-molecular-weight chelates, iron acquisition from the siderophores is ATP independent, induced by multivalent cationic metals, and unaffected by inhibitors of endocytosis and pinocytosis. In vivo, this process could serve as an additional mechanism of host defense to limit iron availability to invading siderophore-producing microbes. PMID:10678937

  17. The effect of soil horizon and mineral type on the distribution of siderophores in soil

    NASA Astrophysics Data System (ADS)

    Ahmed, Engy; Holmström, Sara J. M.

    2014-04-01

    Iron is a key component of the chemical architecture of the biosphere. Due to the low bioavailability of iron in the environment, microorganisms have developed specific uptake strategies like production of siderophores. Siderophores are operationally defined as low-molecular-mass biogenic Fe(III)-binding compounds, that can increase the bioavailability of iron by promoting the dissolution of iron-bearing minerals. In the present study, we investigated the composition of dissolved and adsorbed siderophores of the hydroxamate family in the soil horizons of podzol and the effect of specific mineral types on siderophores. Three polished mineral specimens of 3 cm × 4 cm × 3 mm (apatite, biotite and oligioclase) were inserted in the soil horizons (O (organic), E (eluvial) and B (upper illuvial)). After two years, soil samples were collected from both the bulk soil of the whole profile and from the soil attached to the mineral surfaces. The concentration of ten different fungal tri-hydroxamates within ferrichromes, fusigen and coprogens families, and five bacterial hydroxamates within the ferrioxamine family were detected. All hydroxamate types were determined in both soil water (dissolved) and soil methanol (adsorbed) extracts along the whole soil profile by high-performance liquid chromatography coupled to electrospray ionization mass spectrometry (HPLC-ESI-MS); hence, the study is the most extensive of its kind. We found that coprogens and fusigen were present in much higher concentrations in bulk soil than were ferrioxamines and ferrichromes. On the other hand, the presence of the polished mineral completely altered the distribution of siderophores. In addition, each mineral had a unique interaction with the dissolved and adsorbed hydroxamates in the different soil horizons. Thus siderophore composition in the soil environment is controlled by the chemical, physical and biological characteristics of each soil horizon and also by the available mineral types.

  18. Enzymatic Hydrolysis of Trilactone Siderophores: Where Chiral Recognition Occurs in Enterobactin and Bacillibactin Iron Transport1

    PubMed Central

    Abergel, Rebecca J.; Zawadzka, Anna M.; Hoette, Trisha M.; Raymond, Kenneth N.

    2009-01-01

    Bacillibactin and enterobactin are hexadentate catecholate siderophores produced by bacteria upon iron limitation to scavenge ferric ion and seem to be the ultimate siderophores of their two respective domains: Gram-positive and Gram-negative. Iron acquisition mediated by these trilactone-based ligands necessitates enzymatic hydrolysis of the scaffold for successful intracellular iron delivery. The esterases BesA and Fes hydrolyze bacillibactin and enterobactin, respectively, as well as the corresponding iron complexes. Bacillibactin binds iron through three 2,3-catecholamide moieties linked to a tri-threonine scaffold via glycine spacers, whereas in enterobactin the iron-binding moieties are directly attached to a tri-l-serine backbone; although apparently minor, these structural differences result in markedly different iron coordination properties and iron transport behavior. Comparison of the solution thermodynamic and circular dichroism properties of bacillibactin, enterobactin and the synthetic analogs d-enterobactin, SERGlyCAM and d-SERGlyCAM has determined the role of each different feature in the siderophores' molecular structures in ferric complex stability and metal chirality. While opposite metal chiralities in the different complexes did not affect transport and incorporation in Bacillus subtilis, ferric complexes formed with the various siderophores did not systematically promote growth of the bacteria. The bacillibactin esterase BesA is less specific than the enterobactin esterase Fes; BesA can hydrolyze the trilactones of both siderophores, while only the tri-l-serine trilactone is a substrate of Fes. Both enzymes are stereospecific and cannot cleave tri-d-serine lactones. These data provide a complete picture of the microbial iron transport mediated by these two siderophores, from initial recognition and transport to intracellular iron release. PMID:19673474

  19. Production and Characterization of Desmalonichrome Relative Binding Affinity for Uranyl Ions in Relation to Other Siderophores.

    PubMed

    Mo, Kai-For; Dai, Ziyu; Wunschel, David S

    2016-06-24

    Siderophores are iron (Fe)-binding secondary metabolites that have been investigated for their uranium-binding properties. Previous work has focused on characterizing hydroxamate types of siderophores, such as desferrioxamine B, for their uranyl (UO2)-binding affinity. Carboxylate forms of these metabolites hold potential to be more efficient chelators of UO2, yet they have not been widely studied. Desmalonichrome is a carboxylate siderophore that is not commercially available and so was obtained from the fungus Fusarium oxysporum cultivated under Fe-depleted conditions. The relative affinity for UO2 binding of desmalonichrome was investigated using a competitive analysis of binding affinities between UO2 acetate and different concentrations of Fe(III) chloride using electrospray ionization mass spectrometry. In addition to desmalonichrome, three other siderophores, including two hydroxamates (desferrioxamine B and desferrichrome) and one carboxylate (desferrichrome A), were studied to understand their relative affinities for the UO2(2+) ion at two pH values. The binding affinities of hydroxamate siderophores to UO2(2+) ions were observed to decrease with increasing Fe(III)Cl3 concentration at the lower pH. On the other hand, decreasing the pH has a smaller impact on the binding affinities between carboxylate siderophores and the UO2(2+) ion. Desmalonichrome in particular was shown to have the greatest relative affinity for UO2 at all pH and Fe(III) concentrations examined. These results suggest that acidic functional groups in the ligands are important for strong chelation with UO2 at lower pH. PMID:27232848

  20. Substrate induction of siderophore transport in Bacillus subtilis mediated by a novel one-component regulator

    PubMed Central

    Gaballa, Ahmed; Helmann, John D.

    2011-01-01

    Summary When iron is scarce, Bacillus subtilis expresses genes involved in the synthesis and uptake of the siderophore bacillibactin (BB) and uptake systems to pirate other microbial siderophores. Here, we demonstrate that transcriptional induction of the feuABCybbA operon, encoding the Fe-BB uptake system, is mediated by Btr (formerly YbbB) which is encoded by the immediately upstream gene. Btr contains an AraC-type DNA binding domain fused to a substrate binding protein (SBP) domain related to FeuA, the SBP for Fe-BB uptake. When cells are iron-limited, the Fur-mediated repression of btr is relieved and Btr binds to a conserved direct repeat sequence adjacent to feuA to activate transcription. If BB is present, Btr further activates feuA expression. Btr binds with high affinity to both apo-BB and Fe-BB and the resulting complex displays a significantly increased efficacy as a transcriptional activator relative to Btr alone. Btr can also activate transcription in response to the structurally similar siderophore enterobactin, although genetic analyses indicate that the two siderophores make distinct interactions with the Btr substrate binding domain. Thus, the FeuABC transporter is optimally expressed under conditions of iron starvation, when Fur-mediated repression is relieved, and in the presence of its cognate substrate. PMID:17725565

  1. Interspecies modulation of bacterial development through iron competition and siderophore piracy

    PubMed Central

    Traxler, Matthew F.; Seyedsayamdost, Mohammad R.; Clardy, Jon; Kolter, Roberto

    2012-01-01

    Summary While soil-dwelling actinomycetes are renowned for secreting natural products, little is known about the roles of these molecules in mediating actinomycete interactions. In a previous co-culture screen, we found that one actinomycete, Amycolatopsis sp. AA4, inhibited aerial hyphae formation in adjacent colonies of Streptomyces coelicolor. A siderophore, amychelin, mediated this developmental arrest. Here we present genetic evidence that confirms the role of the amc locus in the production of amychelin and in the inhibition of S. coelicolor development. We further characterize the Amycolatopsis sp. AA4 - S. coelicolor interaction by examining expression of developmental and iron acquisition genes over time in co-culture. Manipulation of iron availability and/or growth near Amycolatopsis sp. AA4 led to alterations in expression of the critical developmental gene bldN, and other key down-stream genes in the S. coelicolor transcriptional cascade. In Amycolatopsis sp. AA4, siderophore genes were down-regulated when grown near S. coelicolor, leading us to find that deferrioxamine E, produced by S. coelicolor, could be readily utilized by Amycolatopsis sp. AA4. Collectively these results suggest that competition for iron via siderophore piracy and species-specific siderophores can alter patterns of gene expression and morphological differentiation during actinomycete interactions. PMID:22931126

  2. Siderophore production by streptomycetes-stability and alteration of ferrihydroxamates in heavy metal-contaminated soil.

    PubMed

    Schütze, Eileen; Ahmed, Engy; Voit, Annekatrin; Klose, Michael; Greyer, Matthias; Svatoš, Aleš; Merten, Dirk; Roth, Martin; Holmström, Sara J M; Kothe, Erika

    2015-12-01

    Heavy metal-contaminated soil derived from a former uranium mining site in Ronneburg, Germany, was used for sterile mesocosms inoculated with the extremely metal-resistant Streptomyces mirabilis P16B-1 or the sensitive control strain Streptomyces lividans TK24. The production and fate of bacterial hydroxamate siderophores in soil was analyzed, and the presence of ferrioxamines E, B, D, and G was shown. While total ferrioxamine concentrations decreased in water-treated controls after 30 days of incubation, the sustained production by the bacteria was seen. For the individual molecules, alteration between neutral and cationic forms and linearization of hydroxamates was observed for the first time. Mesocosms inoculated with biomass of either strain showed changes of siderophore contents compared with the non-treated control indicating for auto-alteration and consumption, respectively, depending on the vital bacteria present. Heat stability and structural consistency of siderophores obtained from sterile culture filtrate were shown. In addition, low recovery (32 %) from soil was shown, indicating adsorption to soil particles or soil organic matter. Fate and behavior of hydroxamate siderophores in metal-contaminated soils may affect soil properties as well as conditions for its inhabiting (micro)organisms. PMID:25414032

  3. Substrate induction of siderophore transport in Bacillus subtilis mediated by a novel one-component regulator.

    PubMed

    Gaballa, Ahmed; Helmann, John D

    2007-10-01

    When iron is scarce, Bacillus subtilis expresses genes involved in the synthesis and uptake of the siderophore bacillibactin (BB) and uptake systems to pirate other microbial siderophores. Here, we demonstrate that transcriptional induction of the feuABCybbA operon, encoding the Fe-BB uptake system, is mediated by Btr (formerly YbbB), which is encoded by the immediately upstream gene. Btr contains an AraC-type DNA binding domain fused to a substrate binding protein (SBP) domain related to FeuA, the SBP for Fe-BB uptake. When cells are iron-limited, the Fur-mediated repression of btr is relieved and Btr binds to a conserved direct repeat sequence adjacent to feuA to activate transcription. If BB is present, Btr further activates feuA expression. Btr binds with high affinity to both apo-BB and Fe-BB, and the resulting complex displays a significantly increased efficacy as a transcriptional activator relative to Btr alone. Btr can also activate transcription in response to the structurally similar siderophore enterobactin, although genetic analyses indicate that the two siderophores make distinct interactions with the Btr substrate binding domain. Thus, the FeuABC transporter is optimally expressed under conditions of iron starvation, when Fur-mediated repression is relieved, and in the presence of its cognate substrate. PMID:17725565

  4. Interspecies modulation of bacterial development through iron competition and siderophore piracy.

    PubMed

    Traxler, Matthew F; Seyedsayamdost, Mohammad R; Clardy, Jon; Kolter, Roberto

    2012-11-01

    While soil-dwelling actinomycetes are renowned for secreting natural products, little is known about the roles of these molecules in mediating actinomycete interactions. In a previous co-culture screen, we found that one actinomycete, Amycolatopsis sp. AA4, inhibited aerial hyphae formation in adjacent colonies of Streptomyces coelicolor. A siderophore, amychelin, mediated this developmental arrest. Here we present genetic evidence that confirms the role of the amc locus in the production of amychelin and in the inhibition of S. coelicolor development. We further characterize the Amycolatopsis sp. AA4 - S. coelicolor interaction by examining expression of developmental and iron acquisition genes over time in co-culture. Manipulation of iron availability and/or growth near Amycolatopsis sp. AA4 led to alterations in expression of the critical developmental gene bldN, and other key downstream genes in the S. coelicolor transcriptional cascade. In Amycolatopsis sp. AA4, siderophore genes were downregulated when grown near S. coelicolor, leading us to find that deferrioxamine E, produced by S. coelicolor, could be readily utilized by Amycolatopsis sp. AA4. Collectively these results suggest that competition for iron via siderophore piracy and species-specific siderophores can alter patterns of gene expression and morphological differentiation during actinomycete interactions. PMID:22931126

  5. Identification and structural characterization of serobactins, a suite of lipopeptide siderophores produced by the grass endophyte Herbaspirillum seropedicae.

    PubMed

    Rosconi, Federico; Davyt, Danilo; Martínez, Verónica; Martínez, Marcela; Abin-Carriquiry, Juan Andrés; Zane, Hannah; Butler, Alison; de Souza, Emanuel M; Fabiano, Elena

    2013-03-01

    Herbaspirillum seropedicae Z67 is a diazotrophic endophyte able to colonize the interior of many economically relevant crops such as rice, wheat, corn and sorghum. Structures of siderophores produced by bacterial endophytes have not yet been elucidated. The aim of this work was to identify and characterize the siderophores produced by this bacterium. In a screening for mutants unable to produce siderophores we found a mutant that had a transposon insertion in a non-ribosomal peptide synthase (NRPS) gene coding for a putative siderophore biosynthetic enzyme. The chemical structure of the siderophore was predicted using computational genomic tools. The predicted structure was confirmed by chemical analysis. We found that siderophores produced by H. seropedicae Z67 are a suite of amphiphilic lipopeptides, named serobactin A, B and C, which vary by the length of the fatty acid chain. We also demonstrated the biological activity of serobactins as nutritional iron sources for H. seropedicae. These are the first structurally described siderophores produced by endophytic bacteria. PMID:23320867

  6. Stereospecificity of the siderophore pyochelin outer membrane transporters in fluorescent pseudomonads.

    PubMed

    Hoegy, Françoise; Lee, Xiaoyun; Noel, Sabrina; Rognan, Didier; Mislin, Gaëtan L A; Reimmann, Cornelia; Schalk, Isabelle J

    2009-05-29

    Pyochelin (Pch) and enantio-pyochelin (EPch) are enantiomer siderophores that are produced by Pseudomonas aeruginosa and Pseudomonas fluorescens, respectively, under iron limitation. Pch promotes growth of P. aeruginosa when iron is scarce, and EPch carries out the same biological function in P. fluorescens. However, the two siderophores are unable to promote growth in the heterologous species, indicating that siderophore-mediated iron uptake is highly stereospecific. In the present work, using binding and iron uptake assays, we found that FptA, the Fe-Pch outer membrane transporter of P. aeruginosa, recognized (K(d) = 2.5 +/- 1.1 nm) and transported Fe-Pch but did not interact with Fe-EPch. Likewise, FetA, the Fe-EPch receptor of P. fluorescens, was specific for Fe-EPch (K(d) = 3.7 +/- 2.1 nm) but did not bind and transport Fe-Pch. Growth promotion experiments performed under iron-limiting conditions confirmed that FptA and FetA are highly specific for Pch and EPch, respectively. When fptA and fetA along with adjacent transport genes involved in siderophore uptake were swapped between the two bacterial species, P. aeruginosa became able to utilize Fe-EPch as an iron source, and P. fluorescens was able to grow with Fe-Pch. Docking experiments using the FptA structure and binding assays showed that the stereospecificity of Pch recognition by FptA was mostly due to the configuration of the siderophore chiral centers C4'' and C2'' and was only weakly dependent on the configuration of the C4' carbon atom. Together, these findings increase our understanding of the stereospecific interaction between Pch and its outer membrane receptor FptA. PMID:19297329

  7. Burkholderia pseudomallei known siderophores and hemin uptake are dispensable for lethal murine melioidosis.

    PubMed

    Kvitko, Brian H; Goodyear, Andrew; Propst, Katie L; Dow, Steven W; Schweizer, Herbert P

    2012-01-01

    Burkholderia pseudomallei is a mostly saprophytic bacterium, but can infect humans where it causes the difficult-to-manage disease melioidosis. Even with proper diagnosis and prompt therapeutic interventions mortality rates still range from >20% in Northern Australia to over 40% in Thailand. Surprisingly little is yet known about how B. pseudomallei infects, invades and survives within its hosts, and virtually nothing is known about the contribution of critical nutrients such as iron to the bacterium's pathogenesis. It was previously assumed that B. pseudomallei used iron-acquisition systems commonly found in other bacteria, for example siderophores. However, our previous discovery of a clinical isolate carrying a large chromosomal deletion missing the entire malleobactin gene cluster encoding the bacterium's major high-affinity siderophore while still being fully virulent in a murine melioidosis model suggested that other iron-acquisition systems might make contributions to virulence. Here, we deleted the major siderophore malleobactin (mba) and pyochelin (pch) gene clusters in strain 1710b and revealed a residual siderophore activity which was unrelated to other known Burkholderia siderophores such as cepabactin and cepaciachelin, and not due to increased secretion of chelators such as citrate. Deletion of the two hemin uptake loci, hmu and hem, showed that Hmu is required for utilization of hemin and hemoglobin and that Hem cannot complement a Hmu deficiency. Prolonged incubation of a hmu hem mutant in hemoglobin-containing minimal medium yielded variants able to utilize hemoglobin and hemin suggesting alternate pathways for utilization of these two host iron sources. Lactoferrin utilization was dependent on malleobactin, but not pyochelin synthesis and/or uptake. A mba pch hmu hem quadruple mutant could use ferritin as an iron source and upon intranasal infection was lethal in an acute murine melioidosis model. These data suggest that B. pseudomallei may employ

  8. Characterization of a Bacillus subtilis transporter for petrobactin, an anthrax stealth siderophore

    SciTech Connect

    Zawadzka, A. M.; Kim, Y.; Maltseva, N; Nichiporuk, R; Fan, Y; Joachimiak, A; Raymond, KN

    2009-12-22

    Iron deprivation activates the expression of components of the siderophore-mediated iron acquisition systems in Bacillus subtilis, including not only the synthesis and uptake of its siderophore bacillibactin but also expression of multiple ABC transporters for iron scavenging using xenosiderophores. The yclNOPQ operon is shown to encode the complete transporter for petrobactin (PB), a photoreactive 3,4-catecholate siderophore produced by many members of the B. cereus group, including B. anthracis. Isogenic disruption mutants in the yclNOPQ transporter, including permease YclN, ATPase YclP, and a substrate-binding protein YclQ, are unable to use either PB or the photoproduct of FePB (FePB{sup {nu}}) for iron delivery and growth, in contrast to the wild-type B. subtilis. Complementation of the mutations with the copies of the respective genes restores this capability. The YclQ receptor binds selectively iron-free and ferric PB, the PB precursor, 3,4-dihydroxybenzoic acid (3,4-DHB), and FePB{sup {nu}} with high affinity; the ferric complexes are seen in ESI-MS, implying strong electrostatic interaction between the protein-binding pocket and siderophore. The first structure of a Gram-positive siderophore receptor is presented. The 1.75-{angstrom} crystal structure of YclQ reveals a bilobal periplasmic binding protein (PBP) fold consisting of two {alpha}/{beta}/{alpha} sandwich domains connected by a long {alpha}-helix with the binding pocket containing conserved positively charged and aromatic residues and large enough to accommodate FePB. Orthologs of the B. subtilis PB-transporter YclNOPQ in PB-producing Bacilli are likely contributors to the pathogenicity of these species and provide a potential target for antibacterial strategies.

  9. Mechanism of Non-Steady State Dissolution of Goethite in the Presence of Siderophores

    NASA Astrophysics Data System (ADS)

    Reichard, P. U.; Kretzschmar, R.; Kraemer, S. M.

    2003-12-01

    Iron is an essential micronutrient for almost all known organisms. Bacteria, fungi, and graminaceous plants are capable of exuding siderophores as part of an iron acquisition strategy. The production of these strong iron chelating ligands is induced by iron limited conditions. Grasses under iron stress, for example, exude phytosiderophores into the rhizosphere in a special diurnal rhythm (Roemheld and Marschner 1986). A few hours after sunrise the exudation starts, culminates around noon and is shut down again until about 4 hours after noon. The phytosiderophores diffuse into the rhizosphere (Marschner et al. 1986) and are passively back transported to the plants by advective flow induced by high transpiration around noon. Despite a fairly short residence time of the phytosiderophores in the rhizosphere, it is a very effective strategy for iron acquisition. To investigate the effect of such pulse inputs of siderophores on iron acquisition, we studied the dissolution mechanism of goethite (alpha-FeOOH), a mineral phase common in soils, under non-steady state conditions. In consideration of the chemical complexity of the rhizosphere, we also investigated the effect of other organic ligands commonly found in the rhizosphere (e. g. oxalate) on the dissolution kinetics. The dissolution experiments were conducted in batch reactors with a constant goethite solids concentration of 2.5 g/l, an ionic strength of 0.01 M, a pH of 6 and 100 microM oxalate. To induce non-steady state conditions, 3 mM phytosiderophores were added to a batch after the goethite-oxalate suspension reacted for a certain time period. Before the siderophore was added to the goethite-oxalate suspension, no dissolution of iron was observed. But, with the addition of the siderophore, a high rate was observed for the iron mobilization under these non-steady state conditions that subsequently was followed by a slow steady state dissolution rate. The results of these non-steady state experiments are very

  10. The Bordetella Bfe System: Growth and Transcriptional Response to Siderophores, Catechols, and Neuroendocrine Catecholamines

    PubMed Central

    Anderson, Mark T.; Armstrong, Sandra K.

    2006-01-01

    Ferric enterobactin utilization by Bordetella bronchiseptica and Bordetella pertussis requires the BfeA outer membrane receptor. Under iron-depleted growth conditions, transcription of bfeA is activated by the BfeR regulator by a mechanism requiring the siderophore enterobactin. In this study, enterobactin-inducible bfeA transcription was shown to be TonB independent. To determine whether other siderophores or nonsiderophore catechols could be utilized by the Bfe system, various compounds were tested for the abilities to promote the growth of iron-starved B. bronchiseptica and induce bfeA transcription. The BfeA receptor transported ferric salmochelin, corynebactin, and the synthetic siderophores TRENCAM and MECAM. Salmochelin and MECAM induced bfeA transcription in iron-starved Bordetella cells, but induction by corynebactin and TRENCAM was minimal. The neuroendocrine catecholamines epinephrine, norepinephrine, and dopamine exhibited a remarkable capacity to induce transcription of bfeA. Norepinephrine treatment of B. bronchiseptica resulted in BfeR-dependent bfeA transcription, elevated BfeA receptor production, and growth stimulation. Pyrocatechol, carbidopa, and isoproterenol were similarly strong inducers of bfeA transcription, whereas tyramine and 3,4-dihydroxymandelic acid demonstrated low inducing activity. The results indicate that the inducer structure requires a catechol group for function and that the ability to induce bfeA transcription does not necessarily correlate with the ability to stimulate bacterial growth. The expanded range of catechol siderophores transported by the BfeA receptor demonstrates the potential versatility of the Bordetella Bfe iron retrieval system. The finding that catecholamine neurotransmitters activate bfeA transcription and promote growth suggests that Bordetella cells can perceive and may benefit from neuroendocrine catecholamines on the respiratory epithelium. PMID:16885441

  11. Effects of the Microbial Siderophore DFO-B on Pb and Cd Speciation in Aqueous Solution

    SciTech Connect

    Mishra, Bhoopesh; Haack, Elizabeth A.; Maurice, Patricia A.; Bunker, Bruce A.

    2009-04-08

    This study investigates the complexation environments of aqueous Pb and Cd in the presence of the trihydroxamate microbial siderophore, desferrioxamine-B (DFO-B) as a function of pH. Complexation of aqueous Pb and Cd with DFO-B was predicted using equilibrium speciation calculation. Synchrotron-based X-ray absorption fine structure (XAFS) spectroscopy at Pb L(III) edge and Cd K edge was used to characterize Pb and Cd-DFO-B complexes at pH values predicted to best represent each of the metal-siderophore complexes. Pb was not found to be complexed measurably by DFO-B at pH 3.0, but was complexed by all three hydroxamate groups to form a totally 'caged' hexadentate structure at pH 7.5-9.0. At the intermediate pH value (pH 4.8), a mixture of Pb-DFOB complexes involving binding of the metal through one and two hydroxamate groups was observed. Cd, on the other hand, remained as hydrated Cd{sup 2+} at pH 5.0, occurred as a mixture of Cd-DFOB and inorganic species at pH 8.0, and was bound by three hydroxamate groups from DFO-B at pH 9.0. Overall, the solution species observed with EXAFS were consistent with those predicted thermodynamically. However, Pb speciation at higher pH values differed from that predicted and suggests that published constants underestimate the binding constant for complexation of Pb with all three hydroxamate groups of the DFO-B ligand. This molecular-level understanding of metal-siderophore solution coordination provides physical evidence for complexes of Pb and Cd with DFO-B, and is an important first step toward understanding processes at the microbial- and/or mineral-water interface in the presence of siderophores.

  12. Breaking a pathogen’s iron will: inhibiting siderophore production as an antimicrobial strategy

    PubMed Central

    Lamb, Audrey L.

    2015-01-01

    The rise of antibiotic resistance is a growing public health crisis. Novel antimicrobials are sought, preferably developing nontraditional chemical scaffolds that do not inhibit standard targets such as cell wall synthesis or the ribosome. Iron scavenging has been proposed as a viable target, because bacterial and fungal pathogens must overcome the nutritional immunity of the host to be virulent. This review highlights the recent work toward exploiting the biosynthetic enzymes of siderophore production for the design of next generation antimicrobials. PMID:25970810

  13. Norepinephrine represses the expression of toxA and the siderophore genes in Pseudomonas aeruginosa.

    PubMed

    Li, Wang; Lyte, Mark; Freestone, Primrose P; Ajmal, Aziba; Colmer-Hamood, Jane A; Hamood, Abdul N

    2009-10-01

    Among the different extracellular virulence factors produced by Pseudomonas aeruginosa are exotoxin A (ETA) and the pyoverdine and pyochelin siderophores. Production of ETA and the siderophores requires the function of the iron-starvation sigma factor PvdS, the transcriptional activator RegA, and the AraC-activator PchR. Iron represses the production of ETA and the siderophores by repressing the expression of pvdS, regA, and pchR. PvdS regulates the expression of the ETA gene, toxA, regA, and the pyoverdine synthesis genes. The catecholamine norepinephrine enhances the growth of pathogenic bacteria by transferring iron from host-binding proteins. In this study, we elucidated the mechanism by which norepinephrine and other catecholamines induce P. aeruginosa growth. We also investigated whether norepinephrine regulates the expression of toxA and the siderophore genes, and the mechanism of this regulation. Norepinephrine enhanced the growth of P. aeruginosa by supplying iron from transferrin. This provision of iron repressed the expression of toxA, the pyoverdine genes pvdD and pvdE, and their regulators, pvdS, regA, and pchR, suggesting that norepinephrine accomplishes this repression through PvdS and PchR. Additionally, norepinephrine bypassed PvdS and supported the growth of a pvdS deletion mutant, indicating that norepinephrine transfers iron to P. aeruginosa independent of pyoverdine. Thus, norepinephrine apparently influences the pathogenesis of P. aeruginosa by affecting its pattern of growth and the production of virulence factors. PMID:19686346

  14. Effects of Injected Iron and Siderophores on Infections in Normal and Immune Mice

    PubMed Central

    Kochan, Ivan; Wasynczuk, John; McCabe, Martha A.

    1978-01-01

    The fate of virulent and avirulent strains of Salmonella typhimurium in untreated and iron-injected mice and in transferrin-containing media demonstrated a direct relationship between bacterial virulence and the ability of bacteria to acquire transferrin-bound iron. Effects of injected iron on the development of infections with virulent and avirulent bacterial strains were determined in normal and immune mice by determinations of bacterial numbers in tissue homogenates and the mortality of infected animals. Results showed that infected and iron-injected mice died much more rapidly and frequently from overwhelming infections than infected and saline-injected mice. The infection-promoting effect of iron varied with the degree of bacterial virulence; the more virulent the bacteria, the more helpful was iron for the development of lethal infections. Siderophores promoted lethal infections in mice infected with virulent but not with avirulent bacteria. Experiments with vaccinated animals showed that iron exerted a deleterious effect on acquired immunity. Immune mice infected with virulent bacteria and injected with iron developed lethal infections as rapidly and nearly as frequently as similarly treated normal mice. Siderophores did not promote the development of lethal infections in immune mice. The effectiveness of iron, but not of siderophores, to promote bacterial infections in vaccinated mice revealed that acquired immunity is dependent upon the activity of an iron-neutralizable antibacterial system. Images PMID:365761

  15. Potential mobilization of platinum-group elements by siderophores in surface environments.

    PubMed

    Dahlheimer, Susan R; Neal, Clive R; Fein, Jeremy B

    2007-02-01

    The emission of platinum-group elements (PGEs) from catalytic converters has led to increased environmental abundances of Pt, Pd, and Rh; however, little is known about the environmental effects and fate of these metals. Organic ligands found in soils have the potential to increase the mobility of PGEs and potentially increase the bioavailability of the metals. Here, we assessed the abilities of microbially produced iron-chelating ligands (siderophores) to complex with the PGEs. Batch experiments using the synthetic siderophore desferrioxamine-B (DFO-B) and powdered metal or oxide forms of Pt, Pd, or Rh showed that DFO-B enhances the solubility of Pt and Pd due to the formation of Pt- and Pd-DFO-B aqueous complexes, with estimated minimum stability constants on the order of 10(17-18) and 10(20-24), respectively. Dissolution rates for Pd are comparable to other mineral dissolution rates with DFO-B. DFO-B had little to no effect on the dissolution of Rh metal or Rh2O3. Our results indicate that siderophores have the potential to increase the mobility of Pt and Pd in environments with limited activities of free trivalent cations. These results have implications for the fate of catalytic converter-emitted Pt and Pd, and support the need for further Pt and Pd toxicity and bioaccumulation studies. PMID:17328196

  16. Adaptation-based resistance to siderophore-conjugated antibacterial agents by Pseudomonas aeruginosa.

    PubMed

    Tomaras, Andrew P; Crandon, Jared L; McPherson, Craig J; Banevicius, Mary Anne; Finegan, Steven M; Irvine, Rebecca L; Brown, Matthew F; O'Donnell, John P; Nicolau, David P

    2013-09-01

    Multidrug resistance in Gram-negative bacteria has become so threatening to human health that new antibacterial platforms are desperately needed to combat these deadly infections. The concept of siderophore conjugation, which facilitates compound uptake across the outer membrane by hijacking bacterial iron acquisition systems, has received significant attention in recent years. While standard in vitro MIC and resistance frequency methods demonstrate that these compounds are potent, broad-spectrum antibacterial agents whose activity should not be threatened by unacceptably high spontaneous resistance rates, recapitulation of these results in animal models can prove unreliable, partially because of the differences in iron availability in these different methods. Here, we describe the characterization of MB-1, a novel siderophore-conjugated monobactam that demonstrates excellent in vitro activity against Pseudomonas aeruginosa when tested using standard assay conditions. Unfortunately, the in vitro findings did not correlate with the in vivo results we obtained, as multiple strains were not effectively treated by MB-1 despite having low MICs. To address this, we also describe the development of new in vitro assays that were predictive of efficacy in mouse models, and we provide evidence that competition with native siderophores could contribute to the recalcitrance of some P. aeruginosa isolates in vivo. PMID:23774440

  17. Bacterial iron transport: coordination properties of azotobactin, the highly fluorescent siderophore of Azotobacter vinelandii.

    PubMed

    Palanché, Tania; Blanc, Sylvie; Hennard, Christophe; Abdallah, Mohamed A; Albrecht-Gary, Anne-Marie

    2004-02-01

    Azotobacter vinelandii, a nitrogen-fixing soil bacterium, secretes in iron deficiency azotobactin delta, a highly fluorescent pyoverdin-like chromopeptidic hexadentate siderophore. The chromophore, derived from 2,3-diamino-6,7 dihydroxyquinoline, is bound to a peptide chain of 10 amino acids: (L)-Asp-(D)-Ser-(L)-Hse-Gly-(D)-beta-threo-HOAsp-(L)-Ser-(D)-Cit-(L)-Hse-(L)-Hse lactone-(D)-N(delta)-Acetyl, N(delta)-HOOrn. Azotobactin delta has three different iron(III) binding sites which are one hydroxamate group at the C-terminal end of the peptidic chain (N(delta)-Acetyl, N(delta)-HOOrn), one alpha-hydroxycarboxylic function in the middle of the chain (beta-threo-hydroxyaspartic acid), and one catechol group on the chromophore. The coordination properties of its iron(III) and iron(II) complexes were measured by spectrophotometry, potentiometry, and voltammetry after the determination of the acid-base functions of the uncomplexed free siderophore. Strongly negatively charged ferric species were observed at neutral p[H]'s corresponding to a predominant absolute configuration Lambda of the ferric complex in solution as deduced from CD measurements. The presence of an alpha-hydroxycarboxylic chelating group does not decrease the stability of the iron(III) complex when compared to the main trishydroxamate siderophores or to pyoverdins. The value of the redox potential of ferric azotobactin is highly consistent with a reductive step by physiological reductants for the iron release. Formation and dissociation kinetics of the azotobactin delta ferric complex point out that both ends of this long siderophore chain get coordinated to Fe(III) before the middle. The most striking result provided by fluorescence measurements is the lasting quenching of the fluorophore in the course of the protonation of the ferric azotobactin delta complex. Despite the release of the hydroxyacid and of the catechol, the fluorescence remains indeed quenched, when iron(III) is bound only to the

  18. Keratan Sulfate Biosynthesis

    PubMed Central

    Funderburgh, James L.

    2010-01-01

    Summary Keratan sulfate was originally identified as the major glycosaminoglycan of cornea but is now known to modify at least a dozen different proteins in a wide variety of tissues. Despite a large body of research documenting keratan sulfate structure, and an increasing interest in the biological functions of keratan sulfate, until recently little was known of the specific enzymes involved in keratan sulfate biosynthesis or of the molecular mechanisms that control keratan sulfate expression. In the last 2 years, however, marked progress has been achieved in identification of genes involved in keratan sulfate biosynthesis and in development of experimental conditions to study keratan sulfate secretion and control in vitro. This review summarizes current understanding of keratan sulfate structure and recent developments in understanding keratan sulfate biosynthesis. PMID:12512857

  19. Rapid identification of siderophores by combined thin-layer chromatography/matrix-assisted laser desorption/ionization mass spectrometry.

    PubMed

    Hayen, Heiko; Volmer, Dietrich A

    2005-01-01

    The investigation of a combined thin-layer chromatography/matrix-assisted laser desorption/ionization mass spectrometry (TLC/MALDI-MS) method for the analysis of siderophores from microbial samples is described. The investigated siderophores were enterobactin, ferrioxamine B, ferrichrome, ferrirhodin, rhodotorulic acid and coprogen. Solid-phase extraction was employed to recover the siderophores from the microbial samples. After visualization of the spots via spraying with ferric chloride or chrome azurol sulfonate assay solution, the MALDI matrix was applied to the gel surface. Several TLC/MALDI experimental parameters were optimized, such as type and concentration of MALDI matrix, as well as the type and composition of solvent to facilitate analyte transport from the inside of the TLC gel to the surface. The impact of these parameters on sensitivity, precision and ion formation of the various siderophores was studied. The detection limits for the investigated siderophores were in the range 1-4 pmol. These values were about 4-24 times higher than the detection limits obtained directly from stainless steel MALDI targets. The differences were most likely due to incomplete transport of the 'trapped' analyte molecules from the deeper layers of the TLC gel to the surface and into the matrix layer. In addition, chromatographic band broadening spread the analyte further in TLC as compared with the steel plates, resulting in less analyte per surface area. The identification of the siderophores was aided by concurrently applying a Ga(III) nitrate solution to the TLC plate during the visualization step. The resulting formation of Ga(III) complexes lead to distinctive (69)Ga/(71)Ga isotope patterns in the mass spectra. The versatility of the TLC/MALDI-MS assay was demonstrated by using it to analyze siderophores in a Pseudomonas aeruginosa sample. An iron-binding compound was identified in the sample, namely pyochelin (2-(2-o-hydroxyphenyl-2-thiazolin-4-yl)-3

  20. Mammalian cardiolipin biosynthesis.

    PubMed

    Mejia, Edgard M; Nguyen, Hieu; Hatch, Grant M

    2014-04-01

    Cardiolipin is a major phospholipid in mitochondria and is involved in the generation of cellular energy in the form of ATP. In mammalian and eukaryotic cells it is synthesized via the cytidine-5'-diphosphate-1,2-diacyl-sn-glycerol phosphate pathway. This brief review will describe some of the more recent studies on mammalian cardiolipin biosynthesis and provide an overview of regulation of cardiolipin biosynthesis. In addition, the important role that this key phospholipid plays in disease processes including heart failure, diabetes, thyroid hormone disease and the genetic disease Barth Syndrome will be discussed. PMID:24144810

  1. Biosynthesis of Polyisoprenoids

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The invention is a process for synthesis of a polymer with the same chemical structure as Natural Rubber (NR) obtained from Hevea brasiliensis and other plant species. The research collaborators recently proposed that NR biosynthesis proceeds via a carbocationic polymerization. Based on this theory...

  2. Iron assimilation and siderophore production by Vibrio ordalii strains isolated from diseased Atlantic salmon Salmo salar in Chile.

    PubMed

    Ruiz, Pamela; Balado, Miguel; Toranzo, Alicia E; Poblete-Morales, Matías; Lemos, Manuel L; Avendaño-Herrera, Ruben

    2016-03-30

    Vibrio ordalii is the causative agent of vibriosis in several cultured salmonid species worldwide. Despite its impact on aquaculture, relatively little information is available about its virulence factors. The present study demonstrates for the first time that V. ordalii possesses different systems of iron acquisition, one involving siderophore synthesis and another one that uses direct binding of heme to use iron. Using 6 strains of V. ordalii from Atlantic salmon Salmo salar and the V. ordalii type strain, we could demonstrate that all strains could grow in presence of the chelating agent 2,2'-dipyridyl and produced siderophores in solid and liquid media. Cross-feeding assays among V. ordalii strains evidenced variability in the siderophores produced. Bioassays and PCR data suggest that V. ordalii could produce a siderophore with a structure similar to piscibactin, although the production of a second siderophore in certain strains cannot be discarded. Furthermore, all strains were able to use hemin and hemoglobin as the only iron sources, although the cell yield was higher when using hemoglobin. A hemin-binding assay indicated the presence of constitutive heme-binding molecules at the cell surface of V. ordalii. Virulence tests using rainbow trout as a model of infection revealed a clear relationship between iron-uptake ability and pathogenicity in V. ordalii. PMID:27025309

  3. In Vitro Cultivation of ‘Unculturable’ Oral Bacteria, Facilitated by Community Culture and Media Supplementation with Siderophores

    PubMed Central

    Vartoukian, Sonia R.; Adamowska, Aleksandra; Lawlor, Megan; Moazzez, Rebecca; Dewhirst, Floyd E.; Wade, William G.

    2016-01-01

    Over a third of oral bacteria are as-yet-uncultivated in-vitro. Siderophores have been previously shown to enable in-vitro growth of previously uncultivated bacteria. The objective of this study was to cultivate novel oral bacteria in siderophore-supplemented culture media. Various compounds with siderophore activity, including pyoverdines-Fe-complex, desferricoprogen and salicylic acid, were found to stimulate the growth of difficult-to-culture strains Prevotella sp. HOT-376 and Fretibacterium fastidiosum. Furthermore, pyrosequencing analysis demonstrated increased proportions of the as-yet-uncultivated phylotypes Dialister sp. HOT-119 and Megasphaera sp. HOT-123 on mixed culture plates supplemented with siderophores. Therefore a culture model was developed, which incorporated 15 μg siderophore (pyoverdines-Fe-complex or desferricoprogen) or 150 μl neat subgingival-plaque suspension into a central well on agar plates that were inoculated with heavily-diluted subgingival-plaque samples from subjects with periodontitis. Colonies showing satellitism were passaged onto fresh plates in co-culture with selected helper strains. Five novel strains, representatives of three previously-uncultivated taxa (Anaerolineae bacterium HOT-439, the first oral taxon from the Chloroflexi phylum to have been cultivated; Bacteroidetes bacterium HOT-365; and Peptostreptococcaceae bacterium HOT-091) were successfully isolated. All novel isolates required helper strains for growth, implying dependence on a biofilm lifestyle. Their characterisation will further our understanding of the human oral microbiome. PMID:26764907

  4. The Legionella pneumophila Siderophore Legiobactin Is a Polycarboxylate That Is Identical in Structure to Rhizoferrin.

    PubMed

    Burnside, Denise M; Wu, Yuyang; Shafaie, Saman; Cianciotto, Nicholas P

    2015-10-01

    Legionella pneumophila, the agent of Legionnaires' disease, secretes a siderophore (legiobactin) that promotes bacterial infection of the lung. In past work, we determined that cytoplasmic LbtA (from Legiobactin gene A) promotes synthesis of legiobactin, inner membrane LbtB aids in export of the siderophore, and outer membrane LbtU and inner membrane LbtC help mediate ferrilegiobactin uptake and assimilation. However, the past studies examined legiobactin contained within bacterial culture supernatants. By utilizing high-pressure liquid chromatography that incorporates hydrophilic interaction-based chemistry, we have now purified legiobactin from supernatants of virulent strain 130b that is suitable for detailed chemical analysis. High-resolution mass spectrometry (MS) revealed that the molecular mass of (protonated) legiobactin is 437.140 Da. On the basis of the results obtained from both MS analysis and various forms of nuclear magnetic resonance, we found that legiobactin is composed of two citric acid residues linked by a putrescine bridge and thus is identical in structure to rhizoferrin, a polycarboxylate-type siderophore made by many fungi and several unrelated bacteria. Both purified legiobactin and rhizoferrin obtained from the fungus Cunninghamella elegans were able to promote Fe(3+) uptake by wild-type L. pneumophila as well as enhance growth of iron-starved bacteria. These results did not occur with 130b mutants lacking lbtU or lbtC, indicating that both endogenously made legiobactin and exogenously derived rhizoferrin are assimilated by L. pneumophila in an LbtU- and LbtC-dependent manner. PMID:26195554

  5. Ferric stability constants of representative marine siderophores: marinobactins, aquachelins, and petrobactin.

    PubMed

    Zhang, Guangping; Amin, Shady A; Küpper, Frithjof C; Holt, Pamela D; Carrano, Carl J; Butler, Alison

    2009-12-01

    The coordination of iron(III) to the marine amphiphilic marinobactin and aquachelin siderophores, as well as to petrobactin, an unusual 3,4-dihydroxybenzoyl siderophore is reported. Potentiometric titrations were performed on the apo siderophore to determine the ligand pK(a) values, as well as the complex formed with addition of 1 equiv of Fe(III). The log K(ML) values for Fe(III)-marinobactin-E and Fe(III)-aquachelin-C are 31.80 and 31.4, respectively, consistent with the similar coordination environment in each complex, while log K(ML) for Fe(III)-petrobactin is estimated to be about 43. The pK(a) of the beta-hydroxyaspartyl hydroxyl group was determined to be 10.8 by (1)H NMR titration. (13)C NMR and IR spectroscopy were used to investigate Ga(III) coordination to the marinobactins. The coordination-induced shifts (CIS) in the (13)C NMR spectrum of Ga(III)-marinobactin-C compared to apo-marinobactin-C indicates that the hydroxamate groups are coordinated to Ga(III); however, the lack of CISs for the carbons of the beta-hydroxyamide group suggests this moiety is not coordinated in the Ga(III) complex. Differences in the IR spectrum of Ga(III)-marinobactin-C and Fe(III)-marinobactin-C in the 1600-1700 cm(-1) region also corroborates Fe(III) is coordinated to the beta-hydroxyamide moiety, whereas Ga(III) is not coordinated. PMID:19902959

  6. Evolutionary dynamics of interlinked public goods traits: an experimental study of siderophore production in Pseudomonas aeruginosa.

    PubMed

    Ross-Gillespie, A; Dumas, Z; Kümmerli, R

    2015-01-01

    Public goods cooperation is common in microbes, and there is much interest in understanding how such traits evolve. Research in recent years has identified several important factors that shape the evolutionary dynamics of such systems, yet few studies have investigated scenarios involving interactions between multiple public goods. Here, we offer general predictions about the evolutionary trajectories of two public goods traits having positive, negative or neutral regulatory influence on one another's expression, and we report on a test of some of our predictions in the context of Pseudomonas aeruginosa's production of two interlinked iron-scavenging siderophores. First, we confirmed that both pyoverdine and pyochelin siderophores do operate as public goods under appropriate environmental conditions. We then tracked their production in lines experimentally evolved under different iron-limitation regimes known to favour different siderophore expression profiles. Under strong iron limitation, where pyoverdine represses pyochelin, we saw a decline in pyoverdine and a concomitant increase in pyochelin - consistent with expansion of pyoverdine-defective cheats derepressed for pyochelin. Under moderate iron limitation, pyochelin declined - again consistent with an expected cheat invasion scenario - but there was no concomitant shift in pyoverdine because cross-suppression between the traits is unidirectional only. Alternating exposure to strong and moderate iron limitation caused qualitatively similar though lesser shifts compared to the constant-environment regimes. Our results confirm that the regulatory interconnections between public goods traits can significantly modulate the course of evolution, yet also suggest how we can start to predict the impacts such complexities will have on phenotypic divergence and community stability. PMID:25421271

  7. The Legionella pneumophila Siderophore Legiobactin Is a Polycarboxylate That Is Identical in Structure to Rhizoferrin

    PubMed Central

    Burnside, Denise M.; Wu, Yuyang; Shafaie, Saman

    2015-01-01

    Legionella pneumophila, the agent of Legionnaires' disease, secretes a siderophore (legiobactin) that promotes bacterial infection of the lung. In past work, we determined that cytoplasmic LbtA (from Legiobactin gene A) promotes synthesis of legiobactin, inner membrane LbtB aids in export of the siderophore, and outer membrane LbtU and inner membrane LbtC help mediate ferrilegiobactin uptake and assimilation. However, the past studies examined legiobactin contained within bacterial culture supernatants. By utilizing high-pressure liquid chromatography that incorporates hydrophilic interaction-based chemistry, we have now purified legiobactin from supernatants of virulent strain 130b that is suitable for detailed chemical analysis. High-resolution mass spectrometry (MS) revealed that the molecular mass of (protonated) legiobactin is 437.140 Da. On the basis of the results obtained from both MS analysis and various forms of nuclear magnetic resonance, we found that legiobactin is composed of two citric acid residues linked by a putrescine bridge and thus is identical in structure to rhizoferrin, a polycarboxylate-type siderophore made by many fungi and several unrelated bacteria. Both purified legiobactin and rhizoferrin obtained from the fungus Cunninghamella elegans were able to promote Fe3+ uptake by wild-type L. pneumophila as well as enhance growth of iron-starved bacteria. These results did not occur with 130b mutants lacking lbtU or lbtC, indicating that both endogenously made legiobactin and exogenously derived rhizoferrin are assimilated by L. pneumophila in an LbtU- and LbtC-dependent manner. PMID:26195554

  8. Multiplicity and specificity of siderophore uptake in the cyanobacterium Anabaena sp. PCC 7120.

    PubMed

    Rudolf, Mareike; Stevanovic, Mara; Kranzler, Chana; Pernil, Rafael; Keren, Nir; Schleiff, Enrico

    2016-09-01

    Many cyanobacteria secrete siderophores to sequester iron. Alternatively, mechanisms to utilize xenosiderophores have evolved. The overall uptake systems are comparable to that of other bacteria involving outer membrane transporters energized by TonB as well as plasma membrane-localized transporters. However, the function of the bioinformatically-inferred components is largely not established and recent studies showed a high diversity of the complexity of the uptake systems in different cyanobacteria. Thus, we approached the systems of the filamentous Anabaena sp. PCC 7120 as a model of a siderophore-secreting cyanobacterium. Anabaena sp. produces schizokinen and uptake of Fe-schizokinen involves the TonB-dependent transporter, schizokinen transporter (SchT), and the ABC-type transport system FhuBCD. We confirm that this system is also relevant for the uptake of structurally similar Fe-siderophore complexes like Fe-aerobactin. Moreover, we demonstrate a function of the TonB-dependent transporter IutA2 in Fe-schizokinen uptake in addition to SchT. The iutA2 mutant shows growth defects upon iron limitation, alterations in Fe-schizokinen uptake and in the transcription profile of the Fe-schizokinen uptake system. The physiological properties of the mutant confirm the importance of iron uptake for cellular function, e.g. for the Krebs cycle. Based on the relative relation of expression of schT and iutA2 as well as of the iron uptake rate to the degree of starvation, a model for the need of the co-existence of two different outer membrane transporters for the same substrate is discussed. PMID:27325117

  9. Stability of YREE complexes with the trihydroxamate siderophore desferrioxamine B at seawater ionic strength

    NASA Astrophysics Data System (ADS)

    Christenson, Emily A.; Schijf, Johan

    2011-11-01

    Organic complexation of yttrium and the rare earth elements (YREEs), although generally believed to be important, is an understudied aspect of YREE solution speciation in the open ocean. We report the first series of stability constants for complexes of YREEs (except Ce and Pm) with the trihydroxamate siderophore desferrioxamine B (DFOB), representing a class of small organic ligands that have an extraordinary selectivity for Fe(III) and are found in surface seawater at low-picomolar concentrations. Constants were measured by potentiometric titration of DFOB (pH 3-10) in the presence of single YREEs, in simple media at seawater ionic strength (NaClO 4 or NaCl, I = 0.7 M). Under these circumstances, the terminal amine of DFOB does not deprotonate. The four acid dissociation constants of the siderophore were determined separately by potentiometric titration of DFOB alone. Values for the bidentate (log β1), tetradentate (log β2), and hexadentate (log β3) complexes of La-Lu range from 4.88 to 6.53, 7.70 to 11.27, and 10.09 to 15.19, respectively, while Y falls between Gd and Tb in each case. Linear free-energy relations of the three stability constants with the first YREE hydrolysis constant, log β1∗, yield regression coefficients of >0.97. On the other hand, plots of the constants vs. the radius of the inner hydration sphere display an increasing deviation from linearity for the lightest REEs (La > Pr > Nd). This may signify steric constraints in DFOB folding around bulkier cations, a larger mismatch in coordination number, or a substantial degree of covalence in the YREE-hydroxamate bond. Complexes of the YREEs with DFOB are many orders of magnitude more stable than those with carbonate, the dominant inorganic YREE ligand in seawater. Speciation modeling with MINEQL indicates that, for an average seawater composition, the hexadentate complex could constitute as much as 28% of dissolved Lu at free DFOB concentrations as low as 10 -13 M. Such conditions might

  10. Synthesis and biological properties of thiazole-analogues of pyochelin, a siderophore of Pseudomonas aeruginosa.

    PubMed

    Noël, Sabrina; Hoegy, Françoise; Rivault, Freddy; Rognan, Didier; Schalk, Isabelle J; Mislin, Gaëtan L A

    2014-01-01

    Pyochelin is a siderophore common to all strains of Pseudomonas aeruginosa utilized by this Gram-negative bacterium to acquire iron(III). FptA is the outer membrane transporter responsible of ferric-pyochelin uptake in P. aeruginosa. We describe in this Letter the synthesis and the biological properties ((55)Fe uptake, binding to FptA) of several thiazole analogues of pyochelin. Among them we report in this Letter the two first pyochelin analogues able to bind FptA without promoting any iron uptake in P. aeruginosa. PMID:24332092

  11. Siderophore-promoted transfer of rare earth elements and iron from volcanic ash into glacial meltwater, river and ocean water

    NASA Astrophysics Data System (ADS)

    Bau, Michael; Tepe, Nathalie; Mohwinkel, Dennis

    2013-02-01

    The rare earth elements (REE) are a group of trace elements that have short marine residence times and that in river, lake and marine surface waters are typically associated with organic and inorganic particles. Explosive volcanic eruptions, such as the 2010 eruptions of Eyjafjallajökull volcano in Iceland, produce volcanic ash particles which can be an important source of iron and other nutrients for aquatic organisms. To become bioavailable, however, this iron needs to be solubilized by complexing agents, such as siderophores. A well-studied example of such a chelator is the biogenic siderophore desferrioxamin-B (DFOB). Based on results from incubation experiments with glacial meltwater-rich river waters from southern Iceland, which are rich in suspended volcanic ash and that had been incubated with and without DFOB, respectively, we here show that siderophores not only enhance the release of iron, but also promote the mobilization of REE from these particles. In the presence of DFOB, partial dissolution of volcanic ash (and presumably other lithic particles) produces a flux of dissolved REE into ambient waters, that is characterized by depletion of the light REE over the middle REE and by selective enrichment of cerium, due to the formation of dissolved Ce(IV)-DFOB complexes. In siderophore-rich environments, this siderophore-bound REE flux has the potential to modify the concentrations and distribution of the dissolved REE and of the isotopic composition of dissolved Nd in glacial meltwaters, river waters and seawater and might be a component of the boundary effects between shelf sediments and seawater, which are assumed to account for the “missing Nd flux” to seawater. Thermodynamic data further suggest that siderophore-promoted element mobilization could also be important for other polyvalent (trace) elements, such as Hf.

  12. The anguibactin biosynthesis and transport genes are encoded in the chromosome of Vibrio harveyi: a possible evolutionary origin for the pJM1 plasmid–encoded system of Vibrio anguillarum?

    PubMed Central

    Naka, Hiroaki; Actis, Luis A; Crosa, Jorge H

    2013-01-01

    Many Vibrio anguillarum serotype O1 strains carry 65-kb pJM1-type plasmids harboring genes involved in siderophore anguibactin biosynthesis and transport. The anguibactin system is an essential factor for V. anguillarum to survive under iron-limiting conditions, and as a consequence, it is a very important virulence factor of this bacterium. Our comparative analysis of genomic data identified a cluster harboring homologs of anguibactin biosynthesis and transport genes in the chromosome of Vibrio harveyi. We have purified the putative anguibactin siderophore and demonstrated that it is indeed anguibactin by mass spectrometry and specific bioassays. Furthermore, we characterized two genes, angR and fatA, in this chromosome cluster that, respectively, participate in anguibactin biosynthesis and transport as determined by mutagenesis analysis. Furthermore, we found that the V. harveyi FatA protein is located in the outer membrane fractions as previously demonstrated in V. anguillarum. Based on our data, we propose that the anguibactin biosynthesis and transport cluster in the V. anguillarum pJM1 plasmid have likely evolved from the chromosome cluster of V. harveyi or vice versa. PMID:23335587

  13. The anguibactin biosynthesis and transport genes are encoded in the chromosome of Vibrio harveyi: a possible evolutionary origin for the pJM1 plasmid-encoded system of Vibrio anguillarum?

    PubMed

    Naka, Hiroaki; Actis, Luis A; Crosa, Jorge H

    2013-02-01

    Many Vibrio anguillarum serotype O1 strains carry 65-kb pJM1-type plasmids harboring genes involved in siderophore anguibactin biosynthesis and transport. The anguibactin system is an essential factor for V. anguillarum to survive under iron-limiting conditions, and as a consequence, it is a very important virulence factor of this bacterium. Our comparative analysis of genomic data identified a cluster harboring homologs of anguibactin biosynthesis and transport genes in the chromosome of Vibrio harveyi. We have purified the putative anguibactin siderophore and demonstrated that it is indeed anguibactin by mass spectrometry and specific bioassays. Furthermore, we characterized two genes, angR and fatA, in this chromosome cluster that, respectively, participate in anguibactin biosynthesis and transport as determined by mutagenesis analysis. Furthermore, we found that the V. harveyi FatA protein is located in the outer membrane fractions as previously demonstrated in V. anguillarum. Based on our data, we propose that the anguibactin biosynthesis and transport cluster in the V. anguillarum pJM1 plasmid have likely evolved from the chromosome cluster of V. harveyi or vice versa. PMID:23335587

  14. Isotope-Assisted Screening for Iron-Containing Metabolites Reveals a High Degree of Diversity among Known and Unknown Siderophores Produced by Trichoderma spp.

    PubMed Central

    Lehner, Sylvia M.; Atanasova, Lea; Neumann, Nora K. N.; Krska, Rudolf; Lemmens, Marc; Druzhinina, Irina S.

    2013-01-01

    Due to low iron availability under environmental conditions, many microorganisms excrete iron-chelating agents (siderophores) to cover their iron demands. A novel screening approach for the detection of siderophores using liquid chromatography coupled to high-resolution tandem mass spectrometry was developed to study the production of extracellular siderophores of 10 wild-type Trichoderma strains. For annotation of siderophores, an in-house library comprising 422 known microbial siderophores was established. After 96 h of cultivation, 18 different iron chelators were detected. Four of those (dimerum acid, fusigen, coprogen, and ferricrocin) were identified by measuring authentic standards. cis-Fusarinine, fusarinine A and B, and des-diserylglycylferrirhodin were annotated based on high-accuracy mass spectral analysis. In total, at least 10 novel iron-containing metabolites of the hydroxamate type were found. On average Trichoderma spp. produced 12 to 14 siderophores, with 6 common to all species tested. The highest number (15) of siderophores was detected for the most common environmental opportunistic and strongly fungicidic species, Trichoderma harzianum, which, however, did not have any unique compounds. The tropical species T. reesei had the most distinctive pattern, producing one unique siderophore (cis-fusarinine) and three others that were present only in T. harzianum and not in other species. The diversity of siderophores did not directly correlate with the antifungal potential of the species tested. Our data suggest that the high diversity of siderophores produced by Trichoderma spp. might be the result of further modifications of the nonribosomal peptide synthetase (NRPS) products and not due to diverse NRPS-encoding genes. PMID:23064341

  15. Role of Catecholate Siderophores in Gram-Negative Bacterial Colonization of the Mouse Gut

    PubMed Central

    Pi, Hualiang; Jones, Shari A.; Mercer, Lynn E.; Meador, Jessica P.; Caughron, Joyce E.; Jordan, Lorne; Newton, Salete M.; Conway, Tyrrell; Klebba, Phillip E.

    2012-01-01

    We investigated the importance of the production of catecholate siderophores, and the utilization of their iron (III) complexes, to colonization of the mouse intestinal tract by Escherichia coli. First, a ΔtonB strain was completely unable to colonize mice. Next, we compared wild type E. coli MG1655 to its derivatives carrying site-directed mutations of genes for enterobactin synthesis (ΔentA::Cm; strain CAT0), ferric catecholate transport (Δfiu, ΔfepA, Δcir, ΔfecA::Cm; CAT4), or both (Δfiu, ΔfepA, ΔfecA, Δcir, ΔentA::Cm; CAT40) during colonization of the mouse gut. Competitions between wild type and mutant strains over a 2-week period in vivo showed impairment of all the genetically engineered bacteria relative to MG1655. CAT0, CAT4 and CAT40 colonized mice 101-, 105-, and 102-fold less efficiently, respectively, than MG1655. Unexpectedly, the additional inability of CAT40 to synthesize enterobactin resulted in a 1000-fold better colonization efficiency relative to CAT4. Analyses of gut mucus showed that CAT4 hyperexcreted enterobactin in vivo, effectively rendering the catecholate transport-deficient strain iron-starved. The results demonstrate that, contrary to prior reports, iron acquisition via catecholate siderophores plays a fundamental role in bacterial colonization of the murine intestinal tract. PMID:23209633

  16. Amonabactin, a novel tryptophan- or phenylalanine-containing phenolate siderophore in Aeromonas hydrophila.

    PubMed Central

    Barghouthi, S; Young, R; Olson, M O; Arceneaux, J E; Clem, L W; Byers, B R

    1989-01-01

    Aeromonas hydrophila 495A2 excreted two forms of amonabactin, a new phenolate siderophore composed of 2,3-dihydroxybenzoic acid, lysine, glycine, and either tryptophan (amonabactin T) or phenylalanine (amonabactin P). Supplementing cultures with L-tryptophan (0.3 mM) caused exclusive synthesis of amonabactin T, whereas supplements of L-phenylalanine (0.3 to 30 mM) gave predominant production of amonabactin P. The two forms of amonabactin were separately purified by a combination of production and polyamide column chromatographic methods. Both forms were biologically active, stimulating growth in iron-deficient medium of an amonabactin-negative mutant. Of 43 additional siderophore-producing isolates of the Aeromonas species that were tested, 76% (19 of 25) of the A. hydrophila isolates were amonabactin positive, whereas only 19% (3 of 16) of the A. sobria isolates and all (3 of 3) of the A. caviae isolates produced amonabactin, suggesting a predominant synthesis of amonabactin in certain Aeromonas species. PMID:2522922

  17. Isolation and characterization of siderophore producing antagonistic rhizobacteria against Rhizoctonia solani.

    PubMed

    Solanki, Manoj Kumar; Singh, Rajesh Kumar; Srivastava, Supriya; Kumar, Sudheer; Kashyap, Prem Lal; Srivastava, Alok K; Arora, Dilip K

    2014-06-01

    Plant protection through siderophore producing rhizobacteria (SPR) has emerged as a sustainable approach for crop health management. In present study, 220 bacteria isolated from tomato rhizosphere were screened for in vitro antagonistic activity against Rhizoctonia solani AG-4. Nine potent antagonistic strains viz., Alcaligenes sp. (MUN1, MB21, and MPF37), Enterobacter sp. (MPM1), Pseudomonas sp. (M10A and MB65), P. aeruginosa (MPF14 and MB123) and P. fluorescens (MPF47) were identified on the basis of physiological characters and 16S rDNA sequencing. These strains were able to produce hydrolytic enzymes, hydrogen cyanide, indole acetic acid, although, only few strains were able to solubilize phosphate. Two strains (MB123 and MPF47) showed significant disease reduction in glasshouse conditions were further evaluated under field conditions using three different application methods. Application of P. fluorescens (MPF47) in nursery as soil mix + seedling root treatments prior to transplantation resulted in significant disease reduction compared to control. Total chlorophyll and available iron were significantly higher in the MPF47 treated plants in contrast to infected control. In conclusion, siderophore producing bacteria MPF47 have strong biocontrol abilities and its application as soil mix + seedling root treatments provided strong shield to plant roots against R. solani and could be used for effective bio-management of pathogen. PMID:23686438

  18. Possible role of bacterial siderophores in inflammation. Iron bound to the Pseudomonas siderophore pyochelin can function as a hydroxyl radical catalyst.

    PubMed Central

    Coffman, T J; Cox, C D; Edeker, B L; Britigan, B E

    1990-01-01

    Tissue injury has been linked to neutrophil associated hydroxyl radical (.OH) generation, a process that requires an exogenous transition metal catalyst such as iron. In vivo most iron is bound in a noncatalytic form. To obtain iron required for growth, many bacteria secrete iron chelators (siderophores). Since Pseudomonas aeruginosa infections are associated with considerable tissue destruction, we examined whether iron bound to the Pseudomonas siderophores pyochelin (PCH) and pyoverdin (PVD) could act as .OH catalysts. Purified PCH and PVD were iron loaded (Fe-PCH, Fe-PVD) and added to a hypoxanthine/xanthine oxidase superoxide- (.O2-) and hydrogen peroxide (H2O2)-generating system. Evidence for .OH generation was then sought using two different spin-trapping agents (5.5 dimethyl-pyrroline-1-oxide or N-t-butyl-alpha-phenylnitrone), as well as the deoxyribose oxidation assay. Regardless of methodology, .OH generation was detected in the presence of Fe-PCH but not Fe-PVD. Inhibition of the process by catalase and/or SOD suggested .OH formation with Fe-PCH occurred via the Haber-Weiss reaction. Similar results were obtained when stimulated neutrophils were used as the source of .O2- and H2O2. Addition of Fe-PCH but not Fe-PVD to stimulated neutrophils yielded .OH as detected by the above assay systems. Since PCH and PVD bind ferric (Fe3+) but not ferrous (Fe2+) iron, .OH catalysis with Fe-PCH would likely involve .O2(-)-mediated reduction of Fe3+ to Fe2+ with subsequent release of "free" Fe2+. This was confirmed by measuring formation of the Fe2(+)-ferrozine complex after exposure of Fe-PCH, but not Fe-PVD, to enzymatically generated .O2-. These data show that Fe-PCH, but not Fe-PVD, is capable of catalyzing generation of .OH. Such a process could represent as yet another mechanism of tissue injury at sites of infection with P. aeruginosa. PMID:2170442

  19. THE APPLICATION OF SIDEROPHORES FOR METAL RECOVERY AND WASTE REMEDIATION: EXAMINATION OF CORRELATIONS FOR PREDICTION OF METAL AFFINITIES

    EPA Science Inventory

    The naturally occurring metal-chelating compounds known as siderophores may be useful in environmental applications but limited metal specificity data is available for this class of compounds. Correlations that predict ligand-metal affinity versus mtal ion charge density and hyd...

  20. Pseudomonas fluorescens CHA0 produces enantio-pyochelin, the optical antipode of the Pseudomonas aeruginosa siderophore pyochelin.

    PubMed

    Youard, Zeb A; Mislin, Gaëtan L A; Majcherczyk, Paul A; Schalk, Isabelle J; Reimmann, Cornelia

    2007-12-01

    The siderophore pyochelin is made by a thiotemplate mechanism from salicylate and two molecules of cysteine. 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, thus determining the stereochemistry of the two interconvertible pyochelin diastereoisomers as 4'R, 2''R, 4''R (pyochelin I) and 4'R, 2''S, 4''R (pyochelin II). Pseudomonas fluorescens CHA0 was found to make a different stereoisomeric mixture, which promoted growth under iron limitation in strain CHA0 and induced the expression of its biosynthetic genes, but was not recognized as a siderophore and signaling molecule by P. aeruginosa. Reciprocally, pyochelin promoted growth and induced pyochelin gene expression in P. aeruginosa, but was not functional in P. fluorescens. The structure of the CHA0 siderophore was determined by mass spectrometry, thin-layer chromatography, NMR, polarimetry, and chiral HPLC as enantio-pyochelin, the optical antipode of the P. aeruginosa siderophore pyochelin. Enantio-pyochelin was chemically synthesized and confirmed to be active in CHA0. Its potential biosynthetic pathway in CHA0 is discussed. PMID:17938167

  1. n-Alkylboronic acid inhibitors reveal determinants of ligand specificity in the quorum-quenching and siderophore biosynthetic enzyme PvdQ.

    PubMed

    Clevenger, Kenneth D; Wu, Rui; Liu, Dali; Fast, Walter

    2014-10-28

    The enzyme PvdQ (E.C. 3.5.1.97) from Pseudomonas aeruginosa is an N-terminal nucleophile hydrolase that catalyzes the removal of an N-myristyl substituent from a biosynthetic precursor of the iron-chelating siderophore pyoverdine. Inhibitors of pyoverdine biosynthesis are potential antibiotics since iron is essential for growth and scarce in most infections. PvdQ also catalyzes hydrolytic amide bond cleavage of selected N-acyl-l-homoserine lactone quorum-sensing signals used by some Gram-negative pathogens to coordinate the transcription of virulence factors. The resulting quorum-quenching activity of PvdQ has potential applications in antivirulence therapies. To inform both inhibitor design and enzyme engineering efforts, a series of n-alkylboronic acid inhibitors of PvdQ was characterized to reveal determinants of ligand selectivity. A simple homologation series results in compounds with Ki values that span from 4.7 mM to 190 pM, with a dependence of ΔGbind values on chain length of -1.0 kcal/mol/CH2. X-ray crystal structures are determined for the PvdQ complexes with 1-ethyl-, 1-butyl-, 1-hexyl-, and 1-octylboronic acids at 1.6, 1.8, 2.0, and 2.1 Å resolution, respectively. The 1-hexyl- and 1-octylboronic acids form tetrahedral adducts with the active-site N-terminal Ser217 in the β-subunit of PvdQ, and the n-alkyl substituents are bound in the acyl-group binding site. The 1-ethyl- and 1-butylboronic acids also form adducts with Ser217 but instead form trigonal planar adducts and extend their n-alkyl substituents into an alternative binding site. These results are interpreted to propose a ligand discrimination model for PvdQ that informs the development of PvdQ-related tools and therapeutics. PMID:25290020

  2. Catecholate-siderophore produced by As-resistant bacterium effectively dissolved FeAsO4 and promoted Pteris vittata growth.

    PubMed

    Liu, Xue; Yang, Guang-Mei; Guan, Dong-Xing; Ghosh, Piyasa; Ma, Lena Q

    2015-11-01

    The impact of siderophore produced by arsenic-resistant bacterium Pseudomonas PG12 on FeAsO4 dissolution and plant growth were examined. Arsenic-hyperaccumulator Pteris vittata was grown for 7 d in 0.2-strength Fe-free Hoagland solution containing FeAsO4 mineral and PG12-siderophore or fungal-siderophore desferrioxamine B (DFOB). Standard siderophore assays indicated that PG12-siderophore was catecholate-type. PG12-siderophore was more effective in promoting FeAsO4 dissolution, and Fe and As plant uptake than DFOB. Media soluble Fe and As in PG12 treatment were 34.6 and 3.07 μM, 1.6- and 1.4-fold of that in DFOB. Plant Fe content increased from 2.93 to 6.24 g kg(-1) in the roots and As content increased from 14.3 to 78.5 mg kg(-1) in the fronds. Besides, P. vittata in PG12 treatment showed 2.6-times greater biomass than DFOB. While P. vittata fronds in PG12 treatment were dominated by AsIII, those in DFOB treatment were dominated by AsV (61-77%). This study showed that siderophore-producing arsenic-resistant rhizobacteria may have potential in enhancing phytoremediation of arsenic-contaminated soils. PMID:26247380

  3. Evaluation of Stress-Induced Microbial Siderophore from Pseudomonas aeruginosa Strain S1 as a Potential Matrix Metalloproteinase Inhibitor in Wound Healing Applications.

    PubMed

    Thyagarajan, Sita Lakshmi; Kandhasamy, S; Ramanathan, Giriprasath; Sivagnanam, Uma Tiruchirapalli; Perumal, P T

    2016-05-01

    Matrix metalloproteinases (MMPs) are zinc-dependent proteolytic enzymes capable of causing various inflammatory and various degenerative diseases if over-expressed. The active site of these enzymes is a zinc binding motif which binds to the specific site on the substrate and induce degradation. Hence an inhibitor is required to form a complex with zinc motif which hampers the binding ability of MMPs. To obtain novel MMPs inhibitor for wound healing, the chelating activity of siderophore from the microbial source was focused. During screening for siderophore production, strain S1 produced the highest amount of siderophore in the minimal salts medium. The isolate was confirmed as Pseudomonas aeruginosa strain S1 based on 16S rRNA gene sequencing and phylogenetic analysis. The activity of the siderophore was assayed using chrome azurol sulphonate and purified by the chromatographic techniques. The structural evidence through Fourier transform infrared and nuclear magnetic resonance spectra revealed that the isolated siderophore is a catecholate type with the distinctive characters. The positive results of calcein and fluozin-3 assays indicate that siderophore could bind to divalent metal ions, namely Fe(2+) and Zn(2+). As the siderophore compound focused on wound healing property, the in vitro studies revealed the viability of NH3T3 fibroblast cells and its efficiency in matrix modulating was confirmed through gelatin zymogram. PMID:26804794

  4. The DNA gyrase inhibitors, nalidixic acid and oxolinic acid, prevent iron-mediated repression of catechol siderophore synthesis in Azotobacter vinelandii.

    PubMed

    Page, W J; Patrick, J

    1988-01-01

    Low concentrations of nalidixic acid and oxolinic acid that were just inhibitory to Azotobacter vinelandii growth promoted the production of the catechol siderophores azotochelin and aminochelin, in the presence of normally repressive concentrations of Fe3+. There was a limited effect on the pyoverdin siderophore, azotobactin, where low concentrations of Fe3+ were rendered less repressive, but the repression by higher concentrations of Fe3+ was normal. These drugs did not induce high-molecular-mass iron-repressible outer-membrane proteins and similar effects on the regulation of catechol siderophore synthesis were not produced by novobiocin, coumermycin, or ethidium bromide. The timing of nalidixic acid and Fe3+ addition to iron-limited cells was critical. Nalidixic acid had to be added before iron-repression of catechol siderophore synthesis and before the onset of iron-sufficient growth. Continued production of the catechol siderophores, however, was not due to interference with normal iron uptake. These data indicated that nalidixic acid prevented normal iron-repression of catechol siderophore synthesis but could not reverse iron repression once it had occurred. The possible roles of DNA gyrase activity in the regulation of catechol siderophore synthesis is discussed. PMID:2856355

  5. Carnitine biosynthesis in mammals.

    PubMed Central

    Vaz, Frédéric M; Wanders, Ronald J A

    2002-01-01

    Carnitine is indispensable for energy metabolism, since it enables activated fatty acids to enter the mitochondria, where they are broken down via beta-oxidation. Carnitine is probably present in all animal species, and in numerous micro-organisms and plants. In mammals, carnitine homoeostasis is maintained by endogenous synthesis, absorption from dietary sources and efficient tubular reabsorption by the kidney. This review aims to cover the current knowledge of the enzymological, molecular, metabolic and regulatory aspects of mammalian carnitine biosynthesis, with an emphasis on the human and rat. PMID:11802770

  6. Increased iron-stress resilience of maize through inoculation of siderophore-producing Arthrobacter globiformis from mine.

    PubMed

    Sharma, Meenakshi; Mishra, Vandana; Rau, Nupur; Sharma, Radhey Shyam

    2016-07-01

    Iron deficiency is common among graminaceous crops. Ecologically successful wild grasses from iron-limiting habitats are likely to harbour bacteria which secrete efficient high-affinity iron-chelating molecules (siderophores) to solubilize and mobilize iron. Such siderophore-producing rhizobacteria may increase the iron-stress resilience of graminaceous crops. Considering this, 51 rhizobacterial isolates of Dichanthium annulatum from iron-limiting abandoned mine (∼84% biologically unavailable iron) were purified and tested for siderophore production; and efficacy of Arthrobacter globiformis inoculation to increase iron-stress resilience of maize and wheat was also evaluated. 16S rRNA sequence analyses demonstrated that siderophore-producing bacteria were taxonomically diverse (seven genera, nineteen species). Among these, Gram-positive Bacillus (eleven species) was prevalent (76.92%). A. globiformis, a commonly found rhizobacterium of graminaceous crops was investigated in detail. Its siderophore has high iron-chelation capacity (ICC: 13.0 ± 2.4 μM) and effectively dissolutes diverse iron-complexes (FeCl3 : 256.13 ± 26.56 μM/ml; Fe2 O3 red: 84.3 ± 4.74 μM/ml; mine spoil: 123.84 ± 4.38 μM/ml). Siderophore production (ICC) of A. globiformis BGDa404 also varied with supplementation of different iron complexes. In plant bioassay with iron-deficiency sensitive species maize, A. globiformis inoculation triggered stress-associated traits (peroxidase and proline) in roots, enhanced plant biomass, uptake of iron and phosphate, and protein and chlorophyll contents. However, in iron deficiency tolerant species wheat, growth improvement was marginal. The present study illustrates: (i) rhizosphere of D. annulatum colonizing abandoned mine as a "hotspot" of siderophore-producing bacteria; and (ii) potential of A. globiformis BGDa404 inoculation to increase iron-stress resilience in maize. A. globiformis BGDa404 has the potential to develop as

  7. Inner-membrane transporters for the siderophores pyochelin in Pseudomonas aeruginosa and enantio-pyochelin in Pseudomonas fluorescens display different enantioselectivities.

    PubMed

    Reimmann, Cornelia

    2012-05-01

    Iron uptake and transcriptional regulation by the enantiomeric siderophores pyochelin (Pch) and enantio-pyochelin (EPch) of Pseudomonas aeruginosa and Pseudomonas fluorescens, respectively, are stereospecific processes. The iron-loaded forms of Pch (ferriPch) and of EPch (ferriEPch) are recognized stereospecifically (i) at the outer membrane by the siderophore receptors FptA in P. aeruginosa and FetA in P. fluorescens and (ii) in the cytoplasm by the two AraC-type regulators PchR, which are activated by their cognate siderophore. Here, stereospecific siderophore recognition is shown to occur at the inner membrane also. In P. aeruginosa, translocation of ferriPch across the inner membrane is carried out by the single-subunit siderophore transporter FptX. In contrast, the uptake of ferriEPch into the cytoplasm of P. fluorescens was found to involve a classical periplasmic binding protein-dependent ABC transporter (FetCDE), which is encoded by the fetABCDEF operon. Expression of a translational fetA-gfp fusion was repressed by ferric ions, and activated by the cognate siderophore bound to PchR, thus resembling the analogous regulation of the P. aeruginosa ferriPch transport operon fptABCX. The inner-membrane transporters FetCDE and FptX were expressed in combination with either of the two siderophore receptors FetA and FptA in a siderophore-negative P. aeruginosa mutant deleted for the fptABCX operon. Growth tests conducted under iron limitation with ferriPch or ferriEPch as the iron source revealed that FptX was able to transport ferriPch as well as ferriEPch, whereas FetCDE specifically transported ferriEPch. Thus, stereospecific siderophore recognition occurs at the inner membrane by the FetCDE transporter. PMID:22343350

  8. Acquisition of Fe from Natural Organic Matter by an Aerobic Pseudomonas Bacterium: Siderophores and Cellular Fe Status

    NASA Astrophysics Data System (ADS)

    Koehn, K.; Dehner, C.; Dubois, J.; Maurice, P. A.

    2010-12-01

    Aerobic microorganisms have evolved various strategies to acquire nutrient Fe, including release of Fe-chelating siderophores. The potential importance of siderophores in Fe acquisition from natural organic matter (NOM) (reverse osmosis, RO; and XAD-8 samples with naturally associated Fe) was investigated using a wild type strain (WT) of aerobic Pseudomonas mendocina that produces siderophore(s) and an engineered mutant that cannot. Microbial growth under Fe-limited batch conditions was monitored via optical density, and a β-galactosidase biosensor assay was used to quantify cellular Fe status. Both WT and mutant strains acquired Fe from NOM. Fe ‘stress’ in the presence of the RO sample decreased with increasing [Fe] (as determined by different [DOC]s) and was consistently less for the WT. For both WT and mutant, maximum growth in the presence of RO sample increased as: 1 mgC/L (0.2μM Fe) < 100 mgC/L (20μM Fe) < 10 mgC/L (2μM Fe). Comparison of XAD-8 and RO samples ([DOC] varied to give 2μM [Fe]total for each), showed that although there were no apparent differences in internal Fe status, growth was better on the XAD-8 sample. Chelex treatment to partially remove metals associated with the RO sample increased Fe stress but did not substantially affect growth. Results demonstrated that: (1) siderophores are useful but not necessary for Fe acquisition from NOM by P. mendocina and (2) NOM may have complex effects on microbial growth, related not just to Fe content but potentially to the presence of other (trace)metals such as Al and/or to effects on biofilm development.

  9. Siderophores, the answer for micro to nanosized asbestos fibre related health hazard

    NASA Astrophysics Data System (ADS)

    Bhattacharya, Shabori; Ledwani, Lalita; John, P. J.

    2016-04-01

    Recent studies on the potential toxicity of High Aspect Ratio Nanoparticles (HARN) has yet once again reinforced the health hazard imposed by asbestos fibres ranging from nano to micro size. Asbestos a naturally occurring fibrous mineral declared a Group I definite carcinogen by IARC (International Agency for Research on Cancer), a unit of WHO in the year 1987, has been extensively used since World War II to the near past for various commercial products. According to the most recent World Health Organization (WHO) estimates, asbestos-related diseases, resulting from exposure at workplace claims more than 107000 lives every year worldwide. The various types of toxic effects induced by asbestos in humans include - i) inflammation and fibrogenesis of lung, ii) mesothelioma iii) asbestosis and iv) bronchogenic carcinoma. The stability of asbestos in natural environment and its biological aggressiveness is related to their fibrous structure and dimensions. The actual risk associated with the exposure to nanosized asbestos, which is still unknown and escapes most regulations worldwide, has been shown in various toxicity assessment studies conducted on various animal models.In an effort to reduce the size of asbestos and therby its toxicity by limiting its biopersistence, oxalic acid treatment of asbestos coupled to power ultrasound treatment was carried out. The nanosized particles formed were still found to retain their hazardous effect. Similar were the results obtained on strong acid treatment of asbestos as well. A probable solution to the asbestos toxicity problem therefore envisaged was bioremediation. This involved the secretion of iron chelating molecules termed siderophores by microbes, which are of significance due to their ability to form very stable and soluble complexes with iron. Iron in asbestos composition is a major factor responsible for its carcinogenicity, removal or extraction of which would prove to be an effective answer to the worldwide problem

  10. Siderophore-Mediated Iron Dissolution from Nontronites Is Controlled by Mineral Cristallochemistry.

    PubMed

    Parrello, Damien; Zegeye, Asfaw; Mustin, Christian; Billard, Patrick

    2016-01-01

    Bacteria living in oxic environments experience iron deficiency due to limited solubility and slow dissolution kinetics of iron-bearing minerals. To cope with iron deprivation, aerobic bacteria have evolved various strategies, including release of siderophores or other organic acids that scavenge external Fe(III) and deliver it to the cells. This research investigated the role of siderophores produced by Pseudomonas aeruginosa in the acquisition of Fe(III) from two iron-bearing colloidal nontronites (NAu-1 and NAu-2), comparing differences in bioavailability related with site occupancy and distribution of Fe(III) in the two lattices. To avoid both the direct contact of the mineral colloids with the bacterial cells and the uncontrolled particle aggregation, nontronite suspensions were homogenously dispersed in a porous silica gel before the dissolution experiments. A multiparametric approach coupling UV-vis spectroscopy and spectral decomposition algorithm was implemented to monitor simultaneously the solubilisation of Fe and the production of pyoverdine in microplate-based batch experiments. Both nontronites released Fe in a particle concentration-dependent manner when incubated with the wild-type P. aeruginosa strain, however iron released from NAu-2 was substantially greater than from NAu-1. The profile of organic acids produced in both cases was similar and may not account for the difference in the iron dissolution efficiency. In contrast, a pyoverdine-deficient mutant was unable to mobilize Fe(III) from either nontronite, whereas iron dissolution occurred in abiotic experiments conducted with purified pyoverdine. Overall, our data provide evidence that P. aeruginosa indirectly mobilize Fe from nontronites primarily through the production of pyoverdine. The structural Fe present on the edges of NAu-2 rather than NAu-1 particles appears to be more bio-accessible, indicating that the distribution of Fe, in the tetrahedron and/or in the octahedron sites, governs

  11. Siderophore-Mediated Iron Dissolution from Nontronites Is Controlled by Mineral Cristallochemistry

    PubMed Central

    Parrello, Damien; Zegeye, Asfaw; Mustin, Christian; Billard, Patrick

    2016-01-01

    Bacteria living in oxic environments experience iron deficiency due to limited solubility and slow dissolution kinetics of iron-bearing minerals. To cope with iron deprivation, aerobic bacteria have evolved various strategies, including release of siderophores or other organic acids that scavenge external Fe(III) and deliver it to the cells. This research investigated the role of siderophores produced by Pseudomonas aeruginosa in the acquisition of Fe(III) from two iron-bearing colloidal nontronites (NAu-1 and NAu-2), comparing differences in bioavailability related with site occupancy and distribution of Fe(III) in the two lattices. To avoid both the direct contact of the mineral colloids with the bacterial cells and the uncontrolled particle aggregation, nontronite suspensions were homogenously dispersed in a porous silica gel before the dissolution experiments. A multiparametric approach coupling UV-vis spectroscopy and spectral decomposition algorithm was implemented to monitor simultaneously the solubilisation of Fe and the production of pyoverdine in microplate-based batch experiments. Both nontronites released Fe in a particle concentration-dependent manner when incubated with the wild-type P. aeruginosa strain, however iron released from NAu-2 was substantially greater than from NAu-1. The profile of organic acids produced in both cases was similar and may not account for the difference in the iron dissolution efficiency. In contrast, a pyoverdine-deficient mutant was unable to mobilize Fe(III) from either nontronite, whereas iron dissolution occurred in abiotic experiments conducted with purified pyoverdine. Overall, our data provide evidence that P. aeruginosa indirectly mobilize Fe from nontronites primarily through the production of pyoverdine. The structural Fe present on the edges of NAu-2 rather than NAu-1 particles appears to be more bio-accessible, indicating that the distribution of Fe, in the tetrahedron and/or in the octahedron sites, governs

  12. Terpene Biosynthesis: Modularity Rules

    PubMed Central

    Oldfield, Eric; Lin, Fu-Yang

    2013-01-01

    Terpenes are the largest class of small molecule natural products on Earth, and the most abundant by mass. Here, we summarize recent developments in elucidating the structure and function of the proteins involved in their biosynthesis. There are 6 main building blocks or modules (α,β,γ,δ,ε and ζ) that make up the structures of these enzymes: the αα and αδ head-to-tail trans-prenyl transferases that produce trans-isoprenoid diphosphates from C5 precursors; the ε head-to-head prenyl transferases that convert these diphosphates into the tri-and tetra-terpene precursors of sterols, hopanoids and carotenoids; the βγ di- and tri-terpene synthases; the ζ head-to-tail cis-prenyl transferases that produce the cis-isoprenoid diphosphates involved in bacterial cell wall biosynthesis, and finally the α, αβ and αβγ terpene synthases that produce plant terpenes, with many of these modular enzymes having originated from ancestral α and β domain proteins. We also review progress in determining the structure and function of the two 4Fe-4S reductases involved in formation of the C5 diphosphates in many bacteria, where again, highly modular structures are found. PMID:22105807

  13. Taxol biosynthesis: an update.

    PubMed

    Hezari, M; Croteau, R

    1997-08-01

    The novel diterpenoid taxol (paclitaxel) is now well-established as a potent chemotherapeutic agent. Total synthesis of the drug is not commercially feasible and, in the foreseeable future, the supply of taxol and its synthetically useful progenitors must rely on biological methods of production. The first three steps of taxol biosynthesis have been defined and the responsible enzymes described. These are the cyclization of the universal diterpenoid precursor geranylgeranyl diphosphate to taxa-4(5),11(12)-diene, the cytochrome P450-catalyzed hydroxylation of this olefin to taxa-4(20), 11(12)-dien-5 alpha-ol, and the acetyl CoA-dependent conversion of the alcohol to the corresponding acetate ester. Demonstration of these early steps of taxol biosynthesis suggests that the complete pathway can be defined by a systematic, stepwise approach at the cell-free enzyme level. When combined with in vivo studies to determine contribution to pathway flux, slow steps can be targeted for gene isolation and subsequent overexpression in Taxus to improve the yield of taxol and related compounds. PMID:9270370

  14. Proteomics of Pseudomonas aeruginosa Australian epidemic strain 1 (AES-1) cultured under conditions mimicking the cystic fibrosis lung reveals increased iron acquisition via the siderophore pyochelin.

    PubMed

    Hare, Nathan J; Soe, Cho Zin; Rose, Barbara; Harbour, Colin; Codd, Rachel; Manos, Jim; Cordwell, Stuart J

    2012-02-01

    Pseudomonas aeruginosa is an opportunistic pathogen that is the major cause of morbidity and mortality in patients with cystic fibrosis (CF). While most CF patients are thought to acquire P. aeruginosa from the environment, person-to-person transmissible strains have been identified in CF clinics worldwide, and the molecular basis for transmissibility remains poorly understood. We undertook a complementary proteomics approach to characterize protein profiles from a transmissible, acute isolate of the Australian epidemic strain 1 (AES-1R), the virulent burns/wound isolate PA14, and the poorly virulent, laboratory-associated strain PAO1 when grown in an artificial medium that mimics the CF lung environment compared to growth in standard laboratory medium. Proteins elevated in abundance in AES-1R included those involved in methionine and S-adenosylmethionine biosynthesis and in the synthesis of phenazines. Proteomic data were validated by measuring culture supernatant levels of the virulence factor pyocyanin, which is the final product of the phenazine pathway. AES-1R and PAO1 released higher extracellular levels of pyocyanin compared to PA14 when grown in conditions that mimic the CF lung. Proteins associated with biosynthesis of the iron-scavenging siderophore pyochelin (PchDEFGH and FptA) were also present at elevated abundance in AES-1R and at much higher levels than in PAO1, whereas they were reduced in PA14. These protein changes resulted phenotypically in increased extracellular iron acquisition potential and, specifically, elevated pyochelin levels in AES-1R culture supernatants as detected by chrome azurol-S assay and fluorometry, respectively. Transcript analysis of pyochelin genes (pchDFG and fptA) showed they were highly expressed during the early stage of growth in artificial sputum medium (18 h) but returned to basal levels following the establishment of microcolony growth (72 h) consistent with that observed in the CF lung. This provides further

  15. Tributyltin chloride (TBTCl)-enhanced exopolysaccharide and siderophore production in an estuarine Alcaligenes faecalis strain.

    PubMed

    Khanolkar, Dnyanada; Dubey, S K; Naik, Milind Mohan

    2015-05-01

    Tributyltin chloride (TBTCl) has been used extensively as an antifouling agent in ship paints, which results in the contamination of aquatic sites. These contaminated sites serve as enrichment areas for TBTCl-resistant bacterial strains. One TBTCl-resistant bacterial strain was isolated from the sediments of Zuari estuary, Goa, India, which is a major hub of various ship-building activities. Based on biochemical characteristics and 16S rDNA sequence analysis, this bacterial strain was identified as Alcaligenes faecalis and designated as strain SD5. It could degrade ≥3 mM TBTCl by using it as a sole carbon source and transform it into the less toxic dibutyltin chloride, which was confirmed by nuclear magnetic resonance and mass spectroscopy. Interestingly, this bacterial strain also showed enhanced exopolysaccharide and siderophore production when cells were exposed to toxic levels of TBTCl, suggesting their involvement in conferring resistance to this antifouling biocide as well as degradative capability respectively. PMID:25612551

  16. Mitophagy confers resistance to siderophore-mediated killing by Pseudomonas aeruginosa.

    PubMed

    Kirienko, Natalia V; Ausubel, Frederick M; Ruvkun, Gary

    2015-02-10

    In the arms race of bacterial pathogenesis, bacteria produce an array of toxins and virulence factors that disrupt core host processes. Hosts mitigate the ensuing damage by responding with immune countermeasures. The iron-binding siderophore pyoverdin is a key virulence mediator of the human pathogen Pseudomonas aeruginosa, but its pathogenic mechanism has not been established. Here we demonstrate that pyoverdin enters Caenorhabditis elegans and that it is sufficient to mediate host killing. Moreover, we show that iron chelation disrupts mitochondrial homeostasis and triggers mitophagy both in C. elegans and mammalian cells. Finally, we show that mitophagy provides protection both against the extracellular pathogen P. aeruginosa and to treatment with a xenobiotic chelator, phenanthroline, in C. elegans. Although autophagic machinery has been shown to target intracellular bacteria for degradation (a process known as xenophagy), our report establishes a role for authentic mitochondrial autophagy in the innate immune defense against P. aeruginosa. PMID:25624506

  17. Siderophore Promoted Dissolution of a Series of Mn-Substituted Goethites

    NASA Astrophysics Data System (ADS)

    Holmstrom, S. J.; Sposito, G.

    2005-12-01

    The presence of organic ligands, like siderophores, can strongly influence mineral dissolution. Recent research suggests that at least some siderophores enhance mineral dissolution by formation of surface complexes with Fe and Mn. The impact of biogeochemical weathering caused by exudates of plants, fungi and bacteria containing siderophores has been discussed. We have studied the dissolution kinetics of Mn-substituted goethites (mol % Mn < 11) in the presences of 80 μM desferrioxamine B (DFO-B), a common and well-studied hydroxamate siderophore that has been identified in both terrestrial and marine environments and which forms very stable 1:1 complexes with Fe(III) or Mn(III). (The stability constants at I = 0.1 are 1030.6 and 1028.3, respectively.) A series of Mn-substituted goethites (α-MnxFe1-xOOH) were synthesized from ferrihydrite in the presence of Mn(II) in alkaline media. The Fe(III) in octahedral positions in the mineral structure was partially replaced by Mn, which was confirmed visually by the change to darker color when the content of Mn increased and proved by infra-red spectroscopy and X-ray diffraction studies of the samples. Substitution of Fe in the goethite by Mn caused a change in the cell dimensions. The calculated unit cell edge lengths a and c decreased, while b increased, for the Mn-goethites compared to pure goethite. The difference of the unit cell parameters between the pure goethite and the Mn-substituted goethites increased with increased Mn content, providing further confirmation that Fe had been substituted by Mn incorporated into the goethite structure. X-ray absorption near-edge structure spectroscopy analysis of the Mn-substituted goethites showed that the oxidation state of Mn in the samples was, as expected, Mn(III), even when Mn-goethites were prepared from Mn(II) solutions. Both SEM and TEM micrographs showed that the Mn-substituted goethite crystals had the same acicular shape as pure goethite. The specific surface area

  18. Mitophagy confers resistance to siderophore-mediated killing by Pseudomonas aeruginosa

    PubMed Central

    Kirienko, Natalia V.; Ausubel, Frederick M.; Ruvkun, Gary

    2015-01-01

    In the arms race of bacterial pathogenesis, bacteria produce an array of toxins and virulence factors that disrupt core host processes. Hosts mitigate the ensuing damage by responding with immune countermeasures. The iron-binding siderophore pyoverdin is a key virulence mediator of the human pathogen Pseudomonas aeruginosa, but its pathogenic mechanism has not been established. Here we demonstrate that pyoverdin enters Caenorhabditis elegans and that it is sufficient to mediate host killing. Moreover, we show that iron chelation disrupts mitochondrial homeostasis and triggers mitophagy both in C. elegans and mammalian cells. Finally, we show that mitophagy provides protection both against the extracellular pathogen P. aeruginosa and to treatment with a xenobiotic chelator, phenanthroline, in C. elegans. Although autophagic machinery has been shown to target intracellular bacteria for degradation (a process known as xenophagy), our report establishes a role for authentic mitochondrial autophagy in the innate immune defense against P. aeruginosa. PMID:25624506

  19. RNA-seq Analysis Reveals That an ECF σ Factor, AcsS, Regulates Achromobactin Biosynthesis in Pseudomonas syringae pv. syringae B728a

    PubMed Central

    Greenwald, Jessica W.; Greenwald, Charles J.; Philmus, Benjamin J.; Begley, Tadhg P.; Gross, Dennis C.

    2012-01-01

    Iron is an essential micronutrient for Pseudomonas syringae pv. syringae strain B728a and many other microorganisms; therefore, B728a has evolved methods of iron acquirement including the use of iron-chelating siderophores. In this study an extracytoplasmic function (ECF) sigma factor, AcsS, encoded within the achromobactin gene cluster is shown to be a major regulator of genes involved in the biosynthesis and secretion of this siderophore. However, production of achromobactin was not completely abrogated in the deletion mutant, implying that other regulators may be involved such as PvdS, the sigma factor that regulates pyoverdine biosynthesis. RNA-seq analysis identified 287 genes that are differentially expressed between the AcsS deletion mutant and the wild type strain. These genes are involved in iron response, secretion, extracellular polysaccharide production, and cell motility. Thus, the transcriptome analysis supports a role for AcsS in the regulation of achromobactin production and the potential activity of both AcsS and achromobactin in the plant-associated lifestyle of strain B728a. PMID:22529937

  20. Fusarinine C, a novel siderophore-based bifunctional chelator for radiolabeling with Gallium-68

    PubMed Central

    Zhai, Chuangyan; Summer, Dominik; Rangger, Christine; Haas, Hubertus; Haubner, Roland; Decristoforo, Clemens

    2015-01-01

    Fusarinine C (FSC), a siderophore-based chelator coupled with the model peptide c(RGDfK) (FSC(succ-RGD)3), revealed excellent targeting properties in vivo using positron emission tomography (PET). Here, we report the details of radiolabeling conditions and specific activity as well as selectivity for 68Ga. 68Ga labeling of FSC(succ-RGD)3 was optimized regarding peptide concentration, pH, temperature, reaction time, and buffer system. Specific activity (SA) of [68Ga]FSC(succ-RGD)3 was compared with 68Ga-1,4,7-triazacyclononane, 1-glutaric acid-4,7 acetic acid RGD ([68Ga]NODAGA-RGD). Stability was evaluated in 1000-fold ethylenediaminetetraacetic acid (EDTA) solution (pH 7) and phosphate-buffered saline (PBS). Metal competition tests (Fe, Cu, Zn, Al, and Ni) were carried out using [68Ga]-triacetylfusarinine C. High radiochemical yield was achieved within 5 min at room temperature, in particular allowing labeling with 68Ga up to pH 8 with excellent stability in 1000-fold EDTA solution and PBS. The 10-fold to 20-fold lower concentrations of FSC(succ-RGD)3 led to the same radiochemical yield compared with [68Ga]NODAGA-RGD with SA up to 1.8 TBq/μmol. Metal competition tests showed high selective binding of 68Ga to FSC. FSC is a multivalent siderophore-based bifunctional chelator allowing fast and highly selective labeling with 68Ga in a wide pH range and results in stable complexes with high SA. Thus it is exceptionally well suited for the development of new 68Ga-tracers for in vivo molecular imaging with PET. PMID:25874571

  1. Structural basis for effectiveness of siderophore-conjugated monocarbams against clinically relevant strains of Pseudomonas aeruginosa

    SciTech Connect

    Han, Seungil; Zaniewski, Richard P.; Marr, Eric S.; Lacey, Brian M.; Tomaras, Andrew P.; Evdokimov, Artem; Miller, J. Richard; Shanmugasundaram, Veerabahu

    2012-02-08

    Pseudomonas aeruginosa is an opportunistic Gram-negative pathogen that causes nosocomial infections for which there are limited treatment options. Penicillin-binding protein PBP3, a key therapeutic target, is an essential enzyme responsible for the final steps of peptidoglycan synthesis and is covalently inactivated by {beta}-lactam antibiotics. Here we disclose the first high resolution cocrystal structures of the P. aeruginosa PBP3 with both novel and marketed {beta}-lactams. These structures reveal a conformational rearrangement of Tyr532 and Phe533 and a ligand-induced conformational change of Tyr409 and Arg489. The well-known affinity of the monobactam aztreonam for P. aeruginosa PBP3 is due to a distinct hydrophobic aromatic wall composed of Tyr503, Tyr532, and Phe533 interacting with the gem-dimethyl group. The structure of MC-1, a new siderophore-conjugated monocarbam complexed with PBP3 provides molecular insights for lead optimization. Importantly, we have identified a novel conformation that is distinct to the high-molecular-weight class B PBP subfamily, which is identifiable by common features such as a hydrophobic aromatic wall formed by Tyr503, Tyr532, and Phe533 and the structural flexibility of Tyr409 flanked by two glycine residues. This is also the first example of a siderophore-conjugated triazolone-linked monocarbam complexed with any PBP. Energetic analysis of tightly and loosely held computed hydration sites indicates protein desolvation effects contribute significantly to PBP3 binding, and analysis of hydration site energies allows rank ordering of the second-order acylation rate constants. Taken together, these structural, biochemical, and computational studies provide a molecular basis for recognition of P. aeruginosa PBP3 and open avenues for future design of inhibitors of this class of PBPs.

  2. Hydroxamate siderophore-promoted reactions between iron(II) and nitroaromatic groundwater contaminants

    NASA Astrophysics Data System (ADS)

    Kim, Dongwook; Duckworth, Owen W.; Strathmann, Timothy J.

    2009-03-01

    Recent studies show that ferrous iron (Fe II), which is often abundant in anaerobic soil and groundwater, is capable of abiotically reducing many subsurface contaminants. However, studies also demonstrate that Fe II redox reactivity in geochemical systems is heavily dependent upon metal speciation. This contribution examines the influence of hydroxamate ligands, including the trihydroxamate siderophore desferrioxamine B (DFOB), on Fe II reactions with nitroaromatic groundwater contaminants (NACs). Experimental results demonstrate that ring-substituted NACs are reduced to the corresponding aniline products in aqueous solutions containing Fe II complexes with DFOB and two monohydroxamate ligands (acetohydroxamic acid and salicylhydroxamic acid). Reaction rates are heavily dependent upon solution conditions and the identities of both the Fe II-complexing hydroxamate ligand and the target NAC. Trends in the observed pseudo-first-order rate constants for reduction of 4-chloronitrobenzene ( kobs, s -1) are quantitatively linked to the formation of Fe II species with standard one-electron reduction potentials, EH0 (Fe III/Fe II), below -0.3 V. Linear free energy relationships correlate reaction rates with the EH0 (Fe III/Fe II) values of different electron-donating Fe II complexes and with the apparent one-electron reduction potentials of different electron-accepting NACs, EH1'(ArNO 2). Experiments describing a redox auto-decomposition mechanism for Fe II-DFOB complexes that occurs at neutral pH and has implications for the stability of hydroxamate siderophores in anaerobic environments are also presented. Results from this study indicate that hydroxamates and other Fe III-stabilizing organic ligands can form highly redox-active Fe II complexes that may contribute to the natural attenuation and remediation of subsurface contaminants.

  3. Biosynthesis of cylindrospermopsin.

    PubMed

    Burgoyne, D L; Hemscheidt, T K; Moore, R E; Runnegar, M T

    2000-01-14

    Studies on the biosynthesis of cylindrospermopsin (1), a potent hepatotoxin associated with the cyanobacterium Cylindrospermopsis raciborskii, indicate that 1 is an acetogenin with guanidinoacetic acid serving as the starter unit of the polyketide chain. Feeding experiments show that C14 and C15 of 1 are derived from C1 and C2 of glycine, respectively, and C4 through C13 arise from five contiguous acetate units attached head to tail. The methyl carbon on C13 originates from the C(1) pool. The starter unit, established by the incorporation of [guanidino-(13)C,alpha-(15)N]-guanidinoacetic acid into N16 and C17 of 1, does not appear to be formed from glycine by known amidination pathways. The origin of the NH-CO-NH segment in the uracil ring is also unknown. PMID:10813909

  4. Biosynthesis of plant sulfolipids

    SciTech Connect

    Kleppinger-Sparace, K.; Mudd, J.B.; Sparace, S. )

    1989-04-01

    The complete biosynthesis of sulfoquinovosyldiacylglycerol (SQDG) remains undetermined although dark synthesis of SQDG by chloroplasts supplied with AP{sup 35}S, PAP{sup 35}S or {sup 35}SO{sub 4} plus ATP suggests the sulfur moiety arises from either APS or sulfite (1). Sulfate incorporation into sulfolipids in isolated chloroplasts and in intact roots is reported here and compared to lipids labelled by {sup 14}C-acetate or {sup 14}C-glycerol. Several unknown {sup 35}S-labelled chloroform-soluble compounds were isolated from sterile roots. These {sup 35}S-labelled compounds differ from those of the chloroplast, identified as elemental sulfur forms. Identification of the unknown root compounds is in progress. Unlike chloroplast, isolated root plastids do not synthesis SQDG from sulfate plus ATP suggesting a requirement for an activated form of sulfate, such as APS or PAPS.

  5. Designer microbes for biosynthesis

    PubMed Central

    Quin, Maureen B.; Schmidt-Dannert, Claudia

    2014-01-01

    Microbes have long been adapted for the biosynthetic production of useful compounds. There is increasing demand for the rapid and cheap microbial production of diverse molecules in an industrial setting. Microbes can now be designed and engineered for a particular biosynthetic purpose, thanks to recent developments in genome sequencing, metabolic engineering, and synthetic biology. Advanced tools exist for the genetic manipulation of microbes to create novel metabolic circuits, making new products accessible. Metabolic processes can be optimized to increase yield and balance pathway flux. Progress is being made towards the design and creation of fully synthetic microbes for biosynthetic purposes. Together, these emerging technologies will facilitate the production of designer microbes for biosynthesis. PMID:24646570

  6. Relative importance of fluorescent siderophores and other factors in biological control of Gaeumannomyces graminis var. tritici by Pseudomonas fluorescens 2-79 and M4-80R.

    PubMed Central

    Hamdan, H; Weller, D M; Thomashow, L S

    1991-01-01

    Pseudomonas fluorescens 2-79 suppresses take-all, a major root disease of wheat caused by Gaeumannomyces graminis var. tritici. The bacteria produce an antibiotic, phenazine-1-carboxylic acid (PCA), and a fluorescent pyoverdin siderophore. Previous studies have established that PCA has an important role in the biological control of take-all but that antibiotic production does not account fully for the suppressiveness of the strain. To define the role of the pyoverdin siderophore more precisely, mutants deficient in production of the antibiotic, the siderophore, or both factors were constructed and compared with the parental strain for control of take-all on wheat roots. In all cases, strains that produced PCA were more suppressive than those that did not, and pyoverdin-deficient mutant derivatives controlled take-all as effectively as their respective fluorescent parental strains. Thus, the phenazine antibiotic was the dominant factor in disease suppression and the fluorescent siderophore had little or no role. The siderophore also was of minor importance in a second strain, P. fluorescens M4-80R, that does not produce PCA. Strains 2-79 and M4-80R both produced substances distinct from the pyoverdin siderophore that were responsible for fungal inhibition in vitro under iron limitation, but these substances also had, at most, a minor role in disease suppression in situ. PMID:1838240

  7. A new fluorimetric method for the detection and quantification of siderophores using Calcein Blue, with potential as a bacterial detection tool.

    PubMed

    Sankaranarayanan, Ranjini; Alagumaruthanayagam, Alagiachidambaram; Sankaran, Krishnan

    2015-03-01

    The presence of microorganisms in biological fluids like urine and blood is an indication of vulnerability to infections. Iron is one of the important micronutrients required for bacterial growth. In an iron-deficit environment, bacteria release high-affinity iron-chelating compounds called siderophores which can be used as non-invasive target molecules for the detection of such pathogens. However, only limited reagents and procedures are available to detect the presence of these organic molecules. The present study aims at detecting the presence of siderophores in the iron-depleted media, exploiting the reversible quenching of Calcein Blue and iron(III) complex. The fluorescence of Calcein Blue is known to be quenched in the presence of iron(III); if a stronger chelator removes this ion from the fluorophore, the fluorescence of the fluorophore is regained. This behaviour of the fluorophore was exploited to detect and quantify siderophores down to 50 and 800 nM equivalent of standard siderophore, deferroxamine mesylate (desferal) in Dulbecco's PBS and siderophore quantification (SPQ) medium, respectively. The siderophores released by pathogens, equivalent to standard desferal, were in the range of 1.29 to 5.00 μM and those for non-pathogens were below 1.19 μM. The simple, sensitive and cost-effective method performed in a 96-well plate was able to detect and quantify iron chelators within 7-8 h of incubation. PMID:25634020

  8. An update on iron acquisition by Legionella pneumophila: new pathways for siderophore uptake and ferric iron reduction

    PubMed Central

    Cianciotto, Nicholas P

    2015-01-01

    Iron acquisition is critical for the growth and pathogenesis of Legionella pneumophila, the causative agent of Legionnaires’ disease. L. pneumophila utilizes two main modes of iron assimilation, namely ferrous iron uptake via the FeoB system and ferric iron acquisition through the action of the siderophore legiobactin. This review highlights recent studies concerning the mechanism of legiobactin assimilation, the impact of c-type cytochromes on siderophore production, the importance of legiobactin in lung infection and a newfound role for a bacterial pyomelanin in iron acquisition. These data demonstrate that key aspects of L. pneumophila iron acquisition are significantly distinct from those of long-studied, ‘model’ organisms. Indeed, L. pneumophila may represent a new paradigm for a variety of other intracellular parasites, pathogens and under-studied bacteria. PMID:26000653

  9. Serine biosynthesis and transport defects.

    PubMed

    El-Hattab, Ayman W

    2016-07-01

    l-serine is a non-essential amino acid that is biosynthesized via the enzymes phosphoglycerate dehydrogenase (PGDH), phosphoserine aminotransferase (PSAT), and phosphoserine phosphatase (PSP). Besides its role in protein synthesis, l-serine is a potent neurotrophic factor and a precursor of a number of essential compounds including phosphatidylserine, sphingomyelin, glycine, and d-serine. Serine biosynthesis defects result from impairments of PGDH, PSAT, or PSP leading to systemic serine deficiency. Serine biosynthesis defects present in a broad phenotypic spectrum that includes, at the severe end, Neu-Laxova syndrome, a lethal multiple congenital anomaly disease, intermediately, infantile serine biosynthesis defects with severe neurological manifestations and growth deficiency, and at the mild end, the childhood disease with intellectual disability. A serine transport defect resulting from deficiency of the ASCT1, the main transporter for serine in the central nervous system, has been recently described in children with neurological manifestations that overlap with those observed in serine biosynthesis defects. l-serine therapy may be beneficial in preventing or ameliorating symptoms in serine biosynthesis and transport defects, if started before neurological damage occurs. Herein, we review serine metabolism and transport, the clinical, biochemical, and molecular aspects of serine biosynthesis and transport defects, the mechanisms of these diseases, and the potential role of serine therapy. PMID:27161889

  10. Small-molecule inhibitors suppress the expression of both type III secretion and amylovoran biosynthesis genes in Erwinia amylovora.

    PubMed

    Yang, Fan; Korban, Schuyler S; Pusey, P Lawrence; Elofsson, Michael; Sundin, George W; Zhao, Youfu

    2014-01-01

    The type III secretion system (T3SS) and exopolysaccharide (EPS) amylovoran are two essential pathogenicity factors in Erwinia amylovora, the causal agent of the serious bacterial disease fire blight. In this study, small molecules that inhibit T3SS gene expression in E. amylovora under hrp (hypersensitive response and pathogenicity)-inducing conditions were identified and characterized using green fluorescent protein (GFP) as a reporter. These compounds belong to salicylidene acylhydrazides and also inhibit amylovoran production. Microarray analysis of E. amylovora treated with compounds 3 and 9 identified a total of 588 significantly differentially expressed genes. Among them, 95 and 78 genes were activated and suppressed by both compounds, respectively, when compared with the dimethylsulphoxide (DMSO) control. The expression of the majority of T3SS genes in E. amylovora, including hrpL and the avrRpt2 effector gene, was suppressed by both compounds. Compound 3 also suppressed the expression of amylovoran precursor and biosynthesis genes. However, both compounds induced significantly the expression of glycogen biosynthesis genes and siderophore biosynthesis, regulatory and transport genes. Furthermore, many membrane, lipoprotein and exported protein-encoding genes were also activated by both compounds. Similar expression patterns were observed for compounds 1, 2 and 4. Using crab apple flower as a model, compound 3 was capable of reducing disease development in pistils. These results suggest a common inhibition mechanism shared by salicylidene acylhydrazides and indicate that small-molecule inhibitors that disable T3SS function could be explored to control fire blight disease. PMID:23915008

  11. Tenacibactins A-D, hydroxamate siderophores from a marine-derived bacterium, Tenacibaculum sp. A4K-17.

    PubMed

    Jang, Jae-Hyuk; Kanoh, Kaneo; Adachi, Kyoko; Matsuda, Satoru; Shizuri, Yoshikazu

    2007-04-01

    Four new hydroxamate siderophores, tenacibactins A-D (1-4), were isolated from a culture broth of the marine-derived bacterium Tenacibaculum sp. A4K-17. The structures of these tenacibactins were determined by NMR analyses and ESIMS/MS experiments. The iron-binding (chelating) activity of 1-4 was evaluated by the chrome azurol sulfonate (CAS) assay. PMID:17319723

  12. BIOSYNTHESIS OF STRESS ETHYLENE IN SOYBEAN SEEDLINGS: SIMILARITIES TO ENDOGENOUS ETHYLENE BIOSYNTHESIS

    EPA Science Inventory

    The similarity of stress ethylene biosynthesis in whole plants to endogenous ethylene biosynthesis was investigated using two inhibitors of ethylene biosynthesis, amino-ethoxyvinylglycine (AVG) and cobalt chloride (Co2+); and the intermediates, methionine, S-adenosylmethionine (S...

  13. Terbium, a fluorescent probe for investigation of siderophore pyochelin interactions with its outer membrane transporter FptA.

    PubMed

    Yang, Binsheng; Hoegy, Françoise; Mislin, Gaëtan L A; Mesini, Philippe J; Schalk, Isabelle J

    2011-10-01

    Pyochelin (Pch) is a siderophore and FptA is its outer membrane transporter produced by Pseudomonas aeruginosa to import iron. The fluorescence of the element terbium is affected by coordinated ligands and it can therefore be used as a probe to investigate the pyochelin-iron uptake pathway in P. aeruginosa. At pH 8.0, terbium fluorescence is greatly enhanced in the presence of pyochelin indicating chelation of the metal by the siderophore. Titration curves showed a 2:1 (Pch:Tb(3+)) stoichiometry and an affinity of K=(2±-1)×10(11)M(-2) was determined. Pch-Tb interaction with the transporter FptA could be followed in vitro and in vivo in P. aeruginosa cells, by Fluorescence Resonance Energy Transfer (FRET) between three partners: the tryptophans of FptA (donor), Pch (acceptor for the Trps and donor for Tb(3+)) and Tb(3+) (acceptor). Pch-Tb binds to the Pch-Fe outer membrane transporter FptA with a dissociation constant (K(d)) of 4.6μM. This three-partner FRET is a potentially valuable tool for investigation of the interactions between FptA and its siderophore Pch. PMID:21861955

  14. Lead-enhanced siderophore production and alteration in cell morphology in a Pb-resistant Pseudomonas aeruginosa strain 4EA.

    PubMed

    Naik, Milind Mohan; Dubey, Santosh Kumar

    2011-02-01

    A lead-resistant bacterial strain 4EA from soil contaminated with car battery waste from Goa, India was isolated and identified as Pseudomonas aeruginosa. This lead-resistant bacterial isolate interestingly revealed lead-enhanced siderophore (pyochelin and pyoverdine) production up to 0.5 mM lead nitrate whereas cells exhibit a significant decline in siderophore production above 0.5 mM lead nitrate. The bacterial cells also revealed significant alteration in cell morphology as size reduction when exposed to 0.8 mM lead nitrate. Enhanced production of siderophore was evidently detected by chrome azurol S agar diffusion (CASAD) assay as increase in diameter of orange halo, and reduction in bacterial size along with significant biosorption of lead was recorded by scanning electron microscopy coupled with energy dispersive X-ray spectrometry (SEM-EDX). Pseudomonas aeruginosa strain 4EA also exhibits cross tolerance to other toxic metals viz. cadmium, mercury, and zinc besides resistance to multiple antibiotics such as ampicillin, erythromycin, amikacin, cephalexin, co-trimoxazole, mecillinam, lincomycin, ciphaloridine, oleondamycin, and nalidixic acid. PMID:20661573

  15. Stereoselectivity in Polyphenol Biosynthesis

    NASA Technical Reports Server (NTRS)

    Lewis, Norman G.; Davin, Laurence B.

    1992-01-01

    Stereoselectivity plays an important role in the late stages of phenyl-propanoid metabolism, affording lignins, lignans, and neolignans. Stereoselectivity is manifested during monolignol (glucoside) synthesis, e.g., where the geometry (E or Z) of the pendant double bond affects the specificity of UDPG:coniferyl alcohol glucosyltransferases in different species. Such findings are viewed to have important ramifications in monolignol transport and storage processes, with roles for both E- and Z-monolignols and their glucosides in lignin/lignan biosynthesis being envisaged. Stereoselectivity is also of great importance in enantiose-lective enzymatic processes affording optically active lignans. Thus, cell-free extracts from Forsythia species were demonstrated to synthesize the enantiomerically pure lignans, (-)-secoisolariciresinol, and (-)-pinoresinol, when NAD(P)H, H2O2 and E-coniferyl alcohol were added. Progress toward elucidating the enzymatic steps involved in such highly stereoselective processes is discussed. Also described are preliminary studies aimed at developing methodologies to determine the subcellular location of late-stage phenylpropanoid metabolites (e.g., coniferyl alcohol) and key enzymes thereof, in intact tissue or cells. This knowledge is essential if questions regarding lignin and lignan tissue specificity and regulation of these processes are to be deciphered.

  16. Biosynthesis of Dolichyl Phosphate

    PubMed Central

    Hopp, H. Esteban; Daleo, Gustavo R.; Romero, Pedro A.; Lezica, Rafael Pont

    1978-01-01

    This is the first report not only on the presence of polyprenyl phosphates and their site of synthesis in algae, but also on the formation of their sugar derivatives in this system. A glucose acceptor lipid was isolated from the nonphotosynthetic alga Prototheca zopfii. The lipid was acidic and resistant to mild acid and alkaline treatments. The glucosylated lipid was labile to mild acid hydrolysis and resistant to phenol treatment and catalytic hydrogenation, as dolichyl phosphate glucose is. These results are consistent with the properties of an α-saturated polyprenyl phosphate. The polyprenylic nature of the lipid was confirmed by biosynthesis from radioactive mevalonate. The [14C]lipid had the same chromatographic properties as dolichyl phosphate in DEAE-cellulose and Sephadex LH-20. Strong alkaline treatment and enzymic hydrolysis liberated free alcohols with chain lengths ranging from C90 to C105, C95 and C100 being the most abundant molecular forms. The glucose acceptor activity of the biosynthesized polyprenyl phosphate was confirmed. The ability of different subcellular fractions to synthesize dolichyl phosphate was studied. Mitochondria and the Golgi apparatus were the sites of dolichyl phosphate synthesis from mevalonate. PMID:16660269

  17. Inter-relationships of MnO 2 precipitation, siderophore-Mn (III) complex formation, siderophore degradation, and iron limitation in Mn (II)-oxidizing bacterial cultures

    NASA Astrophysics Data System (ADS)

    Parker, Dorothy L.; Morita, Takami; Mozafarzadeh, Mylene L.; Verity, Rebecca; McCarthy, James K.; Tebo, Bradley M.

    2007-12-01

    To examine the pathways that form Mn (III) and Mn (IV) in the Mn (II)-oxidizing bacterial strains Pseudomonas putida GB-1 and MnB1, and to test whether the siderophore pyoverdine (PVD) inhibits Mn (IV)O 2 formation, cultures were subjected to various protocols at known concentrations of iron and PVD. Depending on growth conditions, P. putida produced one of two oxidized Mn species - either soluble PVD-Mn (III) complex or insoluble Mn (IV)O 2 minerals - but not both simultaneously. PVD-Mn (III) was present, and MnO 2 precipitation was inhibited, both in iron-limited cultures that had synthesized 26-50 μM PVD and in iron-replete (non-PVD-producing) cultures that were supplemented with 10-550 μM purified PVD. PVD-Mn (III) arose by predominantly ligand-mediated air oxidation of Mn (II) in the presence of PVD, based on the following evidence: (a) yields and rates of this reaction were similar in sterile media and in cultures, and (b) GB-1 mutants deficient in enzymatic Mn oxidation produced PVD-Mn (III) as efficiently as wild type. Only wild type, however, could degrade PVD-Mn (III), a process linked to the production of both MnO 2 and an altered PVD with absorbance and fluorescence spectra markedly different from those of either PVD or PVD-Mn (III). Two conditions, the presence of bioavailable iron and the absence of PVD at concentrations exceeding those of Mn, both had to be satisfied for MnO 2 to appear. These results suggest that P. putida cultures produce soluble Mn (III) or MnO 2 by different and mutually inhibitory pathways: enzymatic catalysis yielding MnO 2 under iron sufficiency or PVD-promoted oxidation yielding PVD-Mn (III) under iron limitation. Since PVD-producing Pseudomonas species are environmentally prevalent Mn oxidizers, these data predict influences of iron (via PVD-Mn (III) versus MnO 2) on the global oxidation/reduction cycling of various pollutants, recalcitrant organic matter, and elements such as C, S, N, Cr, U, and Mn.

  18. Manganese(III) binding to a pyoverdine siderophore produced by a manganese(II)-oxidizing bacterium

    NASA Astrophysics Data System (ADS)

    Parker, Dorothy L.; Sposito, Garrison; Tebo, Bradley M.

    2004-12-01

    The possible roles of siderophores (high affinity chelators of iron(III)) in the biogeochemistry of manganese remain unknown. Here we investigate the interaction of Mn(III) with a pyoverdine-type siderophore (PVD MnB1) produced by the model Mn(II)-oxidizing bacterium Pseudomonas putida strain MnB1. PVD MnB1 confirmed typical pyoverdine behavior with respect to: (a) its absorption spectrum at 350-600 nm, both in the absence and presence of Fe(III), (b) the quenching of its fluorescence by Fe(III), (c) the formation of a 1:1 complex with Fe(III), and (d) the thermodynamic stability constant of its Fe(III) complex. The Mn(III) complex of PVD MnB1 had a 1:1 Mn:pvd molar ratio, showed fluorescence quenching, and exhibited a light absorption spectrum (A max = 408-410 nm) different from that of either PVD MnB1-Fe(III) or uncomplexed PVD MnB1. Mn(III) competed strongly with Fe(III) for binding by PVD MnB1 in culture filtrates (pH 8, 4°C). Equilibration with citrate, a metal-binding ligand, did not detectably release Mn from its PVD MnB1 complex at a citrate/PVD MnB1 molar ratio of 830 (pH 8, 4°C), whereas pyrophosphate under the same conditions removed 55% of the Mn from its PVD MnB1 complex. Most of the PVD MnB1-complexed Mn was released by reaction with ascorbate, a reducing agent, or with EDTA, a ligand that is also oxidized by Mn(III). Data on the competition for binding to PVD MnB1 by Fe(III) vs. Mn(III) were used to determine a thermodynamic stability constant (nominally at 4°C) for the neutral species MnHPVD MnB1 (log K = 47.5 ± 0.5, infinite dilution reference state). This value was larger than that determined for FeHPVD MnB1 (log K = 44.6 ± 0.5). This result has important implications for the metabolism, solubility, speciation, and redox cycling of manganese, as well as for the biologic uptake of iron.

  19. Kinetics of Biosynthesis of Iron-Regulated Membrane Proteins in Escherichia coli

    PubMed Central

    Klebba, Phillip E.; McIntosh, Mark A.; Neilands, J. B.

    1982-01-01

    Using biological iron chelators to control specifically iron availability to Escherichia coli K-12 in conjunction with radioactive pulse-labels, we examined the biosynthesis of six iron-regulated membrane proteins. Iron deprivation induced the synthesis of five proteins, which had molecular weights of 83,000 (83K), 81K (Fep), 78K (TonA), 74K (Cir), and 25K. The kinetics of induction were the same in entA and entA+ strains, but were affected by the initial iron availability in the media. Iron-poor cells induced rapidly (half-time, 10 min), whereas iron-rich cells began induction after a lag and showed a slower induction half-time (30 min). Within this general pattern of induction after iron deprivation, several different kinetic patterns were apparent. The 83K, 81K, and 74K proteins were coordinately controlled under all of the conditions examined. The 78K and 25K proteins were regulated differently. The synthesis of a previously unrecognized 90K inner membrane protein was inhibited by iron deprivation and stimulated by iron repletion. Both ferrichrome and ferric enterobactin completely repressed 81K and 74K synthesis when the siderophores were supplied at concentrations of 5 μM in vivo (half-time, 2.5 min). At concentrations less than 5 μM, however, both siderophores repressed synthesis only temporarily; the duration of repression was proportional to the amount of ferric siderophore added. The half-lives of the 81K and 74K mRNAs, as measured by rifampin treatment, were 1.2 and 1.6 min, respectively. The results of this study suggest that enteric bacteria are capable of instantaneously detecting and reacting to fluctuations in the extracellular iron concentration and that they store iron during periods of iron repletion for utilization during periods of iron stress. Neither iron storage nor iron regulation of envelope protein synthesis is dependent on the ability of the bacteria to form heme. Images PMID:6174499

  20. Burkholderia Diffusible Signal Factor Signals to Francisella novicida To Disperse Biofilm and Increase Siderophore Production

    PubMed Central

    Dean, Scott N.; Chung, Myung-Chul

    2015-01-01

    In many bacteria, the ability to modulate biofilm production relies on specific signaling molecules that are either self-produced or made by neighboring microbes within the ecological niche. We analyzed the potential interspecies signaling effect of the Burkholderia diffusible signal factor (BDSF) on Francisella novicida, a model organism for Francisella tularensis, and demonstrated that BDSF both inhibits the formation and causes the dispersion of Francisella biofilm. Specificity was demonstrated for the cis versus the trans form of BDSF. Using transcriptome sequencing, quantitative reverse transcription-PCR, and activity assays, we found that BDSF altered the expression of many F. novicida genes, including genes involved in biofilm formation, such as chitinases. Using a chitinase inhibitor, the antibiofilm activity of BDSF was also shown to be chitinase dependent. In addition, BDSF caused an increase in RelA expression and increased levels of (p)ppGpp, leading to decreased biofilm production. These results support our observation that exposure of F. novicida to BDSF causes biofilm dispersal. Furthermore, BDSF upregulated the genes involved in iron acquisition (figABCD), increasing siderophore production. Thus, this study provides evidence for a potential role and mechanism of diffusible signal factor (DSF) signaling in the genus Francisella and suggests the possibility of interspecies signaling between Francisella and other bacteria. Overall, this study suggests that in response to the interspecies DSF signal, F. novicida can alter its gene expression and regulate its biofilm formation. PMID:26231649

  1. Siderophore production and membrane alterations by Bordetella pertussis in response to iron starvation.

    PubMed Central

    Agiato, L A; Dyer, D W

    1992-01-01

    Bordetella pertussis was grown in iron (Fe)-free defined medium to limit the growth of the organism. Doubling times of the Fe-starved organism increased by approximately 1 h, and a 40% reduction in the final extent of growth in Fe-depleted medium was observed. Under these conditions, a hydroxamate siderophore named bordetellin was secreted by B. pertussis. Lactoferrin and transferrin supported growth of B. pertussis even when the protein was sequestered inside dialysis tubing. This suggested that binding of lactoferrin and transferrin to B. pertussis was not essential and that bordetellin production plays a major role in Fe uptake. Solid-phase dot blot assays indicated weak binding of lactoferrin to the cell surface, consistent with previous reports of a lactoferrin receptor. Three new proteins of 97, 77, and 63 kDa were synthesized in response to Fe starvation. Fe-inducible proteins of 103, 72, 24, 21, and 18 kDa were also observed. The synthesis of lipopolysaccharide was also altered by Fe availability. Images PMID:1309510

  2. Iron(III) coordination chemistry of alterobactin A: a siderophore from the marine bacterium Alteromonas luteoviolacea.

    PubMed

    Holt, Pamela D; Reid, Richard R; Lewis, Brent L; Luther, George W; Butler, Alison

    2005-10-17

    Alterobactin A is a siderophore produced by the oceanic bacterium Alteromonas luteoviolacea. The thermodynamic stability constant of the ferric alterobactin A (Alt-A) complex was estimated from electrochemical measurements on the basis of a previously reported linear relationship between the reduction potentials and the pH-independent stability constants for known iron(III) complexes. The reduction potential of the ferric alterobactin A complex determined by square wave voltammetry is -0.972 V vs SCE and reversible, corresponding to a thermodynamic stability constant of 10(51+/-2). Potentiometric titration of Fe(III)-Alt-A shows the release of six protons on complexation of Fe(III) to Alt-A. The 1H NMR resonances of the Ga(III)-Alt-A complex show that the C-4, C-5, and C-6 catecholate protons and the C(alpha) and C(beta) protons of both beta-hydroxyaspartate moieties are shifted downfield relative to the free ligand, which along with the potentiometric titration data is consistent with a complex in which Fe(III) is coordinated by both catecholate oxygen atoms and both oxygen atoms of each beta-hydroxyaspartate. The UV-vis spectrum of Fe(III)-Alt-A is invariant over the pH range 4-9, indicating the coordination does not change over a wide pH range. In addition, in the absence of a coordinated metal ion, the serine ester of Alt-A hydrolyzes forming Alt-B. PMID:16212394

  3. Debaryomyces mycophilus sp. nov., a siderophore-dependent yeast isolated from woodlice.

    PubMed

    Thanh, Vu Nguyen; Van Dyk, Martha S; Wingfield, Michael J

    2002-08-01

    Four strains of an ascogenous yeast were isolated from the guts of the woodlice species Armadillidium vulgare (Latreille). This yeast differed from all known yeasts by its inability to grow in culture without the presence of a metabolite produced by some common soil fungi such as Cladosporium cladosporioides, Aspergillus alliaceus, and Penicillium spp. Phylogenetic analysis based on 18S rDNA and 26S rDNA (domain D1/D2) sequences indicated that the yeast represents a new taxon in the genus Debaryomyces. The new species Debaryomyces mycophilus is thus proposed. It was, furthermore, shown that the fungal metabolite necessary for growth of D. mycophilus did not provide the yeast with carbon, nitrogen or vitamins. The active compound was partially purified and it was shown that it is a siderophore used by the yeast as a source of iron. The addition of ferrichrome or high concentrations of FeCl(3) to growth media replaced the obligate dependence on a fungal metabolite. Symbiosis among fungi, based on the availability and utilization of iron, is an aspect of mycology that has not previously been recognized. The addition of chelated iron to isolation media could lead to the discovery of many unknown yeasts and fungi. PMID:12702293

  4. A novel siderophore system is essential for the growth of Pseudomonas aeruginosa in airway mucus.

    PubMed

    Gi, Mia; Lee, Kang-Mu; Kim, Sang Cheol; Yoon, Joo-Heon; Yoon, Sang Sun; Choi, Jae Young

    2015-01-01

    Pseudomonas aeruginosa establishes airway infections in Cystic Fibrosis patients. Here, we investigate the molecular interactions between P. aeruginosa and airway mucus secretions (AMS) derived from the primary cultures of normal human tracheal epithelial (NHTE) cells. PAO1, a prototype strain of P. aeruginosa, was capable of proliferating during incubation with AMS, while all other tested bacterial species perished. A PAO1 mutant lacking PA4834 gene became susceptible to AMS treatment. The ΔPA4834 mutant was grown in AMS supplemented with 100 μM ferric iron, suggesting that the PA4834 gene product is involved in iron metabolism. Consistently, intracellular iron content was decreased in the mutant, but not in PAO1 after the AMS treatment. Importantly, a PAO1 mutant unable to produce both pyoverdine and pyochelin remained viable, suggesting that these two major siderophore molecules are dispensable for maintaining viability during incubation with AMS. The ΔPA4834 mutant was regrown in AMS amended with 100 μM nicotianamine, a phytosiderophore whose production is predicted to be mediated by the PA4836 gene. Infectivity of the ΔPA4834 mutant was also significantly compromised in vivo. Together, our results identify a genetic element encoding a novel iron acquisition system that plays a previously undiscovered role in P. aeruginosa airway infection. PMID:26446565

  5. A novel siderophore system is essential for the growth of Pseudomonas aeruginosa in airway mucus

    PubMed Central

    Gi, Mia; Lee, Kang-Mu; Kim, Sang Cheol; Yoon, Joo-Heon; Yoon, Sang Sun; Choi, Jae Young

    2015-01-01

    Pseudomonas aeruginosa establishes airway infections in Cystic Fibrosis patients. Here, we investigate the molecular interactions between P. aeruginosa and airway mucus secretions (AMS) derived from the primary cultures of normal human tracheal epithelial (NHTE) cells. PAO1, a prototype strain of P. aeruginosa, was capable of proliferating during incubation with AMS, while all other tested bacterial species perished. A PAO1 mutant lacking PA4834 gene became susceptible to AMS treatment. The ΔPA4834 mutant was grown in AMS supplemented with 100 μM ferric iron, suggesting that the PA4834 gene product is involved in iron metabolism. Consistently, intracellular iron content was decreased in the mutant, but not in PAO1 after the AMS treatment. Importantly, a PAO1 mutant unable to produce both pyoverdine and pyochelin remained viable, suggesting that these two major siderophore molecules are dispensable for maintaining viability during incubation with AMS. The ΔPA4834 mutant was regrown in AMS amended with 100 μM nicotianamine, a phytosiderophore whose production is predicted to be mediated by the PA4836 gene. Infectivity of the ΔPA4834 mutant was also significantly compromised in vivo. Together, our results identify a genetic element encoding a novel iron acquisition system that plays a previously undiscovered role in P. aeruginosa airway infection. PMID:26446565

  6. Acceleration of Fe-silicate mineral dissolution for CO2 sequestration via microbial siderophore production

    NASA Astrophysics Data System (ADS)

    Torres, M. A.; Nealson, K. H.; West, A.

    2013-12-01

    While the dissolution of silicate minerals will ultimately neutralize anthropogenic CO2 emissions, the slow natural timescale of this process limits its ability to mitigate any of the societal impacts of high atmospheric pCO2. As a result, much research has been focused on developing ways to significantly accelerate silicate mineral dissolution rates. Harnessing the effects of microbial activity is one particularly attractive strategy because research has shown that microbes can appreciably accelerate mineral dissolution rates and they require little external energy input. At present, one major hurdle in the development of microbe-based CO2 sequestration techniques is the observation that bacteria only accelerate dissolution rates under particular culturing conditions. In this work, natural and genetic mutant strains of the bacterial genera Shewanella, Pseudomonas, and Marinobacter are used to identify the geochemical and genetic factors that underlie the 'accelerated-weathering phenotype' in order to support the development of microbe-based CO2 sequestration techniques using olivine as a model mineral. Preliminary results suggest that microbial siderophore production at circum-neutral pH results in significantly accelerated olivine dissolution rates.

  7. Nanospecific Inhibition of Pyoverdine Siderophore Production in Pseudomonas Chlororaphis O6 by CuO Nanoparticles

    SciTech Connect

    Dimkpa, Christian O.; McLean, Joan E.; Britt, David W.; Johnson, William P.; Arey, Bruce W.; Lea, Alan S.; Anderson, Anne J.

    2012-03-01

    As traditional antibiotics become less effective against a growing number of pathogens, engineered nanoparticles (NPs) are becoming more widely applied as biocides. NPs of Ag, ZnO, and CuO exhibit dose-dependent antimicrobial activity; however, information is scant on the impact of sublethal levels of NPs on bacteria. In this paper, we evaluated the effect of a sublethal concentration (200 mg/L) of commercial CuO NPs on the expression of genes involved in the production of the fluorescent siderophore, pyoverdine (PVD) in the plant-beneficial bacterium Pseudomonas chlororaphis O6. PVDs are important in microbe-microbe and microbe-plant interactions, and are a virulence factor in pathogenic pseudomonads. Cells challenged with the NPs had reduced amounts of PVD in their periplasm and the external medium. The NPs impaired the expression of genes involved in transport of the PVD precursor through the plasmamembrane, PVD maturation in the periplasm, and export through the outer membrane. Also, expression from one of three predicted Fe-PVD receptors was reduced by the NPs. As these effects were not observed for cells challenged with copper ions, this is a nanoparticlespecific phenomenon mediating cellular reprogramming in bacteria, affecting secondary metabolism and thus associated critical microbial processes. The regulation of bacterial genes and secondary metabolites by sublethal doses of a common metal oxide NP has strong environmental and medical implications.

  8. Siderophores in Cloud Waters and Potential Impact on Atmospheric Chemistry: Production by Microorganisms Isolated at the Puy de Dôme Station.

    PubMed

    Vinatier, Virginie; Wirgot, Nolwenn; Joly, Muriel; Sancelme, Martine; Abrantes, Magali; Deguillaume, Laurent; Delort, Anne-Marie

    2016-09-01

    A total of 450 bacteria and yeast strains isolated from cloud waters sampled at the puy de Dôme station in France (1465 m) were screened for their ability to produce siderophores. To achieve this, a high-throughput method in 96-well plates was adapted from the CAS (chrome azurol S) method. Notably, 42% of the isolates were siderophore producers. This production was examined according to the phyla of the tested strains and the type of chelating functional groups (i.e., hydroxamate, catechol, and mixed type). The most active bacteria in the clouds belong to the γ-Proteobacteria class, among which the Pseudomonas genus is the most frequently encountered. γ-Proteobacteria are produced in the majority of mixed function siderophores, such as pyoverdines, which bear a photoactive group. Finally, siderophore production was shown to vary with the origin of the air masses. The organic speciation of iron remains largely unknown in warm clouds. Our results suggest that siderophores could partly chelate Fe(III) in cloud waters and thus potentially impact the chemistry of the atmospheric aqueous phase. PMID:27479540

  9. Auxin biosynthesis and storage forms

    PubMed Central

    Strader, Lucia C.

    2013-01-01

    The plant hormone auxin drives plant growth and morphogenesis. The levels and distribution of the active auxin indole-3-acetic acid (IAA) are tightly controlled through synthesis, inactivation, and transport. Many auxin precursors and modified auxin forms, used to regulate auxin homeostasis, have been identified; however, very little is known about the integration of multiple auxin biosynthesis and inactivation pathways. This review discusses the many ways auxin levels are regulated through biosynthesis, storage forms, and inactivation, and the potential roles modified auxins play in regulating the bioactive pool of auxin to affect plant growth and development. PMID:23580748

  10. Alternate biosynthesis of valerenadiene and related sesquiterpenes.

    PubMed

    Paknikar, Shashikumar K; Kadam, Shahuraj H; Ehrlich, April L; Bates, Robert B

    2013-09-01

    It is proposed that the biosynthesis of the sesquiterpene valerenadiene, a key intermediate in the biosynthesis of a sedative in valerian, involves cyclopropane and not cyclobutane intermediates and includes as a key step a cyclopropylcarbinylcation-cyclopropylcarbinylcation rearrangement analogous to the one observed in the conversion of presqualene to squalene in triterpene and steroid biosynthesis. Similar mechanisms are proposed for the biosynthesis of the related sesquiterpenes pacifigorgiol, tamariscene and (+)-pacifigorgia-1,10-diene. PMID:24273843

  11. Stability of the Cadmium Complex with the Bacterial Trihydroxamate Siderophore Desferrioxamine B at Seawater Ionic Strength

    NASA Astrophysics Data System (ADS)

    Christenson, E. A.; Schijf, J.

    2010-12-01

    The divalent transition metal cadmium occurs in seawater at ultra-trace levels. In the open ocean, dissolved Cd(II) displays a nutrient-like profile characterized by a strong gradient from low picomolar concentrations in surface waters to a mid-depth maximum of around 1 nM. Its vertical distribution is highly correlated with that of dissolved phosphate, seemingly at odds with the general perception that Cd is a very toxic element. On the other hand, in Zn-depleted waters Cd(II) has been found to replace Zn(II) or Co(II) in a functional, albeit less efficient form of carbonic anhydrase, a key enzyme enabling the assimilation of bicarbonate into organic matter. Considering these opposing roles, it is likely that phytoplankton regulates the toxicity and/or bioavailability of Cd(II) through the production of certain strong organic ligands, as it has been shown to do for example in the case of Cu(II). Siderophores are a fascinating class of organic ligands excreted by microorganisms to facilitate the acquisition of micronutrient Fe(III), preciously scarce due to its extremely low solubility in seawater. The linear trihydroxamic acid desferrioxamine B (DFOB) is naturally present in open ocean surface waters at picomolar concentrations and, because of its use as a pharmaceutical agent in the treatment of human iron overload disorders, the only purified siderophore commercially available in practicable quantities. The optimal spacing of three bidentate O-bearing functional groups along a flexible carbon frame allows the molecule to wrap around the Fe3+ ion in a polydentate heterocyclic structure that perfectly matches its ionic radius and preferred coordination. Despite its resultant exceptional affinity and selectivity for Fe3+ (β ~ 1031), DFOB also forms very stable complexes with an array of differently sized and charged cations. The only previous report on the stability constant of the Cd(II)-DFOB complex, dating from 1963, proposes a values of 108 at 0.1 M ionic

  12. Molecular Cloning and Analysis of a Putative Siderophore ABC Transporter from Staphylococcus aureus

    PubMed Central

    Morrissey, Julie A.; Cockayne, Alan; Hill, Philip J.; Williams, Paul

    2000-01-01

    From a mass-excised Staphylococcus aureus λZapII expression library, we cloned an operon encoding a novel ABC transporter with significant homology to bacterial siderophore transporter systems. The operon encodes four genes designated sstA, -B, -C, and -D encoding two putative cytoplasmic membrane proteins (sstA and sstB), an ATPase (sstC), and a membrane-bound 38-kDa lipoprotein (sstD). The sst operon is preceded by two putative Fur boxes, which indicated that expression of the sst operon was likely to be iron dependent. SstD was overexpressed in Escherichia coli, purified by Triton X-114 phase partitioning, and used to generate monospecific antisera in rats. Immunoblotting studies located SstD in the membrane fraction of S. aureus and showed that expression of the lipoprotein was reduced under iron-rich growth conditions. Triton X-114 partitioning studies on isolated membranes provided additional biochemical evidence that SstD in S. aureus is a lipoprotein. Immunoreactive polypeptides of approximately 38 kDa were detected in a wide range of staphylococcal species, but no antigenic homolog was detected in Bacillus subtilis. Expression of SstD in vivo was confirmed by immunoblotting studies with S. aureus recovered from a rat intraperitoneal chamber implant model. To further define the contribution of SstD in promoting growth of S. aureus in vitro and in vivo, we used antisense RNA technology to modulate expression of SstD. Expression of antisense sstD RNA in S. aureus resulted in a decrease in SstD expression under both iron-rich and iron-restricted growth conditions. However, this reduction in SstD levels did not affect the growth of S. aureus in vitro in an iron-limited growth medium or when grown in an intraperitoneal rat chamber implant model in vivo. PMID:11035736

  13. The Evolution of Aflatoxin Biosynthesis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The biosynthesis of aflatoxin (AF) involves over 20 enzymatic reactions in a complex polyketide pathway that converts acetate and malonate to the intermediates sterigmatocystin (ST) and O-methylsterigmatocysin (OMST), the respective penultimate and ultimate precursors of AF. Although ST, OMST, and ...

  14. Pharmacodynamic Profiling of a Siderophore-Conjugated Monocarbam in Pseudomonas aeruginosa: Assessing the Risk for Resistance and Attenuated Efficacy

    PubMed Central

    Kutschke, Amy; Ehmann, David E.; Patey, Sara A.; Crandon, Jared L.; Gorseth, Elise; Miller, Alita A.; McLaughlin, Robert E.; Blinn, Christina M.; Chen, April; Nayar, Asha S.; Dangel, Brian; Tsai, Andy S.; Rooney, Michael T.; Murphy-Benenato, Kerry E.; Eakin, Ann E.; Nicolau, David P.

    2015-01-01

    The objective of this study was to investigate the risk of attenuated efficacy due to adaptive resistance for the siderophore-conjugated monocarbam SMC-3176 in Pseudomonas aeruginosa by using a pharmacokinetic/pharmacodynamic (PK/PD) approach. MICs were determined in cation-adjusted Mueller-Hinton broth (MHB) and in Chelex-treated, dialyzed MHB (CDMHB). Spontaneous resistance was assessed at 2× to 16× the MIC and the resulting mutants sequenced. Efficacy was evaluated in a neutropenic mouse thigh model at 3.13 to 400 mg/kg of body weight every 3 h for 24 h and analyzed for association with free time above the MIC (fT>MIC). To closer emulate the conditions of the in vivo model, we developed a novel assay testing activity mouse whole blood (WB). All mutations were found in genes related to iron uptake: piuA, piuC, pirR, fecI, and pvdS. Against four P. aeruginosa isolates, SMC-3176 displayed predictable efficacy corresponding to the fT>MIC using the MIC in CDMHB (R2 = 0.968 to 0.985), with stasis to 2-log kill achieved at 59.4 to 81.1%. Efficacy did not translate for P. aeruginosa isolate JJ 4-36, as the in vivo responses were inconsistent with fT>MIC exposures and implied a threshold concentration that was greater than the MIC. The results of the mouse WB assay indicated that efficacy was not predictable using the MIC for JJ 4-36 and four additional isolates, against which in vivo failures of another siderophore-conjugated β-lactam were previously reported. SMC-3176 carries a risk of attenuated efficacy in P. aeruginosa due to rapid adaptive resistance preventing entry via the siderophore-mediated iron uptake systems. Substantial in vivo testing is warranted for compounds using the siderophore approach to thoroughly screen for this in vitro-in vivo disconnect in P. aeruginosa. PMID:26438502

  15. (-)-Menthol biosynthesis and molecular genetics.

    PubMed

    Croteau, Rodney B; Davis, Edward M; Ringer, Kerry L; Wildung, Mark R

    2005-12-01

    (-)-Menthol is the most familiar of the monoterpenes as both a pure natural product and as the principal and characteristic constituent of the essential oil of peppermint (Mentha x piperita). In this paper, we review the biosynthesis and molecular genetics of (-)-menthol production in peppermint. In Mentha species, essential oil biosynthesis and storage is restricted to the peltate glandular trichomes (oil glands) on the aerial surfaces of the plant. A mechanical method for the isolation of metabolically functional oil glands, has provided a system for precursor feeding studies to elucidate pathway steps, as well as a highly enriched source of the relevant biosynthetic enzymes and of their corresponding transcripts with which cDNA libraries have been constructed to permit cloning and characterization of key structural genes. The biosynthesis of (-)-menthol from primary metabolism requires eight enzymatic steps, and involves the formation and subsequent cyclization of the universal monoterpene precursor geranyl diphosphate to the parent olefin (-)-(4S)-limonene as the first committed reaction of the sequence. Following hydroxylation at C3, a series of four redox transformations and an isomerization occur in a general "allylic oxidation-conjugate reduction" scheme that installs three chiral centers on the substituted cyclohexanoid ring to yield (-)-(1R, 3R, 4S)-menthol. The properties of each enzyme and gene of menthol biosynthesis are described, as are their probable evolutionary origins in primary metabolism. The organization of menthol biosynthesis is complex in involving four subcellular compartments, and regulation of the pathway appears to reside largely at the level of gene expression. Genetic engineering to up-regulate a flux-limiting step and down-regulate a side route reaction has led to improvement in the composition and yield of peppermint oil. PMID:16292524

  16. (-)-Menthol biosynthesis and molecular genetics

    NASA Astrophysics Data System (ADS)

    Croteau, Rodney B.; Davis, Edward M.; Ringer, Kerry L.; Wildung, Mark R.

    2005-12-01

    (-)-Menthol is the most familiar of the monoterpenes as both a pure natural product and as the principal and characteristic constituent of the essential oil of peppermint ( Mentha x piperita). In this paper, we review the biosynthesis and molecular genetics of (-)-menthol production in peppermint. In Mentha species, essential oil biosynthesis and storage is restricted to the peltate glandular trichomes (oil glands) on the aerial surfaces of the plant. A mechanical method for the isolation of metabolically functional oil glands, has provided a system for precursor feeding studies to elucidate pathway steps, as well as a highly enriched source of the relevant biosynthetic enzymes and of their corresponding transcripts with which cDNA libraries have been constructed to permit cloning and characterization of key structural genes. The biosynthesis of (-)-menthol from primary metabolism requires eight enzymatic steps, and involves the formation and subsequent cyclization of the universal monoterpene precursor geranyl diphosphate to the parent olefin (-)-(4 S)-limonene as the first committed reaction of the sequence. Following hydroxylation at C3, a series of four redox transformations and an isomerization occur in a general “allylic oxidation-conjugate reduction” scheme that installs three chiral centers on the substituted cyclohexanoid ring to yield (-)-(1 R, 3 R, 4 S)-menthol. The properties of each enzyme and gene of menthol biosynthesis are described, as are their probable evolutionary origins in primary metabolism. The organization of menthol biosynthesis is complex in involving four subcellular compartments, and regulation of the pathway appears to reside largely at the level of gene expression. Genetic engineering to up-regulate a flux-limiting step and down-regulate a side route reaction has led to improvement in the composition and yield of peppermint oil.

  17. Presence of the siderophores pyoverdine and pyochelin in the extracellular medium reduces toxic metal accumulation in Pseudomonas aeruginosa and increases bacterial metal tolerance.

    PubMed

    Braud, Armelle; Geoffroy, Valérie; Hoegy, Françoise; Mislin, Gaëtan L A; Schalk, Isabelle J

    2010-06-01

    In order to get access to iron, Pseudomonas aeruginosa strain PAO1 produces two major siderophores pyoverdine (PVD) and pyochelin (PCH). Both siderophores are able to chelate many other metals in addition to iron. However, despite this property, only iron is transported efficiently into the bacteria by the PVD and PCH uptake pathways. Growth studies with P. aeruginosa strains showed a lower sensitivity to toxic metals for the siderophore-producing strain than for the mutants unable to produce siderophores. Moreover, addition of PVD or PCH to the growth medium of a siderophore-deficient strain considerably reduced the toxicity of toxic metals present at concentrations of 100 µM in iron-limited and iron-supplemented growth conditions. Measurement by Inductively Coupled Plasma-Atomic Emission Spectrometry of the concentration of metals present in bacteria incubated with metals in the presence or absence of PVD or PCH indicated that both siderophores were able to sequester metals from the extracellular medium of the bacteria, decreasing metal diffusion into the bacteria. Pyoverdine was able to sequester Al(3+) , Co(2+) , Cu(2+) , Eu(3+) , Ni(2+) , Pb(2+) , Tb(3+) and Zn(2+) from the extracellular medium, and PCH, Al(3+) , Co(2+) , Cu(2+) , Ni(2+) , Pb(2+) and Zn(2+) . Moreover, the presence of 100 µM Cu(2+) and Ni(2+) increased PVD production by 290% and 380%, respectively, in a medium supplemented with iron. All these data suggest that PVD and PCH may contribute to P. aeruginosa resistance to heavy metals. PMID:23766115

  18. A Transmissible Plasmid-Borne Pathogenicity Island Confers Piscibactin Biosynthesis in the Fish Pathogen Photobacterium damselae subsp. piscicida.

    PubMed

    Osorio, Carlos R; Rivas, Amable J; Balado, Miguel; Fuentes-Monteverde, Juan Carlos; Rodríguez, Jaime; Jiménez, Carlos; Lemos, Manuel L; Waldor, Matthew K

    2015-09-01

    The fish pathogen Photobacterium damselae subsp. piscicida produces the siderophore piscibactin. A gene cluster that resembles the Yersinia high-pathogenicity island (HPI) encodes piscibactin biosynthesis. Here, we report that this HPI-like cluster is part of a hitherto-uncharacterized 68-kb plasmid dubbed pPHDP70. This plasmid lacks homologs of genes that mediate conjugation, but we found that it could be transferred at low frequencies from P. damselae subsp. piscicida to a mollusk pathogenic Vibrio alginolyticus strain and to other Gram-negative bacteria, likely dependent on the conjugative functions of the coresident plasmid pPHDP60. Following its conjugative transfer, pPHDP70 restored the capacity of a vibrioferrin mutant of V. alginolyticus to grow under low-iron conditions, and piscibactin became detectable in its supernatant. Thus, pPHDP70 appears to harbor all the genes required for piscibactin biosynthesis and transport. P. damselae subsp. piscicida strains cured of pPHDP70 no longer produced piscibactin, had impaired growth under iron-limited conditions, and exhibited markedly decreased virulence in fish. Collectively, our findings highlight the importance of pPHDP70, with its capacity for piscibactin-mediated iron acquisition, in the virulence of P. damselae subsp. piscicida. Horizontal transmission of this plasmid-borne piscibactin synthesis gene cluster in the marine environment may facilitate the emergence of new pathogens. PMID:26092457

  19. A Transmissible Plasmid-Borne Pathogenicity Island Confers Piscibactin Biosynthesis in the Fish Pathogen Photobacterium damselae subsp. piscicida

    PubMed Central

    Rivas, Amable J.; Balado, Miguel; Fuentes-Monteverde, Juan Carlos; Rodríguez, Jaime; Jiménez, Carlos; Lemos, Manuel L.; Waldor, Matthew K.

    2015-01-01

    The fish pathogen Photobacterium damselae subsp. piscicida produces the siderophore piscibactin. A gene cluster that resembles the Yersinia high-pathogenicity island (HPI) encodes piscibactin biosynthesis. Here, we report that this HPI-like cluster is part of a hitherto-uncharacterized 68-kb plasmid dubbed pPHDP70. This plasmid lacks homologs of genes that mediate conjugation, but we found that it could be transferred at low frequencies from P. damselae subsp. piscicida to a mollusk pathogenic Vibrio alginolyticus strain and to other Gram-negative bacteria, likely dependent on the conjugative functions of the coresident plasmid pPHDP60. Following its conjugative transfer, pPHDP70 restored the capacity of a vibrioferrin mutant of V. alginolyticus to grow under low-iron conditions, and piscibactin became detectable in its supernatant. Thus, pPHDP70 appears to harbor all the genes required for piscibactin biosynthesis and transport. P. damselae subsp. piscicida strains cured of pPHDP70 no longer produced piscibactin, had impaired growth under iron-limited conditions, and exhibited markedly decreased virulence in fish. Collectively, our findings highlight the importance of pPHDP70, with its capacity for piscibactin-mediated iron acquisition, in the virulence of P. damselae subsp. piscicida. Horizontal transmission of this plasmid-borne piscibactin synthesis gene cluster in the marine environment may facilitate the emergence of new pathogens. PMID:26092457

  20. Role of the phosphopantetheinyltransferase enzyme, PswP, in the biosynthesis of antimicrobial secondary metabolites by Serratia marcescens Db10

    PubMed Central

    Gerc, Amy J.; Stanley-Wall, Nicola R.

    2014-01-01

    Phosphopantetheinyltransferase (PPTase) enzymes fulfil essential roles in primary and secondary metabolism in prokaryotes, archaea and eukaryotes. PPTase enzymes catalyse the essential modification of the carrier protein domain of fatty acid synthases, polyketide synthases (PKSs) and non-ribosomal peptide synthetases (NRPSs). In bacteria and fungi, NRPS and PKS enzymes are often responsible for the biosynthesis of secondary metabolites with clinically relevant properties; these secondary metabolites include a variety of antimicrobial peptides. We have previously shown that in the Gram-negative bacterium Serratia marcescens Db10, the PPTase enzyme PswP is essential for the biosynthesis of an NRPS-PKS dependent antibiotic called althiomycin. In this work we utilize bioinformatic analyses to classify PswP as belonging to the F/KES subfamily of Sfp type PPTases and to putatively identify additional NRPS substrates of PswP, in addition to the althiomycin NRPS-PKS, in Ser. marcescens Db10. We show that PswP is required for the production of three diffusible metabolites by this organism, each possessing antimicrobial activity against Staphylococcus aureus. Genetic analyses identify the three metabolites as althiomycin, serrawettin W2 and an as-yet-uncharacterized siderophore, which may be related to enterobactin. Our results highlight the use of an individual PPTase enzyme in multiple biosynthetic pathways, each contributing to the ability of Ser. marcescens to inhibit competitor bacteria by the production of antimicrobial secondary metabolites. PMID:24847000

  1. Pyochelin enantiomers and their outer-membrane siderophore transporters in fluorescent pseudomonads: structural bases for unique enantiospecific recognition.

    PubMed

    Brillet, Karl; Reimmann, Cornelia; Mislin, Gaëtan L A; Noël, Sabrina; Rognan, Didier; Schalk, Isabelle J; Cobessi, David

    2011-10-19

    Pyochelin (Pch) and enantiopyochelin (EPch) are enantiomeric siderophores, with three chiral centers, produced under iron limitation conditions by Pseudomonas aeruginosa and Pseudomonas fluorescens , respectively. After iron chelation in the extracellular medium, Pch-Fe and EPch-Fe are recognized and transported by their specific outer-membrane transporters: FptA in P. aeruginosa and FetA in P. fluorescens . Structural analysis of FetA-EPch-Fe and FptA-Pch-Fe, combined with mutagenesis and docking studies revealed the structural basis of the stereospecific recognition of these enantiomers by their respective transporters. Whereas FetA and FptA have a low sequence identity but high structural homology, the Pch and EPch binding pockets do not share any structural homology, but display similar physicochemical properties. The stereospecific recognition of both enantiomers by their corresponding transporters is imposed by the configuration of the siderophore's C4'' and C2'' chiral centers. This recognition involves specific hydrogen bonds between the Arg91 guanidinium group and EPch-Fe for FetA and between the Leu117-Leu116 main chain and Pch-Fe for FptA. FetA and FptA are the first membrane receptors to be structurally described with opposite binding enantioselectivities for their ligands, giving insights into the structural basis of their enantiospecificity. PMID:21902256

  2. Gibberellin biosynthesis in Gibberlla fujikuroi

    SciTech Connect

    Johnson, S.W.; Coolbaugh, R.C. )

    1989-04-01

    Gibberellins (GAs) are a group of plant growth hormones which were first isolated from the fungus Gibberella fujikuori. We have examined the biosynthesis of GAs in this fungus in liquid cultures using HPLC followed by GC-MS. Furthermore we have used cell-free enzyme extracts with {sup 14}C-labeled intermediates to examine the regulation of specific parts of the biosynthetic pathway. GA{sub 3} is the predominant GA in well aerated cultures. GA{sub 4} and GA{sub 7}, intermediates in GA{sub 3} biosynthesis, accumulate in cultures with low levels of dissolved oxygen, but are not detectable in more aerated cultures. Light stimulates GA production in G. fujikuroi cultures grown from young stock. Cell-free enzyme studies indicate that light has no effect on incorporation of mevalonic acid into kaurene, but does significantly stimulate the oxidation of kaurenoic acid.

  3. Benzodiazepine biosynthesis in Streptomyces refuineus.

    PubMed

    Hu, Yunfeng; Phelan, Vanessa; Ntai, Ioanna; Farnet, Chris M; Zazopoulos, Emmanuel; Bachmann, Brian O

    2007-06-01

    Anthramycin is a benzodiazepine alkaloid with potent antitumor and antibiotic activity produced by the thermophilic actinomycete Streptomyces refuineus sbsp. thermotolerans. In this study, the complete 32.5 kb gene cluster for the biosynthesis of anthramycin was identified by using a genome-scanning approach, and cluster boundaries were estimated via comparative genomics. A lambda-RED-mediated gene-replacement system was developed to provide supporting evidence for critical biosynthetic genes and to validate the boundaries of the proposed anthramycin gene cluster. Sequence analysis reveals that the 25 open reading frame anthramycin cluster contains genes consistent with the biosynthesis of the two halves of anthramycin: 4 methyl-3-hydroxyanthranilic acid and a "dehydroproline acrylamide" moiety. These nonproteinogenic amino acid precursors are condensed by a two-module nonribosomal peptide synthetase (NRPS) terminated by a reductase domain, consistent with the final hemiaminal oxidation state of anthramycin. PMID:17584616

  4. Lignification: Flexibility, Biosynthesis and Regulation.

    PubMed

    Zhao, Qiao

    2016-08-01

    Lignin is a complex phenolic polymer that is deposited in the secondary cell wall of all vascular plants. The evolution of lignin is considered to be a critical event during vascular plant development, because lignin provides mechanical strength, rigidity, and hydrophobicity to secondary cell walls to allow plants to grow tall and transport water and nutrients over a long distance. In recent years, great research efforts have been made to genetically alter lignin biosynthesis to improve biomass degradability for the production of second-generation biofuels. This global focus on lignin research has significantly advanced our understanding of the lignification process. Based on these advances, here I provide an overview of lignin composition, the biosynthesis pathway and its regulation. PMID:27131502

  5. Biosynthesis of enediyne antitumor antibiotics.

    PubMed

    Van Lanen, Steven G; Shen, Ben

    2008-01-01

    The enediyne polyketides are secondary metabolites isolated from a variety of Actinomycetes. All members share very potent anticancer and antibiotic activity, and prospects for the clinical application of the enediynes has been validated with the recent marketing of two enediyne derivatives as anticancer agents. The biosynthesis of these compounds is of interest because of the numerous structural features that are unique to the enediyne family. The gene cluster for five enediynes has now been cloned and sequenced, providing the foundation to understand natures' means to biosynthesize such complex, exotic molecules. Presented here is a review of the current progress in delineating the biosynthesis of the enediynes with an emphasis on the model enediyne, C-1027. PMID:18397168

  6. Synthesis and iron sequestration equilibria of novel exocyclic 3-hydroxy-2-pyridinone donor group siderophore mimics.

    PubMed

    Harrington, James M; Chittamuru, Sumathi; Dhungana, Suraj; Jacobs, Hollie K; Gopalan, Aravamudan S; Crumbliss, Alvin L

    2010-09-20

    The synthesis of a novel class of exocyclic bis- and tris-3,2-hydroxypyridinone (HOPO) chelators built on N(2) and N(3) aza-macrocyclic scaffolds and the thermodynamic solution characterization of their complexes with Fe(III) are described. The chelators for this study were prepared by reaction of either piperazine or N,N',N''-1,4,7-triazacyclononane with a novel electrophilic HOPO iminium salt in good yields. Subsequent removal of the benzyl protecting groups using HBr/acetic acid gave bis-HOPO chelators N(2)(etLH)(2) and N(2)(prLH)(2), and tris-HOPO chelator N(3)(etLH)(3) in excellent yields. Solution thermodynamic characterization of their complexes with Fe(III) was accomplished using spectrophotometric, potentiometric, and electrospray ionization-mass spectrometry (ESI-MS) methods. The pK(a)'s of N(2)(etLH)(2), N(2)(prLH)(2), and N(3)(etLH)(3), were determined spectrophotometrically and potentiometrically. The Fe(III) complex stability constants for the tetradentate N(2)(etLH)(2) and N(2)(prLH)(2), and hexadentate N(3)(etLH)(3), were measured by spectrophotometric and potentiometric titration, and by competition with ethylenediaminetetraacetic acid (EDTA). N(3)(etLH)(3) forms a 1:1 complex with Fe(III) with log β(110) = 27.34 ± 0.04. N(2)(prLH)(2) forms a 3:2 L:Fe complex with Fe(III) where log β(230) = 60.46 ± 0.04 and log β(110) = 20.39 ± 0.02. While N(2)(etLH)(2) also forms a 3:2 L:Fe complex with Fe(III), solubility problems precluded determining log β(230); log β(110) was found to be 20.45 ± 0.04. The pFe values of 26.5 for N(3)(etLH)(3) and 24.78 for N(2)(prLH)(2) are comparable to other siderophore molecules used in the treatment of iron overload, suggesting that these hydroxypyridinone ligands may be useful in the development of new chelation therapy agents. PMID:20715813

  7. Microbial mobilization of cesium from illite: Role of organic acids and siderophores

    NASA Astrophysics Data System (ADS)

    Hazotte, Alice; Peron, Olivier; Abdelouas, Abdesselam; Lebeau, Thierry

    2015-04-01

    Understanding the behavior of cesium (Cs) in soils and geological formations is interesting in the context of nuclear accidents and nuclear waste disposals. Indeed, this radionuclide with a 30-years half-life can contaminate crops and more generally the food chain. Cs with properties similar to potassium is known to be strongly accumulated in the clays of upper soil horizons. While excavation of contaminated soil cannot be feasible for the whole contaminated surfaces (huge volumes to be cleaned-up), in situ methods could provide a sustainable and low cost solution. Phytoextraction is one of a few solutions for in situ remediation of soils contaminated by trace elements and it preserves the quality of agricultural soils. However, many improvements are still needed to enhance phytoextraction effectiveness. The combination of bioaugmentation (soil inoculation with exogenous microorganisms) with phytoextraction is likely to increase the bioaccessibility of radionuclides and their accumulation in plants. The role of bacteria on soil-pollutants can be direct (direct metal complexation) and/or indirect (weathering of clays adsorbing Cs). This study aims to provide more specifically a mechanistic understanding of the bacterial mobilization of Cs from soil with the prospect of soil bioremediation. Bacterial metabolites of Pseudomonas fluorescens (ATCC 17400) were supplied to illite spiked with 0.1 and 1 mM of Cs. Purified siderophores including pyoverdine from P. fluorescens, or the whole metabolites from the bacterial culture supernatant were compared to low molecular weight organic acids (LMWOA) (citric and oxalic acids) at 0.04 mM, or synthetic chelants, i.e., acetohydroxamic acid (AHA) and desferrioxamine mesylate (DFOM) ranging from 50 µM up to 250 µM. The release of Cs and the structural alteration of illite (release of Al, Fe and Si) were monitored. When compared to the control, no release of Cs from illite was observed with LMWOA. On the contrary, a slight release

  8. Biosynthesis of Ochratoxin A1

    PubMed Central

    Searcy, J. W.; Davis, N. D.; Diener, U. L.

    1969-01-01

    Biosynthesis of ochratoxin A by Aspergillus ochraceus Wilh. was investigated by radiolabeling experiments in which phenylalanine-1-14C and sodium acetate-2-14C were supplied to the fungus in sucrose-yeast extract medium. Results showed that phenylalanine was incorporated unaltered into the phenylalanine moiety of ochratoxin A, whereas the isocoumarin moiety of ochratoxin A was mostly derived via acetate condensation. PMID:5369298

  9. Biosynthesis of Fungal Indole Alkaloids

    PubMed Central

    Xu, Wei; Gavia, Diego J.; Tang, Yi

    2014-01-01

    This review provides a summary of recent research advances in elucidating the biosynthesis of fungal indole alkaloids. Different strategies used to incorporate and derivatize the indole/indoline moieties in various families of fungal indole alkaloids will be discussed, including tryptophan-containing nonribosomal peptides and polyketide-nonribosomal peptide hybrids; and alkaloids derived from other indole building blocks. This review also includes discussion regarding the downstream modifications that generate chemical and structural diversity among indole alkaloids. PMID:25180619

  10. Salicylate Biosynthesis: Overexpression, Purification, and Characterization of Irp9, a Bifunctional Salicylate Synthase from Yersinia enterocolitica

    PubMed Central

    Kerbarh, Olivier; Ciulli, Alessio; Howard, Nigel I.; Abell, Chris

    2005-01-01

    In some bacteria, salicylate is synthesized using the enzymes isochorismate synthase and isochorismate pyruvate lyase. In contrast, gene inactivation and complementation experiments with Yersinia enterocolitica suggest the synthesis of salicylate in the biosynthesis of the siderophore yersiniabactin involves a single protein, Irp9, which converts chorismate directly into salicylate. In the present study, Irp9 was for the first time heterologously expressed in Escherichia coli as a hexahistidine fusion protein, purified to near homogeneity, and characterized biochemically. The recombinant protein was found to be a dimer, each subunit of which has a molecular mass of 50 kDa. Enzyme assays, reverse-phase high-pressure liquid chromatography and 1H nuclear magnetic resonance (NMR) spectroscopic analyses confirmed that Irp9 is a salicylate synthase and converts chorismate to salicylate with a Km for chorismate of 4.2 μM and a kcat of 8 min−1. The reaction was shown to proceed through the intermediate isochorismate, which was detected directly using 1H NMR spectroscopy. PMID:16030197

  11. Accurate Mass MS/MS/MS Analysis of Siderophores Ferrioxamine B and E1 by Collision-Induced Dissociation Electrospray Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Sidebottom, Ashley M.; Karty, Jonathan A.; Carlson, Erin E.

    2015-11-01

    Siderophores are bacterially secreted, small molecule iron chelators that facilitate the binding of insoluble iron (III) for reuptake and use in various biological processes. These compounds are classified by their iron (III) binding geometry, as dictated by subunit composition and include groups such as the trihydroxamates (hexadentate ligand) and catecholates (bidentate). Small modifications to the core structure such as acetylation, lipid tail addition, or cyclization, make facile characterization of new siderophores difficult by molecular ion detection alone (MS1). We have expanded upon previous fragmentation-directed studies using electrospray ionization collision-induced dissociation tandem mass spectrometry (ESI-CID-MS/MS/MS) and identified diagnostic MS3 features from the trihydroxamate siderophore class for ferrioxamine B and E1 by accurate mass. Diagnostic features for MS3 include C-C, C-N, amide, and oxime cleavage events with proposed losses of water and -CO from the iron (III) coordination sites. These insights will facilitate the discovery of novel trihydroxamate siderophores from complex sample matrices.

  12. Identification of extracellular siderophores and a related peptide from the endophytic fungus Epichloë festucae in culture and endophyte-infected Lolium perenne

    PubMed Central

    Koulman, Albert; Lee, T. Verne; Fraser, Karl; Johnson, Linda; Arcus, Vickery; Lott, J. Shaun; Rasmussen, Susanne; Lane, Geoffrey

    2012-01-01

    A number of genes encoding non-ribosomal peptide synthetases (NRPSs) have been identified in fungi of Epichloë/Neotyphodium species, endophytes of Pooid grasses, including sidN, putatively encoding a ferrichrome siderophore-synthesizing NRPS. Targeted gene replacement and complementation of sidN in Epichloë festucae has established that extracellular siderophore epichloënin A is the major product of the SidN enzyme complex (Johnson et al., 2007a). We report here high resolution mass spectrometric fragmentation experiments and NMR analysis of an isolated fraction establishing that epichloënin A is a siderophore of the ferrichrome family, comprising a cyclic sequence of four glycines, a glutamine and three Nδ-trans-anhydromevalonyl–Nδ-hydroxyornithine (AMHO) moieties. Epichloënin A is unusual among ferrichrome siderophores in comprising an octapeptide rather than hexapeptide sequence, and in incorporating a glutamine residue. During this investigation we have established that desferrichrome siderophores with pendant trans-AMHO groups can be distinguished from those with pendant cis-AMHO groups by the characteristic neutral loss of an hydroxyornithine moiety in the MS/MS spectrum. A minor component, epichloënin B, has been characterized as the triglycine variant by mass spectrometry. A peptide characterized by mass spectrometry as the putative deoxygenation product, epichloëamide has been detected together with ferriepichloënin A in guttation fluid from ryegrass (Lolium perenne) plants infected with wild-type E. festucae, but not in plants infected with the ΔsidN mutant strain, and also detected at trace levels in wild-type E. festucae fungal culture. PMID:22196939

  13. Bacterial siderophores that evade or overwhelm lipocalin 2 induce hypoxia inducible factor 1α and proinflammatory cytokine secretion in cultured respiratory epithelial cells.

    PubMed

    Holden, Victoria I; Lenio, Steven; Kuick, Rork; Ramakrishnan, Sadeesh K; Shah, Yatrik M; Bachman, Michael A

    2014-09-01

    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. Furthermore, the combination of Ent and Lcn2 (Ent+Lcn2) leads to enhanced secretion of interleukin-8 (IL-8) compared to that induced by either stimulus alone. Modified or structurally distinct siderophores, including yersiniabactin (Ybt) and glycosylated Ent (GlyEnt, or salmochelin), deliver iron to bacteria despite the presence of Lcn2. We hypothesized that the robust immune response to Ent and Lcn2 requires iron chelation rather than the Ent+Lcn2 complex itself and also can be stimulated by Lcn2-evasive siderophores. To test this hypothesis, cultured respiratory epithelial cells were stimulated with combinations of purified siderophores and Lcn2 and analyzed by gene expression microarrays, quantitative PCR, and cytokine immunoassays. Ent caused HIF-1α protein stabilization, induced the expression of genes regulated by hypoxia-inducible factor 1α (HIF-1α), and repressed genes involved in cell cycle and DNA replication, whereas Lcn2 induced expression of proinflammatory cytokines. Iron chelation by excess Ent or Ybt significantly increased Lcn2-induced secretion of IL-8, IL-6, and CCL20. Stabilization of HIF-1α was sufficient to enhance Lcn2-induced IL-6 secretion. These data indicate that respiratory epithelial cells can respond to bacterial siderophores that evade or overwhelm Lcn2 binding by increasing proinflammatory cytokine production. PMID:24980968

  14. Bacterial ability in AsIII oxidation and AsV reduction: Relation to arsenic tolerance, P uptake, and siderophore production.

    PubMed

    Ghosh, Piyasa; Rathinasabapathi, Bala; Teplitski, Max; Ma, Lena Q

    2015-11-01

    The relationship between bacterial ability in arsenic transformation, siderophore production, and P uptake was investigated using six arsenic-resistant bacteria isolated from the rhizosphere of arsenic-hyperaccumulator Pteris vittata. Bacterial strains of PG5 and 12 were better arsenite (AsIII) oxidizers (31-46 vs. 6.2-21% of 1 mM AsIII) whereas PG 6, 9, 10 and 16 were better arsenate (AsV) reducers (58-95 vs. 7.5-46% of 1 mM AsV). Increase in AsV concentration from 0 to 1 mM induced 3.0-8.4 times more P uptake by bacteria but increase in P concentration from 0.1 to 1 mM reduced AsV uptake by 17-71%, indicating that P and AsV were taken up by P transporters. Bacteria producing more siderophores (PG5 and 12; >73 μM equiv) showed greater AsIII oxidation and AsIII resistance than those producing less siderophore (PG 6, 9, 10 and 16; <23 μM equiv). This observation was further supported by results obtained from mutants of Pseudomonas fluorescens impaired in siderophore production, as they were 23-25% less tolerant to AsIII than the wild-type. Arsenic-resistant bacteria increased their arsenic tolerance by retaining less arsenic in cells via efficient AsIII oxidation and AsV reduction, which were impacted by P uptake and siderophore production. PMID:25576133

  15. Microbial Siderophores Exert a Subtle Role in Arabidopsis during Infection by Manipulating the Immune Response and the Iron Status1[W

    PubMed Central

    Dellagi, Alia; Segond, Diego; Rigault, Martine; Fagard, Mathilde; Simon, Clara; Saindrenan, Patrick; Expert, Dominique

    2009-01-01

    Siderophores (ferric ion chelators) are secreted by organisms in response to iron deficiency. The pathogenic enterobacterium Erwinia chrysanthemi produces two siderophores, achromobactin and chrysobactin (CB), which are required for systemic dissemination in host plants. Previous studies have shown that CB is produced in planta and can trigger the up-regulation of the plant ferritin gene AtFER1. To further investigate the function of CB during pathogenesis, we analyzed its effect in Arabidopsis (Arabidopsis thaliana) plants following leaf infiltration. CB activates the salicylic acid (SA)-mediated signaling pathway, while the CB ferric complex is ineffective, suggesting that the elicitor activity of this siderophore is due to its iron-binding property. We confirmed this hypothesis by testing the effect of siderophores structurally unrelated to CB, including deferrioxamine. There was no activation of SA-dependent defense in plants grown under iron deficiency before CB treatment. Transcriptional analysis of the genes encoding the root ferrous ion transporter and ferric chelate reductase, and determination of the activity of this enzyme in response to CB or deferrioxamine, showed that these compounds induce a leaf-to-root iron deficiency signal. This root response as well as ferritin gene up-regulation in the leaf were not compromised in a SA-deficient mutant line. Using the Arabidopsis-E. chrysanthemi pathosystem, we have shown that CB promotes bacterial growth in planta and can modulate plant defenses through an antagonistic mechanism between SA and jasmonic acid signaling cascades. Collectively, these data reveal a new link between two processes mediated by SA and iron in response to microbial siderophores. PMID:19448037

  16. Tetrahydrobiopterin biosynthesis, regeneration and functions.

    PubMed Central

    Thöny, B; Auerbach, G; Blau, N

    2000-01-01

    Tetrahydrobiopterin (BH(4)) cofactor is essential for various processes, and is present in probably every cell or tissue of higher organisms. BH(4) is required for various enzyme activities, and for less defined functions at the cellular level. The pathway for the de novo biosynthesis of BH(4) from GTP involves GTP cyclohydrolase I, 6-pyruvoyl-tetrahydropterin synthase and sepiapterin reductase. Cofactor regeneration requires pterin-4a-carbinolamine dehydratase and dihydropteridine reductase. Based on gene cloning, recombinant expression, mutagenesis studies, structural analysis of crystals and NMR studies, reaction mechanisms for the biosynthetic and recycling enzymes were proposed. With regard to the regulation of cofactor biosynthesis, the major controlling point is GTP cyclohydrolase I, the expression of which may be under the control of cytokine induction. In the liver at least, activity is inhibited by BH(4), but stimulated by phenylalanine through the GTP cyclohydrolase I feedback regulatory protein. The enzymes that depend on BH(4) are the phenylalanine, tyrosine and tryptophan hydroxylases, the latter two being the rate-limiting enzymes for catecholamine and 5-hydroxytryptamine (serotonin) biosynthesis, all NO synthase isoforms and the glyceryl-ether mono-oxygenase. On a cellular level, BH(4) has been found to be a growth or proliferation factor for Crithidia fasciculata, haemopoietic cells and various mammalian cell lines. In the nervous system, BH(4) is a self-protecting factor for NO, or a general neuroprotecting factor via the NO synthase pathway, and has neurotransmitter-releasing function. With regard to human disease, BH(4) deficiency due to autosomal recessive mutations in all enzymes (except sepiapterin reductase) have been described as a cause of hyperphenylalaninaemia. Furthermore, several neurological diseases, including Dopa-responsive dystonia, but also Alzheimer's disease, Parkinson's disease, autism and depression, have been suggested to be

  17. Evolution of rosmarinic acid biosynthesis.

    PubMed

    Petersen, Maike; Abdullah, Yana; Benner, Johannes; Eberle, David; Gehlen, Katja; Hücherig, Stephanie; Janiak, Verena; Kim, Kyung Hee; Sander, Marion; Weitzel, Corinna; Wolters, Stefan

    2009-01-01

    Rosmarinic acid and chlorogenic acid are caffeic acid esters widely found in the plant kingdom and presumably accumulated as defense compounds. In a survey, more than 240 plant species have been screened for the presence of rosmarinic and chlorogenic acids. Several rosmarinic acid-containing species have been detected. The rosmarinic acid accumulation in species of the Marantaceae has not been known before. Rosmarinic acid is found in hornworts, in the fern family Blechnaceae and in species of several orders of mono- and dicotyledonous angiosperms. The biosyntheses of caffeoylshikimate, chlorogenic acid and rosmarinic acid use 4-coumaroyl-CoA from the general phenylpropanoid pathway as hydroxycinnamoyl donor. The hydroxycinnamoyl acceptor substrate comes from the shikimate pathway: shikimic acid, quinic acid and hydroxyphenyllactic acid derived from l-tyrosine. Similar steps are involved in the biosyntheses of rosmarinic, chlorogenic and caffeoylshikimic acids: the transfer of the 4-coumaroyl moiety to an acceptor molecule by a hydroxycinnamoyltransferase from the BAHD acyltransferase family and the meta-hydroxylation of the 4-coumaroyl moiety in the ester by a cytochrome P450 monooxygenase from the CYP98A family. The hydroxycinnamoyltransferases as well as the meta-hydroxylases show high sequence similarities and thus seem to be closely related. The hydroxycinnamoyltransferase and CYP98A14 from Coleus blumei (Lamiaceae) are nevertheless specific for substrates involved in RA biosynthesis showing an evolutionary diversification in phenolic ester metabolism. Our current view is that only a few enzymes had to be "invented" for rosmarinic acid biosynthesis probably on the basis of genes needed for the formation of chlorogenic and caffeoylshikimic acid while further biosynthetic steps might have been recruited from phenylpropanoid metabolism, tocopherol/plastoquinone biosynthesis and photorespiration. PMID:19560175

  18. The Biogeochemistry of Pu and U: Distribution of Radionuclides Affected by Micro-Organisms and Their Siderophores, Reductants, and Exopolymers

    SciTech Connect

    Traina, Samuel J.

    2003-06-01

    Investigations to date focused on studying the dissolution of oxides and desorption of metals by the siderophore, Desferrioxamine B (DFB), with different metal ions adsorbed onto the solids. X-ray absorption spectroscopy (XAS) was used to probe the surface structural environment of sorbed metal ions. Results indicated that while DFB effectively dissolved iron oxides with different adsorbed metals, this process was also affected by the type of the metal adsorbed. For pure hematite, samples with adsorbed metals had less dissolved Fe by DFB than the one without. Different type of metal ion seemed to have no significant effects on Fe dissolution under these experimental conditions. This result suggested that while adsorbed metals blocked available surface sites on hematite surfaces for DFB causing less Fe release, Fe dissolution by DFB from the well crystalline structure of hematite was not affected by the adsorbed metal ions.

  19. DtxR, an iron-dependent transcriptional repressor that regulates the expression of siderophore gene clusters in Thermobifida fusca.

    PubMed

    Deng, Yu; Zhang, Xiaojuan

    2015-01-01

    Thermobifida fusca is an aerobic, moderately thermophilic, filamentous soil bacterium. Iron is an essential metal involved in the vital metabolic functions in microorganisms. Thermobifida fusca was found to survive in the iron-deficiency condition and produce siderophores which synthesized to acquire iron. The iron transport was regulated by DtxR, a prototypic member of a superfamily of transition metal ion-activated transcriptional regulators. Tfu_0249 was found to be working as DtxR in T. fusca. The promoter regions of the three gene clusters related to the iron transport bound and regulated by Tfu_0249 were identified by bioinformatics analysis. The gel shift assays confirmed that Tfu_0249 bound the binding sites firmly on the upstream of three gene clusters. By comparing the binding sites of three clusters and gel shift assay, TWAGGTWAGSCTWACCTWA was found to be recognized and bound by Tfu_0249 to control iron transport in T. fusca. PMID:25673661

  20. Cellulose biosynthesis inhibitors - a multifunctional toolbox.

    PubMed

    Tateno, Mizuki; Brabham, Chad; DeBolt, Seth

    2016-01-01

    In the current review, we examine the growing number of existing Cellulose Biosynthesis Inhibitors (CBIs) and based on those that have been studied with live cell imaging we group their mechanism of action. Attention is paid to the use of CBIs as tools to ask fundamental questions about cellulose biosynthesis. PMID:26590309

  1. BIOSYNTHESIS OF NITRO COMPOUNDS I.

    PubMed Central

    Shaw, Paul D.; Wang, Nancy

    1964-01-01

    Shaw, Paul D. (University of Illinois, Urbana), and Nancy Wang. Biosynthesis of nitro compounds. I. Nitrogen and carbon requirements for the biosynthesis of β-nitropropionic acid by Penicillium atrovenetum. J. Bacteriol. 88:1629–1635. 1964.—β-Nitropropionic acid was produced by Penicillium atrovenetum when this fungus was grown on a Raulin-Thom medium in shake flasks. The nitro compound was formed in the early stages of growth, and the total amount in the medium decreased when the fungus reached the end of the log phase. When increasing amounts of nitrate were substituted for the ammonia in the growth medium, production of β-nitropropionic acid decreased. Aspartic acid did not promote the synthesis of the nitro compound unless either ammonium chloride or sodium tartrate was also added to the medium. The addition of small amounts of hydroxylamine or sodium nitrite to the Raulin-Thom medium stimulated β-nitropropionic acid production to a greater degree on a molar basis than the amount of hydroxylamine or nitrite added. The nature of possible precursors to the nitro group of β-nitropropionic acid is discussed. PMID:14240949

  2. Oleic acid biosynthesis in cyanobacteria

    SciTech Connect

    VanDusen, W.J.; Jaworski, J.G.

    1986-05-01

    The biosynthesis of fatty acids in cyanobacteria is very similar to the well characterized system found in green plants. However, the initial desaturation of stearic acid in cyanobacteria appears to represent a significant departure from plant systems in which stearoyl-ACP is the exclusive substrate for desaturation. In Anabaena variabilis, the substrate appears to be monoglucosyldiacylglycerol, a lipid not found in plants. The authors examined five different cyanobacteria to determine if the pathway in A. variabilis was generally present in other cyanobacteria. The cyanobacteria studied were A. variabilis, Chlorogloeopsis sp., Schizothrix calcicola, Anacystis marina, and Anacystis nidulans. Each were grown in liquid culture, harvested, and examined for stearoyl-ACP desaturase activity or incubated with /sup 14/CO/sub 2/. None of the cyanobacteria contained any stearoyl-ACP desaturase activity in whole homogenates or 105,000g supernatants. All were capable of incorporating /sup 14/CO/sub 2/ into monoglucosyldiacylglycerol and results from incubations of 20 min, 1 hr, 1 hr + 10 hr chase were consistent with monoglucosyldiacylglycerol serving as precursor for monogalctosyldiacylglycerol. Thus, initial evidence is consistent with oleic acid biosynthesis occurring by desaturation of stearoyl-monoglucosyldiacylglycerol in all cyanobacteria.

  3. Current aspects of auxin biosynthesis in plants.

    PubMed

    Kasahara, Hiroyuki

    2015-01-01

    Auxin is an important plant hormone essential for many aspects of plant growth and development. Indole-3-acetic acid (IAA) is the most studied auxin in plants, and its biosynthesis pathway has been investigated for over 70 years. Although the complete picture of auxin biosynthesis remains to be elucidated, remarkable progress has been made recently in understanding the mechanism of IAA biosynthesis. Genetic and biochemical studies demonstrate that IAA is mainly synthesized from l-tryptophan (Trp) via indole-3-pyruvate by two-step reactions in Arabidopsis. While IAA is also produced from Trp via indole-3-acetaldoxime in Arabidopsis, this pathway likely plays an auxiliary role in plants of the family Brassicaceae. Recent studies suggest that the Trp-independent pathway is not a major route for IAA biosynthesis, but they reveal an important role for a cytosolic indole synthase in this pathway. In this review, I summarize current views and future prospects of IAA biosynthesis research in plants. PMID:26364770

  4. Regulation of cardiolipin biosynthesis in the heart.

    PubMed

    Hatch, G M

    1996-06-21

    Cardiolipin is one of the principle phospholipids in the mammalian heart comprising as much as 15-20% of the entire phospholipid phosphorus mass of that organ. Cardiolipin is localized primarily in the mitochondria and appears to be essential for the function of several enzymes of oxidative phosphorylation. Thus, cardiolipin is essential for production of energy for the heart to beat. Cardiac cardiolipin is synthesized via the cytidine-5'-diphosphate-1,2-diacyl-sn-glycerol pathway. The properties of the four enzymes of the cytidine-5'-diphosphate-1,2-diacyl-sn-glycerol pathway have been characterized in the heart. The rate-limiting step of this pathway is catalyzed by the phosphatidic acid: cytidine-5'-triphosphate cytidylyltransferase. Several regulatory mechanisms that govern cardiolipin biosynthesis in the heart have been uncovered. Current evidence suggests that cardiolipin biosynthesis is regulated by the energy status (adenosine-5'-triphosphate and cytidine-5'-triphosphate level) of the heart. Thyroid hormone and unsaturated fatty acids may regulate cardiolipin biosynthesis at the level of three key enzymes of the cytidine-5'-diphosphate-1,2-diacyl-sn-glycerol pathway, phosphatidylglycerol phosphate synthase, phosphatidyl-glycerolphosphate phosphatase and cardiolipin synthase. Newly synthesized phosphatidic acid and phosphatidylglycerol may be preferentially utilized for cardiolipin biosynthesis in the heart. In addition, separate pools of phosphatidylglycerol, including an exogenous (extra-mitochondrial) pool not derived from de novo phosphatidylglycerol biosynthesis, may be utilized for cardiac cardiolipin biosynthesis. In several mammalian tissues a significant number of studies on polyglycerophospholipid biosynthesis have been documented, including detailed studies in the lung and liver. However, in spite of the important role of cardiolipin in the maintenance of mitochondrial function and membrane integrity, studies on the control of cardiolipin

  5. Isoprenoid Biosynthesis in Plasmodium falciparum

    PubMed Central

    Guggisberg, Ann M.; Amthor, Rachel E.

    2014-01-01

    Malaria kills nearly 1 million people each year, and the protozoan parasite Plasmodium falciparum has become increasingly resistant to current therapies. Isoprenoid synthesis via the methylerythritol phosphate (MEP) pathway represents an attractive target for the development of new antimalarials. The phosphonic acid antibiotic fosmidomycin is a specific inhibitor of isoprenoid synthesis and has been a helpful tool to outline the essential functions of isoprenoid biosynthesis in P. falciparum. Isoprenoids are a large, diverse class of hydrocarbons that function in a variety of essential cellular processes in eukaryotes. In P. falciparum, isoprenoids are used for tRNA isopentenylation and protein prenylation, as well as the synthesis of vitamin E, carotenoids, ubiquinone, and dolichols. Recently, isoprenoid synthesis in P. falciparum has been shown to be regulated by a sugar phosphatase. We outline what is known about isoprenoid function and the regulation of isoprenoid synthesis in P. falciparum, in order to identify valuable directions for future research. PMID:25217461

  6. Biosynthesis of mycobacterial phosphatidylinositol mannosides.

    PubMed Central

    Morita, Yasu S; Patterson, John H; Billman-Jacobe, Helen; McConville, Malcolm J

    2004-01-01

    All mycobacterial species, including pathogenic Mycobacterium tuberculosis, synthesize an abundant class of phosphatidylinositol mannosides (PIMs) that are essential for normal growth and viability. These glycolipids are important cell-wall and/or plasma-membrane components in their own right and can also be hyperglycosylated to form other wall components, such as lipomannan and lipoarabinomannan. We have investigated the steps involved in the biosynthesis of the major PIM species in a new M. smegmatis cell-free system. A number of apolar and polar PIM intermediates were labelled when this system was continuously labelled or pulse-chase-labelled with GDP-[3H]Man, and the glycan head groups and the acylation states of these species were determined by chemical and enzymic treatments and octyl-Sepharose chromatography respectively. These analyses showed that (1) the major apolar PIM species, acyl-PIM2, can be synthesized by at least two pathways that differ in the timing of the first acylation step, (2) early PIM intermediates containing a single mannose residue can be modified with two fatty acid residues, (3) formation of polar PIM species from acyl-PIM2 is amphomycin-sensitive, indicating that polyprenol phosphate-Man, rather than GDP-Man, is the donor for these reactions, (4) modification of acylated PIM4 with alpha1-2- or alpha1-6-linked mannose residues is probably the branch point in the biosyntheses of polar PIM and lipoarabinomannan respectively and (5) GDP strongly inhibits the synthesis of early PIM intermediates and increases the turnover of polyprenol phosphate-Man. These findings are incorporated into a revised pathway for mycobacterial PIM biosynthesis. PMID:14627436

  7. Acylphloroglucinol Biosynthesis in Strawberry Fruit.

    PubMed

    Song, Chuankui; Ring, Ludwig; Hoffmann, Thomas; Huang, Fong-Chin; Slovin, Janet; Schwab, Wilfried

    2015-11-01

    Phenolics have health-promoting properties and are a major group of metabolites in fruit crops. Through reverse genetic analysis of the functions of four ripening-related genes in the octoploid strawberry (Fragaria × ananassa), we discovered four acylphloroglucinol (APG)-glucosides as native Fragaria spp. fruit metabolites whose levels were differently regulated in the transgenic fruits. The biosynthesis of the APG aglycones was investigated by examination of the enzymatic properties of three recombinant Fragaria vesca chalcone synthase (FvCHS) proteins. CHS is involved in anthocyanin biosynthesis during ripening. The F. vesca enzymes readily catalyzed the condensation of two intermediates in branched-chain amino acid metabolism, isovaleryl-Coenzyme A (CoA) and isobutyryl-CoA, with three molecules of malonyl-CoA to form phlorisovalerophenone and phlorisobutyrophenone, respectively, and formed naringenin chalcone when 4-coumaroyl-CoA was used as starter molecule. Isovaleryl-CoA was the preferred starter substrate of FvCHS2-1. Suppression of CHS activity in both transient and stable CHS-silenced fruit resulted in a substantial decrease of APG glucosides and anthocyanins and enhanced levels of volatiles derived from branched-chain amino acids. The proposed APG pathway was confirmed by feeding isotopically labeled amino acids. Thus, Fragaria spp. plants have the capacity to synthesize pharmaceutically important APGs using dual functional CHS/(phloriso)valerophenone synthases that are expressed during fruit ripening. Duplication and adaptive evolution of CHS is the most probable scenario and might be generally applicable to other plants. The results highlight that important promiscuous gene function may be missed when annotation relies solely on in silico analysis. PMID:26169681

  8. Oxidative mobilization of cerium and uranium and enhanced release of "immobile" high field strength elements from igneous rocks in the presence of the biogenic siderophore desferrioxamine B

    NASA Astrophysics Data System (ADS)

    Kraemer, Dennis; Kopf, Sebastian; Bau, Michael

    2015-09-01

    Polyvalent trace elements such as the high field strength elements (HFSE) are commonly considered rather immobile during low-temperature water-rock interaction. Hence, they have become diagnostic tools that are widely applied in geochemical studies. We present results of batch leaching experiments focused on the mobilization of certain HFSE (Y, Zr, Hf, Th, U and rare earth elements) from mafic, intermediate and felsic igneous rocks in the presence and absence, respectively, of the siderophore desferrioxamine B (DFOB). Our data show that DFOB strongly enhances the mobility of these trace elements during low-temperature water-rock interaction. The presence of DFOB produces two distinct features in the Rare Earths and Yttrium (REY) patterns of leaching solutions, regardless of the mineralogical and chemical composition or the texture of the rock type studied. Bulk rock-normalized REY patterns of leaching solutions with DFOB show (i) a very distinct positive Ce anomaly and (ii) depletion of La and other light REY relative to the middle REY, with a concave downward pattern between La and Sm. These features are not observed in experiments with hydrochloric acid, acetic acid or deionized water. In DFOB-bearing leaching solutions Ce and U are decoupled from and selectively enriched relative to light REY and Th, respectively, due to oxidation to Ce(IV) and U(VI). Oxidation of Ce3+ and U4+ is promoted by the significantly higher stability of the Ce(IV) and U(VI) DFOB complexes as compared to the Ce(III) and U(IV) DFOB complexes. This is similar to the relationship between the Ce(IV)- and Ce(III)-pentacarbonate complexes that cause positive Ce anomalies in alkaline lakes. However, while formation of Ce(IV) carbonate complexes is confined to alkaline environments, Ce(IV) DFOB complexes may produce positive Ce anomalies even in mildly acidic and near-neutral natural waters. Siderophore-promoted dissolution processes also significantly enhance mobility of other 'immobile' HFSE

  9. The ferrichrome uptake pathway in Pseudomonas aeruginosa involves an iron release mechanism with acylation of the siderophore and recycling of the modified desferrichrome.

    PubMed

    Hannauer, Mélissa; Barda, Yaniv; Mislin, Gaëtan L A; Shanzer, Abraham; Schalk, Isabelle J

    2010-03-01

    The uptake of iron into Pseudomonas aeruginosa is mediated by two major siderophores produced by the bacterium, pyoverdine and pyochelin. The bacterium is also able of utilize several heterologous siderophores of bacterial or fungal origin. In this work, we have investigated the iron uptake in P. aeruginosa PAO1 by the heterologous ferrichrome siderophore. (55)Fe uptake assays showed that ferrichrome is transported across the outer membrane primarily (80%) by the FiuA receptor and to a lesser extent (20%) by a secondary transporter. Moreover, we demonstrate that like in the uptake of ferripyoverdine and ferripyochelin, the energy required for both pathways of ferrichrome uptake is provided by the inner membrane protein TonB1. Desferrichrome-(55)Fe uptake in P. aeruginosa was also dependent on the expression of the permease FiuB, suggesting that this protein is the inner membrane transporter of the ferrisiderophore. A biomimetic fluorescent analogue of ferrichrome, RL1194, was used in vivo to monitor the kinetics of iron release from ferrichrome in P. aeruginosa in real time. This dissociation involves acylation of ferrichrome and its biomimetic analogue RL1194 and recycling of both modified siderophores into the extracellular medium. FiuC, an N-acetyltransferase, is certainly involved in this mechanism of iron release, since its mutation abolished desferrichrome-(55)Fe uptake. The acetylated derivative reacts with iron in the extracellular medium and is able to be taken up again by the cells. All these observations are discussed in light of the current knowledge concerning ferrichrome uptake in P. aeruginosa and in Escherichia coli. PMID:20047910

  10. Alpha-keto acids are novel siderophores in the genera Proteus, Providencia, and Morganella and are produced by amino acid deaminases.

    PubMed Central

    Drechsel, H; Thieken, A; Reissbrodt, R; Jung, G; Winkelmann, G

    1993-01-01

    Growth promotion and iron transport studies revealed that certain alpha-keto acids generated by amino acid deaminases, by enterobacteria of the Proteus-Providencia-Morganella group (of the tribe Proteeae), show significant siderophore activity. Their iron-binding properties were confirmed by the chrome azurol S assay and UV spectra. These compounds form ligand-to-metal charge transfer bands in the range of 400 to 500 nm. Additional absorption bands of the enolized ligands at 500 to 700 nm are responsible for color formation. Siderophore activity was most pronounced with alpha-keto acids possessing an aromatic or heteroaromatic side chain, like phenylpyruvic acid and indolylpyruvic acid, resulting from deamination of phenylalanine and tryptophan, respectively. In addition, alpha-keto acids possessing longer nonpolar side chains, like alpha-ketoisocaproic acid or alpha-ketoisovaleric acid and even alpha-ketoadipic acid, also showed siderophore activity which was absent or negligible with smaller alpha-keto acids or those possessing polar functional groups, like pyruvic acid, alpha-ketobutyric acid, or alpha-ketoglutaric acid. The fact that deaminase-negative enterobacteria, like Escherichia coli and Salmonella spp., could not utilize alpha-keto acids supports the view that specific iron-carboxylate transport systems have evolved in members of the tribe Proteeae and are designed to recognize ferric complexes of both alpha-hydroxy acids and alpha-keto acids, of which the latter can easily be generated by L-amino acid deaminases in an amino acid-rich medium. Exogenous siderophores, like ferric hydroxamates (ferrichromes) and ferric polycarboxylates (rhizoferrin and citrate), were also utilized by members of the tribe Proteeae. Images PMID:8478334

  11. The Major Cow Milk Allergen Bos d 5 Manipulates T-Helper Cells Depending on Its Load with Siderophore-Bound Iron

    PubMed Central

    Roth-Walter, Franziska; Pacios, Luis F.; Gomez-Casado, Cristina; Hofstetter, Gerlinde; Roth, Georg A.; Singer, Josef; Diaz-Perales, Araceli; Jensen-Jarolim, Erika

    2014-01-01

    The mechanisms of allergic sensitization to milk are still elusive. The major allergen Bos d 5 belongs to the lipocalin-family and thus is able to transport numerous ligands. In this study we investigated its ability to bind to iron-siderophore complexes and tested the immune-modulatory properties of Bos d 5 in either forms. Structural and in silico docking analysis of Bos d 5 revealed that Bos d 5 is able to bind to iron via catechol-based flavonoids (quercetin, myricetin, luteolin) that act as siderophores as confirmed by spectral-analysis and iron staining. Calculated dissociation constants of docking analyses were below 1 µM by virtual addition of iron. When incubated with human peripheral blood mononuclear cells (PBMCs), only the apo-form of Bos d 5 led to an increase of CD4+positive cells and significantly elevated IL13 and IFNγ-levels. In contrast, holo-Bos d 5 decreased numbers of CD4 expressing cells and induced apoptosis. Taken together, our data give evidence that Bos d 5 is capable of binding iron via siderophores. Moreover, our data support for the first time the notion that the form of application (apo- or holo-form) is decisive for the subsequent immune response. The apo-form promotes Th2 cells and inflammation, whereas the holo-form appears to be immunosuppressive. PMID:25117976

  12. Advances in Understanding the Biosynthesis of Fumonisins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fumonisins are a group of economically important mycotoxins that are derived polyketides. Since the cloning of the fumonisin polyketide synthase (PKS) gene from Fusarium verticillioides in 1999, significant advances have been made in understanding the molecular mechanisms for fumonisin biosynthesis...

  13. Sterols of the fungi - Distribution and biosynthesis.

    NASA Technical Reports Server (NTRS)

    Weete, J. D.

    1973-01-01

    The importance of sterols in the growth and reproduction in fungi is becoming increasingly apparent. This article concerns the composition and biosynthesis of ergosterol in these organisms. Comparison to plant and animal sterol formation are made.

  14. Sterols of the fungi - Distribution and biosynthesis

    NASA Technical Reports Server (NTRS)

    Weete, J. D.

    1973-01-01

    The importance of sterols in the growth and reproduction in fungi is becoming increasingly apparent. This article concerns the composition and biosynthesis of ergosterol in these organisms. Comparison to plant and animal sterol formation are made.

  15. Carotenoid Biosynthesis in Daucus carota.

    PubMed

    Simpson, Kevin; Cerda, Ariel; Stange, Claudia

    2016-01-01

    Carrot (Daucus carota) is one of the most important vegetable cultivated worldwide and the main source of dietary provitamin A. Contrary to other plants, almost all carrot varieties accumulate massive amounts of carotenoids in the root, resulting in a wide variety of colors, including those with purple, yellow, white, red and orange roots. During the first weeks of development the root, grown in darkness, is thin and pale and devoid of carotenoids. At the second month, the thickening of the root and the accumulation of carotenoids begins, and it reaches its highest level at 3 months of development. This normal root thickening and carotenoid accumulation can be completely altered when roots are grown in light, in which chromoplasts differentiation is redirected to chloroplasts development in accordance with an altered carotenoid profile. Here we discuss the current evidence on the biosynthesis of carotenoid in carrot roots in response to environmental cues that has contributed to our understanding of the mechanism that regulates the accumulation of carotenoids, as well as the carotenogenic gene expression and root development in D. carota. PMID:27485223

  16. Biosynthesis of trichothecenes and apotrichothecenes.

    PubMed

    Zamir, L O; Nikolakakis, A; Sauriol, F; Mamer, O

    1999-05-01

    Fusarium culmorum produces two major trichothecenes, 3-acetyldeoxynivalenol and sambucinol, and some minor apotrichothecenes. It was desired to investigate if during their biosynthesis a C-11-keto intermediate was involved. To verify this postulate, trichodiene, a known precursor to trichothecenes, was synthesized with two deuteriums at C-11 and one at C-15. It was then fed to F. culmorum cultures, and the derived metabolites were purified and analyzed. The results ruled out the involvement of an 11-keto intermediate but revealed two novel apotrichothecenes. The characterization of their structures suggested that one of the 2-hydroxy-11alpha-apotrichothecene stereoisomers (2alpha or 2beta) could be converted to sambucinol. These apotrichothecenes were therefore synthesized labeled specifically with two deuteriums at C-4 and C-15 and fed to F. culmorum cultures. Indeed, the result established for the first time that 2alpha-hydroxy-11alpha-apotrichothecene was a precursor to sambucinol. A biosynthetic scheme for the production of trichothecenes and apotrichothecenes is described. PMID:10552458

  17. Salicylic Acid Biosynthesis and Metabolism

    PubMed Central

    Dempsey, D'Maris Amick; Vlot, A. Corina; Wildermuth, Mary C.; Klessig, Daniel F.

    2011-01-01

    Salicylic acid (SA) has been shown to regulate various aspects of growth and development; it also serves as a critical signal for activating disease resistance in Arabidopsis thaliana and other plant species. This review surveys the mechanisms involved in the biosynthesis and metabolism of this critical plant hormone. While a complete biosynthetic route has yet to be established, stressed Arabidopsis appear to synthesize SA primarily via an isochorismate-utilizing pathway in the chloroplast. A distinct pathway utilizing phenylalanine as the substrate also may contribute to SA accumulation, although to a much lesser extent. Once synthesized, free SA levels can be regulated by a variety of chemical modifications. Many of these modifications inactivate SA; however, some confer novel properties that may aid in long distance SA transport or the activation of stress responses complementary to those induced by free SA. In addition, a number of factors that directly or indirectly regulate the expression of SA biosynthetic genes or that influence the rate of SA catabolism have been identified. An integrated model, encompassing current knowledge of SA metabolism in Arabidopsis, as well as the influence other plant hormones exert on SA metabolism, is presented. PMID:22303280

  18. Reporting a new siderophore based Ca(2+) selective chemosensor that works as a staining agent in the live organism Artemia.

    PubMed

    Raju, M; Nair, Ratish R; Raval, Ishan H; Haldar, Soumya; Chatterjee, Pabitra B

    2015-11-21

    A Ca(2+)-specific chemosensor involving acyclic non-ether and non-carboxylato-type metal chelating ligands is rare. The tetradentate OONO artificial receptor, HL, possessing a sulfur-containing intermediate siderophore aeruginic acid, tethered to a rhodamine 6G based signalling unit in a single molecule has been synthesized. The fluoroionophore required excitation in the visible wavelength (510 nm) and showed highly selective and sensitive detection of Ca(2+) ions in 100% water solution in HEPES buffer at physiological pH (7.4). The probe HL, with LOD as low as 70 nM, behaves reversibly and showed nearly 17-fold enhanced selectivity for Ca(2+) over other cell abundant alkali and alkaline metal ions such as Na(+), K(+), Li(+), and Mg(2+) without any intervention. Job's plot, (1)H NMR titration and ESI-MS data provided corroborative evidence in support of 1 : 1 association between HL and Ca(2+). From a wide range of transition and heavy metal ions series, HL also binds Cu(2+). However, the use of l-cysteine removes the interference from Cu(2+) and results in highly selective detection specificity of HL for Ca(2+). As a reversible "off-on-off" fluorescent chemosensor, it is possible to detect Ca(2+) at as low as 5 μM in the midgut region of the gastrointestinal tract of the live animal Artemia, a brine shrimp. PMID:26460620

  19. Disruption of Transporters Affiliated with Enantio-Pyochelin Biosynthesis Gene Cluster of Pseudomonas protegens Pf-5 Has Pleiotropic Effects

    PubMed Central

    Lim, Chee Kent; Penesyan, Anahit; Hassan, Karl A.; Loper, Joyce E.; Paulsen, Ian T.

    2016-01-01

    Pseudomonas protegens Pf-5 (formerly Pseudomonas fluorescens) is a biocontrol bacterium that produces the siderophore enantio-pyochelin under conditions of iron starvation in a process that is often accompanied by the secretion of its biosynthesis intermediates, salicylic acid and dihydroaeruginoic acid. In this study, we investigated whether several transporters that are encoded by genes within or adjacent to the enantio-pyochelin biosynthetic cluster, serve as efflux systems for enantio-pyochelin and/or its intermediates. In addition, we determined whether these transporters have broad substrates range specificity using a Phenotype Microarray system. Intriguingly, knockouts of the pchH and fetF transporter genes resulted in mutant strains that secrete higher levels of enantio-pyochelin as well as its intermediates salicylic acid and dihydroaeruginoic acid. Analyses of these mutants did not indicate significant change in transcription of biosynthetic genes involved in enantio-pyochelin production. In contrast, the deletion mutant of PFL_3504 resulted in reduced transcription of the biosynthetic genes as well as decreased dihydroaeruginoic acid concentrations in the culture supernatant, which could either point to regulation of gene expression by the transporter or its role in dihydroaeruginoic acid transport. Disruption of each of the transporters resulted in altered stress and/or chemical resistance profile of Pf-5, which may reflect that these transporters could have specificity for rather a broad range of substrates. PMID:27442435

  20. Disruption of Transporters Affiliated with Enantio-Pyochelin Biosynthesis Gene Cluster of Pseudomonas protegens Pf-5 Has Pleiotropic Effects.

    PubMed

    Lim, Chee Kent; Penesyan, Anahit; Hassan, Karl A; Loper, Joyce E; Paulsen, Ian T

    2016-01-01

    Pseudomonas protegens Pf-5 (formerly Pseudomonas fluorescens) is a biocontrol bacterium that produces the siderophore enantio-pyochelin under conditions of iron starvation in a process that is often accompanied by the secretion of its biosynthesis intermediates, salicylic acid and dihydroaeruginoic acid. In this study, we investigated whether several transporters that are encoded by genes within or adjacent to the enantio-pyochelin biosynthetic cluster, serve as efflux systems for enantio-pyochelin and/or its intermediates. In addition, we determined whether these transporters have broad substrates range specificity using a Phenotype Microarray system. Intriguingly, knockouts of the pchH and fetF transporter genes resulted in mutant strains that secrete higher levels of enantio-pyochelin as well as its intermediates salicylic acid and dihydroaeruginoic acid. Analyses of these mutants did not indicate significant change in transcription of biosynthetic genes involved in enantio-pyochelin production. In contrast, the deletion mutant of PFL_3504 resulted in reduced transcription of the biosynthetic genes as well as decreased dihydroaeruginoic acid concentrations in the culture supernatant, which could either point to regulation of gene expression by the transporter or its role in dihydroaeruginoic acid transport. Disruption of each of the transporters resulted in altered stress and/or chemical resistance profile of Pf-5, which may reflect that these transporters could have specificity for rather a broad range of substrates. PMID:27442435

  1. Gene and protein expression profiles of Shewanella oneidensis during anaerobic growth with different electron acceptors.

    SciTech Connect

    Beliaev, A. S.; Thompson, D. K.; Khare, T.; Lim, H.; Brandt, C. C.; Li, G.; Murray, A. E.; Heidelberg, J. F.; Giometti, C. S.; Yates, J., III; Nealson, K. H.; Tiedje, J. M.; Zhou, J.; Biosciences Division; ORNL; Scripps Research Inst.; Michigan State Univ.; The Inst. for Genomic Research; Jet Propulsion Laboratory; California Inst. of Tech.

    2002-01-01

    Changes in mRNA and protein expression profiles of Shewanella oneidenesis MR-1 during switch from aerobic to fumarate-, Fe(III)-, or nitrate-reducing conditions were examined using DNA microarrays and two-dimensional polyacrylamide gel electrophoresis (2-D PAGE). In response to changes in growth conditions, 121 of the 691 arrayed genes displayed at least a two-fold difference in transcript abundance as determined by microarray analysis. Genes involved in aerobic respiration encoding cytochrome c and d oxidases and TCA cycle enzymes were repressed under anaerobic conditions. Genes induced during anaerobic respiration included those involved in cofactor biosynthesis and assembly (moaACE, ccmHF, nosD, cysG), substrate transport (cysUP, cysTWA, dcuB), and anaerobic energy metabolism (dmsAB, psrC, pshA, hyaABC, hydA). Transcription of genes encoding a periplasmic nitrate reductase (napBHGA), cytochrome c{sub 552}, and prismane was elevated 8- to 56-fold in response to the presence of nitrate, while cymA, ifcA, and frdA were specifically induced three- to eightfold under fumarate-reducing conditions. The mRNA levels for two oxidoreductase-like genes of unknown function and several cell envelope genes involved in multidrug resistance increased two- to fivefold specifically under Fe(III)-reducing conditions. Analysis of protein expression profiles under aerobic and anaerobic conditions revealed 14 protein spots that showed significant differences in abundance on 2-D gels. Protein identification by mass spectrometry indicated that the expression of prismane, dihydrolipoamide succinyltransferase, and alcaligin siderophore biosynthesis protein correlated with the microarray data.

  2. Veratrole biosynthesis in white campion.

    PubMed

    Akhtar, Tariq A; Pichersky, Eran

    2013-05-01

    White campion (Silene latifolia) is a dioecious plant that emits 1,2-dimethoxybenzene (veratrole), a potent pollinator attractant to the nocturnal moth Hadena bicruris. Little is known about veratrole biosynthesis, although methylation of 2-methoxyphenol (guaiacol), another volatile emitted from white campion flowers, has been proposed. Here, we explore the biosynthetic route to veratrole. Feeding white campion flowers with [(13)C9]l-phenylalanine increased guaiacol and veratrole emission, and a significant portion of these volatile molecules contained the stable isotope. When white campion flowers were treated with the phenylalanine ammonia lyase inhibitor 2-aminoindan-2-phosphonic acid, guaiacol and veratrole levels were reduced by 50% and 63%, respectively. Feeding with benzoic acid (BA) or salicylic acid (SA) increased veratrole emission 2-fold, while [(2)H5]BA and [(2)H6]SA feeding indicated that the benzene ring of both guaiacol and veratrole is derived from BA via SA. We further report guaiacol O-methyltransferase (GOMT) activity in the flowers of white campion. The enzyme was purified to apparent homogeneity, and the peptide sequence matched that encoded by a recently identified complementary DNA (SlGOMT1) from a white campion flower expressed sequence tag database. Screening of a small population of North American white campion plants for floral volatile emission revealed that not all plants emitted veratrole or possessed GOMT activity, and SlGOMT1 expression was only observed in veratrole emitters. Collectively these data suggest that veratrole is derived by the methylation of guaiacol, which itself originates from phenylalanine via BA and SA, and therefore implies a novel branch point of the general phenylpropanoid pathway. PMID:23547102

  3. Biosynthesis of gold nanoparticles: A green approach.

    PubMed

    Ahmed, Shakeel; Annu; Ikram, Saiqa; Yudha S, Salprima

    2016-08-01

    Nanotechnology is an immensely developing field due to its extensive range of applications in different areas of technology and science. Different types of methods are employed for synthesis of nanoparticles due to their wide applications. The conventional chemical methods have certain limitations with them either in the form of chemical contaminations during their syntheses procedures or in later applications and use of higher energy. During the last decade research have been focussed on developing simple, clean, non-toxic, cost effective and eco-friendly protocols for synthesis of nanoparticles. In order to get this objective, biosynthesis methods have been developed in order to fill this gap. The biosynthesis of nanoparticles is simple, single step, eco-friendly and a green approach. The biochemical processes in biological agents reduce the dissolved metal ions into nano metals. The various biological agents like plant tissues, fungi, bacteria, etc. are used for biosynthesis for metal nanoparticles. In this review article, we summarised recent literature on biosynthesis of gold nanoparticles which have revolutionised technique of synthesis for their applications in different fields. Due to biocompatibility of gold nanoparticles, it has find its applications in biomedical applications. The protocol and mechanism of biosynthesis of gold nanoparticles along with various applications have also been discussed. PMID:27236049

  4. Light-controlled flavonoid biosynthesis in fruits

    PubMed Central

    Zoratti, Laura; Karppinen, Katja; Luengo Escobar, Ana; Häggman, Hely; Jaakola, Laura

    2014-01-01

    Light is one of the most important environmental factors affecting flavonoid biosynthesis in plants. The absolute dependency of light to the plant development has driven evolvement of sophisticated mechanisms to sense and transduce multiple aspects of the light signal. Light effects can be categorized in photoperiod (duration), intensity (quantity), direction and quality (wavelength) including UV-light. Recently, new information has been achieved on the regulation of light-controlled flavonoid biosynthesis in fruits, in which flavonoids have a major contribution on quality. This review focuses on the effects of the different light conditions on the control of flavonoid biosynthesis in fruit producing plants. An overview of the currently known mechanisms of the light-controlled flavonoid accumulation is provided. R2R3 MYB transcription factors are known to regulate by differential expression the biosynthesis of distinct flavonoids in response to specific light wavelengths. Despite recent advances, many gaps remain to be understood in the mechanisms of the transduction pathway of light-controlled flavonoid biosynthesis. A better knowledge on these regulatory mechanisms is likely to be useful for breeding programs aiming to modify fruit flavonoid pattern. PMID:25346743

  5. A Single Sfp-Type Phosphopantetheinyl Transferase Plays a Major Role in the Biosynthesis of PKS and NRPS Derived Metabolites in Streptomyces ambofaciens ATCC23877

    PubMed Central

    Bunet, Robert; Riclea, Ramona; Laureti, Luisa; Hôtel, Laurence; Paris, Cédric; Girardet, Jean-Michel; Spiteller, Dieter; Dickschat, Jeroen S.; Leblond, Pierre; Aigle, Bertrand

    2014-01-01

    The phosphopantetheinyl transferases (PPTases) are responsible for the activation of the carrier protein domains of the polyketide synthases (PKS), non ribosomal peptide synthases (NRPS) and fatty acid synthases (FAS). The analysis of the Streptomyces ambofaciens ATCC23877 genome has revealed the presence of four putative PPTase encoding genes. One of these genes appears to be essential and is likely involved in fatty acid biosynthesis. Two other PPTase genes, samT0172 (alpN) and samL0372, are located within a type II PKS gene cluster responsible for the kinamycin production and an hybrid NRPS-PKS cluster involved in antimycin production, respectively, and their products were shown to be specifically involved in the biosynthesis of these secondary metabolites. Surprisingly, the fourth PPTase gene, which is not located within a secondary metabolite gene cluster, appears to play a pleiotropic role. Its product is likely involved in the activation of the acyl- and peptidyl-carrier protein domains within all the other PKS and NRPS complexes encoded by S. ambofaciens. Indeed, the deletion of this gene affects the production of the spiramycin and stambomycin macrolide antibiotics and of the grey spore pigment, all three being PKS-derived metabolites, as well as the production of the nonribosomally produced compounds, the hydroxamate siderophore coelichelin and the pyrrolamide antibiotic congocidine. In addition, this PPTase seems to act in concert with the product of samL0372 to activate the ACP and/or PCP domains of the antimycin biosynthesis cluster which is also responsible for the production of volatile lactones. PMID:24498152

  6. Biosynthesis and metabolism of salicylic acid

    SciTech Connect

    Lee, H.; Leon, J.; Raskin, I.

    1995-05-09

    Pathways of salicylic acid (SA) biosynthesis and metabolism in tobacco have been recently identified. SA, an endogenous regulator of disease resistance, is a product of phenylpropanoid metabolism formed via decarboxylation of trans-cinnamic acid to benzoic acid and its subsequent 2-hydroxylation to SA. In tobacco mosaic virus-inoculated tobacco leaves, newly synthesized SA is rapidly metabolized to SA O-{beta}-D-glucoside and methyl salicylate. Two key enzymes involved in SA biosynthesis and metabolism: benzoic acid 2-hydroxylase, which converts benzoic acid to SA, and UDPglucose:SA glucosyltransferase (EC 2.4.1.35), which catalyzes conversion of SA to SA glucoside have been partially purified and characterized. Progress in enzymology and molecular biology of SA biosynthesis and metabolism will provide a better understanding of signal transduction pathway involved in plant disease resistance. 62 refs., 1 fig.

  7. Bacterial exopolysaccharides: biosynthesis pathways and engineering strategies

    PubMed Central

    Schmid, Jochen; Sieber, Volker; Rehm, Bernd

    2015-01-01

    Bacteria produce a wide range of exopolysaccharides which are synthesized via different biosynthesis pathways. The genes responsible for synthesis are often clustered within the genome of the respective production organism. A better understanding of the fundamental processes involved in exopolysaccharide biosynthesis and the regulation of these processes is critical toward genetic, metabolic and protein-engineering approaches to produce tailor-made polymers. These designer polymers will exhibit superior material properties targeting medical and industrial applications. Exploiting the natural design space for production of a variety of biopolymer will open up a range of new applications. Here, we summarize the key aspects of microbial exopolysaccharide biosynthesis and highlight the latest engineering approaches toward the production of tailor-made variants with the potential to be used as valuable renewable and high-performance products for medical and industrial applications. PMID:26074894

  8. Flavonoids: biosynthesis, biological functions, and biotechnological applications

    PubMed Central

    Falcone Ferreyra, María L.; Rius, Sebastián P.; Casati, Paula

    2012-01-01

    Flavonoids are widely distributed secondary metabolites with different metabolic functions in plants. The elucidation of the biosynthetic pathways, as well as their regulation by MYB, basic helix-loop-helix (bHLH), and WD40-type transcription factors, has allowed metabolic engineering of plants through the manipulation of the different final products with valuable applications. The present review describes the regulation of flavonoid biosynthesis, as well as the biological functions of flavonoids in plants, such as in defense against UV-B radiation and pathogen infection, nodulation, and pollen fertility. In addition, we discuss different strategies and achievements through the genetic engineering of flavonoid biosynthesis with implication in the industry and the combinatorial biosynthesis in microorganisms by the reconstruction of the pathway to obtain high amounts of specific compounds. PMID:23060891

  9. The Terpenoid Biosynthesis Toolkit of Trichoderma.

    PubMed

    Bansal, Ravindra; Mukherjee, Prasun Kumar

    2016-04-01

    The widely used biotechnologically important fungi belonging to the genus Trichoderma are rich sources of secondary metabolites. Even though the genomes of several Trichoderma spp. have been published, and data are available on the genes involved in biosynthesis of non-ribosomal peptide synthetases and polyketide synthases, no genome-wide data are available for the terpenoid biosynthesis machinery in these organisms. In the present study, we have identified the genes involved in terpene biosynthesis in the genomes of three Trichoderma spp., viz., T. virens, T. atroviride and T. reesei. While the genes involved in the condensation steps are highly conserved across the three species, these fungi differed in the number and organization of terpene cyclases. T. virens genome harbours eleven terpene cyclases, while T. atroviride harbours seven, and T. reeseisix in their genomes; seven, three and two being part of putative secondary metabolism related gene clusters. PMID:27396184

  10. Purification and Structural Characterization of “Simple Catechol”, the NGAL-Siderocalin Siderophore in Human Urine

    PubMed Central

    Bao, Guan-Hu; Barasch, Jonathan; Xu, Jie; Wang, Wei; Hu, Feng-Lin; Deng, Shi-Xian

    2015-01-01

    The identification of ligands that bind the protein Neutrophil Gelatinase-Associated Lipocalin (NGAL, Siderocalin, Lipocalin-2) have helped to elucidate its function. NGAL-Siderocalin binds and sequesters the iron loaded bacterial siderophore enterochelin (Ent), defining the protein as an innate immune effector. Simple metabolic catechols can also form tight complexes with NGAL-Siderocalin and ferric iron, suggesting that the protein may act as an iron scavenger even in the absence of Ent. While different catechols have been detected in human urine, they have not been directly purified from a biofluid and demonstrated to ligate iron with NGAL-Siderocalin. This paper describes a “natural products” approach to identify small molecules that mediate iron binding to NGAL-Siderocalin. A 10K filtrate of human urine was subjected to multiple steps of column chromatography and reverse-phase HPLC, guided by NGAL-Siderocalin-iron binding assays and LC-MS detection. The co-factor forming a ternary structure with iron and NGAL-Siderocalin was identified as authentic simple catechol (dihydroxybenze) by ESI-HR-Mass, UV, and NMR spectrometric analysis. Comparison of the binding strengths of different catechols demonstrated that the vicinal-dihydroxyl groups were the key functional groups and that steric compatibilities of the catechol ring have the strongest effect on binding. Although catechol was a known NGAL-Siderocalin co-factor, our purification directly confirmed its presence in urine as well as its capacity to serve as an iron trap with NGAL-Siderocalin. PMID:26257890

  11. The Pseudomonas fluorescens Siderophore Pyoverdine Weakens Arabidopsis thaliana Defense in Favor of Growth in Iron-Deficient Conditions.

    PubMed

    Trapet, Pauline; Avoscan, Laure; Klinguer, Agnès; Pateyron, Stéphanie; Citerne, Sylvie; Chervin, Christian; Mazurier, Sylvie; Lemanceau, Philippe; Wendehenne, David; Besson-Bard, Angélique

    2016-05-01

    Pyoverdines are siderophores synthesized by fluorescent Pseudomonas spp. Under iron-limiting conditions, these high-affinity ferric iron chelators are excreted by bacteria in the soil to acquire iron. Pyoverdines produced by beneficial Pseudomonas spp. ameliorate plant growth. Here, we investigate the physiological incidence and mode of action of pyoverdine from Pseudomonas fluorescens C7R12 on Arabidopsis (Arabidopsis thaliana) plants grown under iron-sufficient or iron-deficient conditions. Pyoverdine was provided to the medium in its iron-free structure (apo-pyoverdine), thus mimicking a situation in which it is produced by bacteria. Remarkably, apo-pyoverdine abolished the iron-deficiency phenotype and restored the growth of plants maintained in the iron-deprived medium. In contrast to a P. fluorescens C7R12 strain impaired in apo-pyoverdine production, the wild-type C7R12 reduced the accumulation of anthocyanins in plants grown in iron-deficient conditions. Under this condition, apo-pyoverdine modulated the expression of around 2,000 genes. Notably, apo-pyoverdine positively regulated the expression of genes related to development and iron acquisition/redistribution while it repressed the expression of defense-related genes. Accordingly, the growth-promoting effect of apo-pyoverdine in plants grown under iron-deficient conditions was impaired in iron-regulated transporter1 and ferric chelate reductase2 knockout mutants and was prioritized over immunity, as highlighted by an increased susceptibility to Botrytis cinerea This process was accompanied by an overexpression of the transcription factor HBI1, a key node for the cross talk between growth and immunity. This study reveals an unprecedented mode of action of pyoverdine in Arabidopsis and demonstrates that its incidence on physiological traits depends on the plant iron status. PMID:26956666

  12. Triterpenoid biosynthesis in Euphorbia lathyris latex

    SciTech Connect

    Hawkins, D.R.

    1987-11-01

    The structures of triterpenols, not previously been known, from Euphorbia lathyris latex are reported. A method for quantifying very small amounts of these compounds was developed. Concerning the biochemistry of the latex, no exogenous cofactors were required for the biosynthesis and the addition of compounds such as NADPAH and ATP do not stimulate the biosynthesis. The addition of DTE or a similar anti-oxidant was found to help reduce the oxidation of the latex, thus increasing the length of time that the latex remains active. The requirement of a divalent cation and the preference for Mn in the pellet was observed. The effect of several inhibitors on the biosynthesis of the triterpenoids was examined. Mevinolin was found to inhibit the biosynthesis of the triterpenoids from acetate, but not mevalonate. A dixon plot of the inhibition of acetate incorporation showed an I/sub 50/ concentration of 3.2 ..mu..M. Fenpropimorph was found to have little or no effect on the biosynthesis. Tridemorph was found to inhibit the biosynthesis of all of the triterpenoids with an I/sub 50/ of 4 ..mu..M. It was also observed that the cyclopropyl containing triterpenols, cycloartenol and 24-methylenecycloartenol were inhibited much more strongly than those containing an 8-9 double bond, lanosterol and 24-methylenelanosterol. The evidence indicates, but does not definetely prove, that lanosterol and 24-methylenelanosterol are not made from cycloartenol and 24-methylenecycloartenol via a ring-opening enzyme such as cycloeucalenol-obtusifoliol isomerase. The possibilty that cycloartenol is made via lanosterol was investigated by synthesizing 4-R-4-/sup 3/H-mevalonic acid and incubating latex with a mixture of this and /sup 14/C-mevalonic acid. From the /sup 3/H//sup 14/C ratio it was shown that cycloartenol and 24-methylenecycloartenol are not made via an intermediate containing as 8-9 double bond. 88 refs., 15 figs., 30 tabs.

  13. Pyochelin, a siderophore of Pseudomonas aeruginosa: physicochemical characterization of the iron(III), copper(II) and zinc(II) complexes.

    PubMed

    Brandel, Jérémy; Humbert, Nicolas; Elhabiri, Mourad; Schalk, Isabelle J; Mislin, Gaëtan L A; Albrecht-Gary, Anne-Marie

    2012-03-01

    Pseudomonas aeruginosa is an opportunistic pathogen, synthesizing two major siderophores, pyoverdine (Pvd) and pyochelin (Pch), to cover its needs in iron(III). If the high affinity and specificity of Pvd toward iron(III) (pFe = 27.0) was well described in the literature, the physicochemical and coordination properties of Pch toward biologically relevant metals (Fe(III), Cu(II) or Zn(II)) have been only scarcely investigated. We report a thorough physico-chemical investigation of Pch (potentiometry, spectrophotometries, ESI/MS) that highlighted its moderate but significantly higher affinity for Fe(3+) (pFe = 16.0 at p[H] 7.4) than reported previously. We also demonstrated that Pch strongly chelates divalent metals such as Zn(II) (pZn = 11.8 at p[H] 7.4) and Cu(II) (pCu = 14.9 at p[H] 7.4) and forms predominantly 1 : 2 (M(2+)/Pch) complexes. Kinetic studies revealed that the formation of the ferric Pch complexes proceeds through a Eigen-Wilkins dissociative ligand interchange mechanism involving two protonated species of Pch and the Fe(OH)(2+) species of Fe(III). Our physico-chemical parameters supports the previous biochemical studies which proposed that siderophores are not only devoted to iron(III) shuttling but most likely display other specific biological role in the subtle metals homeostasis in microorganisms. This work also represents a step toward deciphering the role of siderophores throughout evolution. PMID:22261733

  14. Biosynthesis of pyochelin and dihydroaeruginoic acid requires the iron-regulated pchDCBA operon in Pseudomonas aeruginosa.

    PubMed

    Serino, L; Reimmann, C; Visca, P; Beyeler, M; Chiesa, V D; Haas, D

    1997-01-01

    The high-affinity siderophore salicylate is an intermediate in the biosynthetic pathway of pyochelin, another siderophore and chelator of transition metal ions, in Pseudomonas aeruginosa. The 2.5-kb region upstream of the salicylate biosynthetic genes pchBA was sequenced and found to contain two additional, contiguous genes, pchD and pchC, having the same orientation. The deduced amino acid sequence of the 60-kDa PchD protein was similar to those of the EntE protein (2,3-dihydroxybenzoate-AMP ligase) of Escherichia coli and other adenylate-forming enzymes, suggesting that salicylate might be adenylated at the carboxyl group by PchD. The 28-kDa PchC protein showed similarities to thioesterases of prokaryotic and eukaryotic origin and might participate in the release of the product(s) formed from activated salicylate. One potential product, dihydroaeruginoate (Dha), was identified in culture supernatants of iron-limited P. aeruginosa cells. The antifungal antibiotic Dha is thought to arise from the reaction of salicylate with cysteine, followed by cyclization of cysteine. Inactivation of the chromosomal pchD gene by insertion of the transcription and translation stop element omega Sm/Sp abolished the production of Dha and pyochelin, implying that PchD-mediated activation of salicylate may be a common first step in the synthesis of both metabolites. Furthermore, the pchD::omega Sm/Sp mutation had a strong polar effect on the expression of the pchBA genes, i.e., on salicylate synthesis, indicating that the pchDCBA genes constitute a transcriptional unit. A full-length pchDCBA transcript of ca. 4.4 kb could be detected in iron-deprived, growing cells of P. aeruginosa. Transcription of pchD started at tandemly arranged promoters, which overlapped with two Fur boxes (binding sites for the ferric uptake regulator) and the promoter of the divergently transcribed pchR gene encoding an activator of pyochelin biosynthesis. This promoter arrangement allows tight iron

  15. The structural biology of phenazine biosynthesis

    PubMed Central

    Blankenfeldt, Wulf; Parsons, James F.

    2014-01-01

    The phenazines are a class of over 150 nitrogen-containing aromatic compounds of bacterial and archeal origin. Their redox properties not only explain their activity as broad-specificity antibiotics and virulence factors but also enable them to function as respiratory pigments, thus extending their importance to the primary metabolism of phenazine-producing species. Despite their discovery in the mid-19th century, the molecular mechanisms behind their biosynthesis have only been unraveled in the last decade. Here, we review the contribution of structural biology that has led to our current understanding of phenazine biosynthesis. PMID:25215885

  16. The structural biology of phenazine biosynthesis.

    PubMed

    Blankenfeldt, Wulf; Parsons, James F

    2014-12-01

    The phenazines are a class of over 150 nitrogen-containing aromatic compounds of bacterial and archeal origin. Their redox properties not only explain their activity as broad-specificity antibiotics and virulence factors but also enable them to function as respiratory pigments, thus extending their importance to the primary metabolism of phenazine-producing species. Despite their discovery in the mid-19th century, the molecular mechanisms behind their biosynthesis have only been unraveled in the last decade. Here, we review the contribution of structural biology that has led to our current understanding of phenazine biosynthesis. PMID:25215885

  17. Biosynthesis and biodegradation of wood components

    SciTech Connect

    Higuchi, T.

    1985-01-01

    A textbook containing 22 chapters by various authors covers the structure of wood, the localization of polysaccharides and lignins in wood cell walls, metabolism and synthetic function of cambial tissue, cell organelles and their function in the biosynthesis of cell wall components, biosynthesis of plant cell wall polysaccharides, lignin, cutin, suberin and associated waxes, phenolic acids and monolignols, quinones, flavonoids, tannins, stilbenes and terpenoid wood extractives, the occurrence of extractives, the metabolism of phenolic acids, wood degradation by micro-organisms and fungi, and biodegradation of cellulose, hemicelluloses, lignin, and aromatic extractives of wood. An index is included.

  18. Solid-to-solid oxidation of a vanadium(IV) to a vanadium(V) compound: chemisty of a sulfur-containing siderophore.

    PubMed

    Chatterjee, Pabitra B; Crans, Debbie C

    2012-09-01

    Visible light facilitates a solid-to-solid photochemical aerobic oxidation of a hunter-green microcrystalline oxidovanadium(IV) compound (1) to form a black powder of cis-dioxidovanadium(V) (2) at ambient temperature. The siderophore ligand pyridine-2,6-bis(thiocarboxylic acid), H(2)L, is secreted by a microorganism from the Pseudomonas genus. This irreversible transformation of a metal monooxo to a metal dioxo complex in the solid state in the absence of solvent is unprecedented. It serves as a proof-of-concept reaction for green chemistry occurring in solid matrixes. PMID:22880634

  19. Biosynthesis of coenzyme Q in eukaryotes.

    PubMed

    Kawamukai, Makoto

    2015-01-01

    Coenzyme Q (CoQ) is a component of the electron transport chain that participates in aerobic cellular respiration to produce ATP. In addition, CoQ acts as an electron acceptor in several enzymatic reactions involving oxidation-reduction. Biosynthesis of CoQ has been investigated mainly in Escherichia coli and Saccharomyces cerevisiae, and the findings have been extended to various higher organisms, including plants and humans. Analyses in yeast have contributed greatly to current understanding of human diseases related to CoQ biosynthesis. To date, human genetic disorders related to mutations in eight COQ biosynthetic genes have been reported. In addition, the crystal structures of a number of proteins involved in CoQ synthesis have been solved, including those of IspB, UbiA, UbiD, UbiX, UbiI, Alr8543 (Coq4 homolog), Coq5, ADCK3, and COQ9. Over the last decade, knowledge of CoQ biosynthesis has accumulated, and striking advances in related human genetic disorders and the crystal structure of proteins required for CoQ synthesis have been made. This review focuses on the biosynthesis of CoQ in eukaryotes, with some comparisons to the process in prokaryotes. PMID:26183239

  20. Cyclopiazonic acid biosynthesis by Aspergillus flavus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cyclopiazonic acid (CPA) is an indole-tetramic acid mycotoxin produced by some strains of Aspergillus flavus. Characterization of the CPA biosynthesis gene cluster confirmed that formation of CPA is via a three-enzyme pathway. This review examines the structure and organization of the CPA genes, elu...

  1. Control of aflatoxin biosynthesis in Aspergilli

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Expression of the genes in the AF biosynthesis cluster is mainly controlled by the pathway specific Cys6Zn2 DNA binding protein, AflR. While AflR appears to be necessary for the activation, a number of coactivators are important for fine-tuning of the timing of AflR’s activity. These proteins, AflJ,...

  2. Siderophore-mediated oxidation of Ce and fractionation of HREE by Mn (hydr)oxide-coprecipitation and sorption on MnO2: Experimental evidence for negative Ce-anomalies in abiogenic manganese precipitates

    NASA Astrophysics Data System (ADS)

    Krämer, Dennis; Tepe, Nathalie; Bau, Michael

    2014-05-01

    We conducted experiments with Rare Earths and Yttrium (REY), where the REY were sorbed on synthetic manganese dioxide as well as on coprecipitating manganese (hydr)oxide in the presence and absence of the siderophore desferrioxamine-B (DFOB). Siderophores are a group of globally abundant biogenic complexing agents which are excreted by plants and bacteria to enhance the bioavailability of Fe in oxic environments. The model siderophore used in this study, DFOB, is a hydroxamate siderophore occurring in almost all environmental settings with concentrations in the nanomolar to millimolar range and is one of the most thoroughly studied siderophores. In the absence of siderophores and other organic ligands, trivalent Ce is usually surface-oxidized to tetravalent Ce during sorption onto manganese (hydr)oxides. Such Mn precipitates, therefore, often show positive Ce anomalies, whereas the ambient solutions exhibit negative Ce anomalies (Ohta and Kawabe, 2001). In marked contrast, however, REY sorption in the presence of DFOB produces negative Ce anomalies in the Mn precipitates and a distinct and characteristic positive Ce anomaly in the residual siderophore-bearing solution. Furthermore, the heavy REY with ionic radii larger than the radius of Sm are also almost completely prevented from sorption onto the Mn solid phases. Sorption of REY onto Mn (hydr)oxides in the presence of DFOB creates a distinct and pronounced fractionation of Ce and the heavy REY from the light and middle REY. Apart from Ce, which is oxidized in solution by the siderophore, the distribution of the other REY mimics the stability constants for multi-dentate complexes of REY with DFOB, as determined by Christenson & Schijf (2011). Heavier REY are forming stronger complexes (and are hence better "protected" from sorption) than light REY, excluding Ce. Preferential partitioning of Ce into the liquid phase during the precipitation of Mn (hydr)oxides has only rarely been described for natural Mn (hydr

  3. Evidence for Hydroxamate Siderophores and Other N-Containing Organic Compounds Controlling (239,240)Pu Immobilization and Remobilization in a Wetland Sediment.

    PubMed

    Xu, Chen; Zhang, Saijin; Kaplan, Daniel I; Ho, Yi-Fang; Schwehr, Kathleen A; Roberts, Kimberly A; Chen, Hongmei; DiDonato, Nicole; Athon, Matthew; Hatcher, Patrick G; Santschi, Peter H

    2015-10-01

    Pu concentrations in wetland surface sediments collected downstream of a former nuclear processing facility in F-Area of the Savannah River Site (SRS), USA, were ∼2.5 times greater than those measured in the associated upland aquifer sediments; similarly, the Pu concentration solid/water ratios were orders of magnitude greater in the wetland than in the low-organic matter content aquifer soils. Sediment Pu concentrations were correlated to total organic carbon and total nitrogen contents and even more strongly to hydroxamate siderophore (HS) concentrations. The HS were detected in the particulate or colloidal phases of the sediments but not in the low molecular weight fractions (<1000 Da). Macromolecules which scavenged the majority of the potentially mobile Pu were further separated from the bulk mobile organic matter fraction ("water extract") via an isoelectric focusing experiment (IEF). An electrospray ionization Fourier-transform ion cyclotron resonance ultrahigh resolution mass spectrometry (ESI FTICR-MS) spectral comparison of the IEF extract and a siderophore standard (desferrioxamine; DFO) suggested the presence of HS functionalities in the IEF extract. This study suggests that while HS are a very minor component in the sediment particulate/colloidal fractions, their concentrations greatly exceed those of ambient Pu, and HS may play an especially important role in Pu immobilization/remobilization in wetland sediments. PMID:26313339

  4. Detection of a Serum Siderophore by LC-MS/MS as a Potential Biomarker of Invasive Aspergillosis

    PubMed Central

    Carroll, Cassandra S.; Amankwa, Lawrence N.; Pinto, Linda J.; Fuller, Jeffrey D.; Moore, Margo M.

    2016-01-01

    Invasive aspergillosis (IA) is a life-threatening systemic mycosis caused primarily by Aspergillus fumigatus. Early diagnosis of IA is based, in part, on an immunoassay for circulating fungal cell wall carbohydrate, galactomannan (GM). However, a wide range of sensitivity and specificity rates have been reported for the GM test across various patient populations. To obtain iron in vivo, A. fumigatus secretes the siderophore, N,N',N"-triacetylfusarinine C (TAFC) and we hypothesize that TAFC may represent a possible biomarker for early detection of IA. We developed an ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for TAFC analysis from serum, and measured TAFC in serum samples collected from patients at risk for IA. The method showed lower and upper limits of quantitation (LOQ) of 5 ng/ml and 750 ng/ml, respectively, and complete TAFC recovery from spiked serum. As proof of concept, we evaluated 76 serum samples from 58 patients with suspected IA that were investigated for the presence of GM. Fourteen serum samples obtained from 11 patients diagnosed with probable or proven IA were also analyzed for the presence of TAFC. Control sera (n = 16) were analyzed to establish a TAFC cut-off value (≥6 ng/ml). Of the 36 GM-positive samples (≥0.5 GM index) from suspected IA patients, TAFC was considered positive in 25 (69%). TAFC was also found in 28 additional GM-negative samples. TAFC was detected in 4 of the 14 samples (28%) from patients with proven/probable aspergillosis. Log-transformed TAFC and GM values from patients with proven/probable IA, healthy individuals and SLE patients showed a significant correlation with a Pearson r value of 0.77. In summary, we have developed a method for the detection of TAFC in serum that revealed this fungal product in the sera of patients at risk for invasive aspergillosis. A prospective study is warranted to determine whether this method provides improved early detection of IA. PMID:26974544

  5. Complete biosynthesis of opioids in yeast.

    PubMed

    Galanie, Stephanie; Thodey, Kate; Trenchard, Isis J; Filsinger Interrante, Maria; Smolke, Christina D

    2015-09-01

    Opioids are the primary drugs used in Western medicine for pain management and palliative care. Farming of opium poppies remains the sole source of these essential medicines, despite diverse market demands and uncertainty in crop yields due to weather, climate change, and pests. We engineered yeast to produce the selected opioid compounds thebaine and hydrocodone starting from sugar. All work was conducted in a laboratory that is permitted and secured for work with controlled substances. We combined enzyme discovery, enzyme engineering, and pathway and strain optimization to realize full opiate biosynthesis in yeast. The resulting opioid biosynthesis strains required the expression of 21 (thebaine) and 23 (hydrocodone) enzyme activities from plants, mammals, bacteria, and yeast itself. This is a proof of principle, and major hurdles remain before optimization and scale-up could be achieved. Open discussions of options for governing this technology are also needed in order to responsibly realize alternative supplies for these medically relevant compounds. PMID:26272907

  6. Chemical genetics to examine cellulose biosynthesis

    PubMed Central

    Brabham, Chad; DeBolt, Seth

    2013-01-01

    Long-term efforts to decode plant cellulose biosynthesis via molecular genetics and biochemical strategies are being enhanced by the ever-expanding scale of omics technologies. An alternative approach to consider are the prospects for inducing change in plant metabolism using exogenously supplied chemical ligands. Cellulose biosynthesis inhibitors (CBIs) have been identified among known herbicides, during diverse combinatorial chemical libraries screens, and natural chemical screens from microbial agents. In this review, we summarize the current knowledge of the inhibitory effects of CBIs and further group them by how they influence fluorescently tagged cellulose synthase A proteins. Additional attention is paid to the continuing development of the CBI toolbox to explore the cell biology and genetic mechanisms underpinning effector molecule activity. PMID:23372572

  7. Structural basis for phosphatidylinositol-phosphate biosynthesis

    PubMed Central

    Clarke, Oliver B.; Tomasek, David; Jorge, Carla D.; Dufrisne, Meagan Belcher; Kim, Minah; Banerjee, Surajit; Rajashankar, Kanagalaghatta R.; Shapiro, Lawrence; Hendrickson, Wayne A.; Santos, Helena; Mancia, Filippo

    2015-01-01

    Phosphatidylinositol is critical for intracellular signalling and anchoring of carbohydrates and proteins to outer cellular membranes. The defining step in phosphatidylinositol biosynthesis is catalysed by CDP-alcohol phosphotransferases, transmembrane enzymes that use CDP-diacylglycerol as donor substrate for this reaction, and either inositol in eukaryotes or inositol phosphate in prokaryotes as the acceptor alcohol. Here we report the structures of a related enzyme, the phosphatidylinositol-phosphate synthase from Renibacterium salmoninarum, with and without bound CDP-diacylglycerol to 3.6 and 2.5 Å resolution, respectively. These structures reveal the location of the acceptor site, and the molecular determinants of substrate specificity and catalysis. Functional characterization of the 40%-identical ortholog from Mycobacterium tuberculosis, a potential target for the development of novel anti-tuberculosis drugs, supports the proposed mechanism of substrate binding and catalysis. This work therefore provides a structural and functional framework to understand the mechanism of phosphatidylinositol-phosphate biosynthesis. PMID:26510127

  8. Ceramide biosynthesis and metabolism in trophoblast syncytialization.

    PubMed

    Singh, Ambika T; Dharmarajan, Arunasalam; Aye, Irving L M H; Keelan, Jeffrey A

    2012-10-15

    Sphingolipid mediators such as ceramide are pleiotropic regulators of cellular growth, differentiation and apoptosis. We investigated the role of ceramide biosynthesis, metabolism and actions in term human cytotrophoblasts syncytialized over 7 days in culture. Intracellular C16 ceramide levels increased modestly after 3 days in culture, then declined. Ceramidase was present at particularly high levels in syncytialized trophoblasts; inhibition of ceramidase reduced the degree of cell fusion. Exposure to short chain C8 ceramide or aSMase enhanced secretion of the differentiation marker hCG without affecting fusion or cell viability. In contrast, pharmacological inhibition of ceramidase reduced the extent of fusion. Inhibition of the ceramide-responsive JNK and PP2A pathways did not abolish the effects of ceramide, and JNK phosphorylation was unresponsive to ceramide; however, ceramide significantly inhibited phosphorylation of Akt. This study suggests that changes in ceramide biosynthesis and metabolism play a differential role in the biochemical and morphological features of trophoblast differentiation. PMID:22652149

  9. Structural basis for phosphatidylinositol-phosphate biosynthesis.

    PubMed

    Clarke, Oliver B; Tomasek, David; Jorge, Carla D; Dufrisne, Meagan Belcher; Kim, Minah; Banerjee, Surajit; Rajashankar, Kanagalaghatta R; Shapiro, Lawrence; Hendrickson, Wayne A; Santos, Helena; Mancia, Filippo

    2015-01-01

    Phosphatidylinositol is critical for intracellular signalling and anchoring of carbohydrates and proteins to outer cellular membranes. The defining step in phosphatidylinositol biosynthesis is catalysed by CDP-alcohol phosphotransferases, transmembrane enzymes that use CDP-diacylglycerol as donor substrate for this reaction, and either inositol in eukaryotes or inositol phosphate in prokaryotes as the acceptor alcohol. Here we report the structures of a related enzyme, the phosphatidylinositol-phosphate synthase from Renibacterium salmoninarum, with and without bound CDP-diacylglycerol to 3.6 and 2.5 Å resolution, respectively. These structures reveal the location of the acceptor site, and the molecular determinants of substrate specificity and catalysis. Functional characterization of the 40%-identical ortholog from Mycobacterium tuberculosis, a potential target for the development of novel anti-tuberculosis drugs, supports the proposed mechanism of substrate binding and catalysis. This work therefore provides a structural and functional framework to understand the mechanism of phosphatidylinositol-phosphate biosynthesis. PMID:26510127

  10. Structural basis for phosphatidylinositol-phosphate biosynthesis

    NASA Astrophysics Data System (ADS)

    Clarke, Oliver B.; Tomasek, David; Jorge, Carla D.; Dufrisne, Meagan Belcher; Kim, Minah; Banerjee, Surajit; Rajashankar, Kanagalaghatta R.; Shapiro, Lawrence; Hendrickson, Wayne A.; Santos, Helena; Mancia, Filippo

    2015-10-01

    Phosphatidylinositol is critical for intracellular signalling and anchoring of carbohydrates and proteins to outer cellular membranes. The defining step in phosphatidylinositol biosynthesis is catalysed by CDP-alcohol phosphotransferases, transmembrane enzymes that use CDP-diacylglycerol as donor substrate for this reaction, and either inositol in eukaryotes or inositol phosphate in prokaryotes as the acceptor alcohol. Here we report the structures of a related enzyme, the phosphatidylinositol-phosphate synthase from Renibacterium salmoninarum, with and without bound CDP-diacylglycerol to 3.6 and 2.5 Å resolution, respectively. These structures reveal the location of the acceptor site, and the molecular determinants of substrate specificity and catalysis. Functional characterization of the 40%-identical ortholog from Mycobacterium tuberculosis, a potential target for the development of novel anti-tuberculosis drugs, supports the proposed mechanism of substrate binding and catalysis. This work therefore provides a structural and functional framework to understand the mechanism of phosphatidylinositol-phosphate biosynthesis.

  11. Complete biosynthesis of opioids in yeast

    PubMed Central

    Galanie, Stephanie; Thodey, Kate; Trenchard, Isis J.; Interrante, Maria Filsinger; Smolke, Christina D.

    2016-01-01

    Opioids are the primary drugs used in Western medicine for pain management and palliative care. Farming of opium poppies remains the sole source of these essential medicines despite diverse market demands and uncertainty in crop yields due to weather, climate change, and pests. Here, we engineered yeast to produce the selected opioid compounds thebaine and hydrocodone starting from sugar. All work was conducted in a laboratory that is permitted and secured for work with controlled substances. We combined enzyme discovery, enzyme engineering, and pathway and strain optimization to realize full opiate biosynthesis in yeast. The resulting opioid biosynthesis strains required expression of 21 (thebaine) and 23 (hydrocodone) enzyme activities from plants, mammals, bacteria, and yeast itself. This is a proof-of-principle, and major hurdles remain before optimization and scale up could be achieved. Open discussions of options for governing this technology are also needed in order to responsibly realize alternative supplies for these medically relevant compounds. PMID:26272907

  12. Moss cell walls: structure and biosynthesis

    PubMed Central

    Roberts, Alison W.; Roberts, Eric M.; Haigler, Candace H.

    2012-01-01

    The genome sequence of the moss Physcomitrella patens has stimulated new research examining the cell wall polysaccharides of mosses and the glycosyl transferases that synthesize them as a means to understand fundamental processes of cell wall biosynthesis and plant cell wall evolution. The cell walls of mosses and vascular plants are composed of the same classes of polysaccharides, but with differences in side chain composition and structure. Similarly, the genomes of P. patens and angiosperms encode the same families of cell wall glycosyl transferases, yet, in many cases these families have diversified independently in each lineage. Our understanding of land plant evolution could be enhanced by more complete knowledge of the relationships among glycosyl transferase functional diversification, cell wall structural and biochemical specialization, and the roles of cell walls in plant adaptation. As a foundation for these studies, we review the features of P. patens as an experimental system, analyses of cell wall composition in various moss species, recent studies that elucidate the structure and biosynthesis of cell wall polysaccharides in P. patens, and phylogenetic analysis of P. patens genes potentially involved in cell wall biosynthesis. PMID:22833752

  13. Tetrahydrobiopterin biosynthesis, utilization and pharmacological effects.

    PubMed

    Werner-Felmayer, G; Golderer, G; Werner, E R

    2002-04-01

    Tetrahydrobiopterin (H4-biopterin) is an essential cofactor of a set of enzymes that are of central metabolic importance, i.e. the hydroxylases of the three aromatic amino acids phenylalanine, tyrosine, and tryptophan, of ether lipid oxidase, and of the three nitric oxide synthase (NOS) isoenzymes. As a consequence, H4-biopterin plays a key role in a vast number of biological processes and pathological states associated with neurotransmitter formation, vasorelaxation, and immune response. In mammals, its biosynthesis is controlled by hormones, cytokines and certain immune stimuli. This review aims to summarize recent developments concerning regulation of H4-biopterin biosynthetic and regulatory enzymes and pharmacological effects of H4-biopterin in various conditions, e.g. endothelial dysfunction or apoptosis of neuronal cells. Also, approaches towards gene therapy of diseases like the different forms of phenylketonuria or of Parkinson's disease are reviewed. Additional emphasis is given to H4-biopterin biosynthesis and function in non-mammalian species such as fruit fly, zebra fish, fungi, slime molds, the bacterium Nocardia as well as to the parasitic protozoan genus of Leishmania that is not capable of pteridine biosynthesis but has evolved a sophisticated salvage network for scavenging various pteridine compounds, notably folate and biopterin. PMID:12003348

  14. Regulation of Leucine Biosynthesis in Bacillus subtilis

    PubMed Central

    Ward, Jonathan B.; Zahler, Stanley A.

    1973-01-01

    The biosynthesis of α-isopropylmalate (αIPM) synthetase, IPM isomerase, and βIPM dehydrogenase in Bacillus subtilis can be derepressed in leucine auxotrophs by limiting them for leucine. The derepression of the three enzymes is apparently coordinate. A class of mutants resistant to 4-azaleucine excretes leucine and has derepressed levels of all three enzymes. The azaleucine-resistance mutations may lie in a gene (azlA) encoding a repressor. Efforts to find mutations characteristic of a constitutive operator have been unsuccessful. No polar mutations have been found among nine leucine auxotrophs that have characteristics of frameshift mutations. The enzyme catalyzing the first step in leucine biosynthesis, αIPM synthetase, is sensitive to feedback inhibition by leucine. We conclude that leucine biosynthesis is controlled by the inhibition of the activity of the first biosynthetic enzyme by leucine, and by the repression of the synthesis of the first three biosynthetic enzymes by leucine. The repression of the three enzymes may be under the control of a single repressor and a single operator, or of a single repressor and a separate operator for each structural gene. PMID:4200854

  15. Urinary Concentrations and Antibacterial Activity of BAL30072, a Novel Siderophore Monosulfactam, against Uropathogens after Intravenous Administration in Healthy Subjects.

    PubMed

    Straubinger, Marion; Blenk, Holger; Naber, Kurt G; Wagenlehner, Florian M E

    2016-06-01

    This annex study to a phase 1 study aimed to correlate urinary concentrations and bactericidal titers (UBTs) of BAL30072, a novel siderophore monosulfactam, in healthy subjects in order to evaluate which dosage of BAL30072 should be investigated in a clinical study on complicated urinary tract infection (UTI). Three cohorts of a total of 19 healthy male subjects were included in the add-on study and received the following BAL30072 dosages. The 1st cohort received 1 g once a day (q.d.) intravenously (i.v.) (1 h) on day 1 and 1 g thrice daily (t.i.d.) on day 2, the 2nd cohort received 2 g q.d. i.v. (1 h) on day 1 and 2 g t.i.d. on day 2, and the 3rd cohort received 1 g q.d. i.v. (4-h infusion) on day 8. Urine was collected up to 24 h after drug administration. UBTs were determined for seven Escherichia coli isolates (three wild type [WT], CTX-M-15, TEM-3, TEM-5, NDM-1), two Klebsiella pneumoniae isolates (WT, KPC), one Proteus mirabilis isolate (WT), and two Pseudomonas aeruginosa isolates (WT, VIM-1 plus AmpC). Urine drug concentrations were measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The median urinary excretions of BAL30072 ranged between 38% and 46% (3 cohorts). The median UBTs after i.v. administration of 1 or 2 g q.d. and after 1 or 2 g t.i.d. showed positive UBTs for 24 h after the lowest dosage (1 g q.d.) for 5 of 7 of the Enterobacteriaceae strains and after the higher dosage of 2 g administered i.v. t.i.d. for all strains tested. After i.v. infusion of 1 g over 4 h, positive UBTs were demonstrated for three E. coli strains for up to 12 h, for the K. pneumoniae (KPC) strain for up to 8 h, and for the P. aeruginosa (VIM-1 plus AmpC) strain for up to only 4 h. The minimal bactericidal concentrations (MBCs) of the E. coli (NDM-1) strain and the K. pneumoniae (WT) strain correlated well between broth and urine but did not correlate well for the two P. aeruginosa strains. BAL30072 exhibits positive UBTs for 24 h even after a dosage of 1

  16. A putative siderophore-interacting protein from the marine bacterium Shewanella frigidimarina NCIMB 400: cloning, expression, purification, crystallization and X-ray diffraction analysis

    PubMed Central

    Trindade, Inês B.; Fonseca, Bruno M.; Matias, Pedro M.; Louro, Ricardo O.; Moe, Elin

    2016-01-01

    Siderophore-binding proteins (SIPs) perform a key role in iron acquisition in multiple organisms. In the genome of the marine bacterium Shewanella frigidimarina NCIMB 400, the gene tagged as SFRI_RS12295 encodes a protein from this family. Here, the cloning, expression, purification and crystallization of this protein are reported, together with its preliminary X-ray crystallographic analysis to 1.35 Å resolution. The SIP crystals belonged to the monoclinic space group P21, with unit-cell parameters a = 48.04, b = 78.31, c = 67.71 Å, α = 90, β = 99.94, γ = 90°, and are predicted to contain two molecules per asymmetric unit. Structure determination by molecular replacement and the use of previously determined ∼2 Å resolution SIP structures with ∼30% sequence identity as templates are ongoing. PMID:27599855

  17. Direct Detection of Fungal Siderophores on Bats with White-Nose Syndrome via Fluorescence Microscopy-Guided Ambient Ionization Mass Spectrometry

    PubMed Central

    Mascuch, Samantha J.; Moree, Wilna J.; Hsu, Cheng-Chih; Turner, Gregory G.; Cheng, Tina L.; Blehert, David S.; Kilpatrick, A. Marm; Frick, Winifred F.; Meehan, Michael J.; Dorrestein, Pieter C.; Gerwick, Lena

    2015-01-01

    White-nose syndrome (WNS) caused by the pathogenic fungus Pseudogymnoascus destructans is decimating the populations of several hibernating North American bat species. Little is known about the molecular interplay between pathogen and host in this disease. Fluorescence microscopy ambient ionization mass spectrometry was used to generate metabolic profiles from the wings of both healthy and diseased bats of the genus Myotis. Fungal siderophores, molecules that scavenge iron from the environment, were detected on the wings of bats with WNS, but not on healthy bats. This work is among the first examples in which microbial molecules are directly detected from an infected host and highlights the ability of atmospheric ionization methodologies to provide direct molecular insight into infection. PMID:25781976

  18. Direct detection of fungal siderophores on bats with white-nose syndrome via fluorescence microscopy-guided ambient ionization mass spectrometry

    USGS Publications Warehouse

    Mascuch, Samantha J.; Moree, Wilna J.; Cheng-Chih Hsu, Cheng-Chih; Turner, Gregory G.; Cheng, Tina L.; Blehert, David S.; Kilpatrick, A. Marm; Frick, Winifred F.; Meehan, Michael J.; Dorrestein, Pieter C.; Gerwick, Lena

    2015-01-01

    White-nose syndrome (WNS) caused by the pathogenic fungus Pseudogymnoascus destructans is decimating the populations of several hibernating North American bat species. Little is known about the molecular interplay between pathogen and host in this disease. Fluorescence microscopy ambient ionization mass spectrometry was used to generate metabolic profiles from the wings of both healthy and diseased bats of the genus Myotis. Fungal siderophores, molecules that scavenge iron from the environment, were detected on the wings of bats with WNS, but not on healthy bats. This work is among the first examples in which microbial molecules are directly detected from an infected host and highlights the ability of atmospheric ionization methodologies to provide direct molecular insight into infection.

  19. Direct detection of fungal siderophores on bats with white-nose syndrome via fluorescence microscopy-guided ambient ionization mass spectrometry.

    PubMed

    Mascuch, Samantha J; Moree, Wilna J; Hsu, Cheng-Chih; Turner, Gregory G; Cheng, Tina L; Blehert, David S; Kilpatrick, A Marm; Frick, Winifred F; Meehan, Michael J; Dorrestein, Pieter C; Gerwick, Lena

    2015-01-01

    White-nose syndrome (WNS) caused by the pathogenic fungus Pseudogymnoascus destructans is decimating the populations of several hibernating North American bat species. Little is known about the molecular interplay between pathogen and host in this disease. Fluorescence microscopy ambient ionization mass spectrometry was used to generate metabolic profiles from the wings of both healthy and diseased bats of the genus Myotis. Fungal siderophores, molecules that scavenge iron from the environment, were detected on the wings of bats with WNS, but not on healthy bats. This work is among the first examples in which microbial molecules are directly detected from an infected host and highlights the ability of atmospheric ionization methodologies to provide direct molecular insight into infection. PMID:25781976

  20. A putative siderophore-interacting protein from the marine bacterium Shewanella frigidimarina NCIMB 400: cloning, expression, purification, crystallization and X-ray diffraction analysis.

    PubMed

    Trindade, Inês B; Fonseca, Bruno M; Matias, Pedro M; Louro, Ricardo O; Moe, Elin

    2016-09-01

    Siderophore-binding proteins (SIPs) perform a key role in iron acquisition in multiple organisms. In the genome of the marine bacterium Shewanella frigidimarina NCIMB 400, the gene tagged as SFRI_RS12295 encodes a protein from this family. Here, the cloning, expression, purification and crystallization of this protein are reported, together with its preliminary X-ray crystallographic analysis to 1.35 Å resolution. The SIP crystals belonged to the monoclinic space group P21, with unit-cell parameters a = 48.04, b = 78.31, c = 67.71 Å, α = 90, β = 99.94, γ = 90°, and are predicted to contain two molecules per asymmetric unit. Structure determination by molecular replacement and the use of previously determined ∼2 Å resolution SIP structures with ∼30% sequence identity as templates are ongoing. PMID:27599855

  1. Metabolic flux analysis of diterpene biosynthesis pathway in rice.

    PubMed

    Chang, Yung-Jin; Kim, Bo-Ra; Kim, Soo-Un

    2005-09-01

    Relative transcript levels of eight rice diterpene cyclases at the branch points of gibberellins and phytoalexins biosynthesis pathway were measured by reverse transcription quantitative PCR. Metabolic flux analysis by the distribution ratio of common substrate showed that UV-irradiation of etiolated rice seedlings decreased the flux for primary metabolism of gibberellins biosynthesis by half (from 62 to 27%) and 41% of geranylgeranyl pyrophosphate was used for induction of pimaradiene intermediate as the major phytoalexin. In comparison, light-illumination used almost all geranylgeranyl pyrophosphate (96%) for gibberellin biosynthesis to stimulate the plant growth and strongly repressed the metabolic flux for phytoalexins biosynthesis. PMID:16215852

  2. The role of the siderophore pyridine-2,6-bis (thiocarboxylic acid) (PDTC) in zinc utilization by Pseudomonas putida DSM 3601.

    PubMed

    Leach, Lynne H; Morris, James C; Lewis, Thomas A

    2007-10-01

    Previous work had suggested that in addition to serving the function of a siderophore, pyridine-2,6-bis(thiocarboxylic acid) (PDTC) may also provide producing organisms with the ability to assimilate other divalent transition metals. This was tested further by examining regulation of siderophore production, expression of pdt genes, and growth in response to added zinc. In media containing 10-50 microM ZnCl2, the production of PDTC was found to be differentially repressed, as compared with the production of pyoverdine. The expression of PdtK, the outer membrane receptor involved in PDTC transport, was also reduced in response to added zinc whereas other iron-regulated outer membrane proteins were not. Expression of a chromosomal pdtI: xylE fusion was repressed to a similar extent in response to zinc or iron. Mutants that cannot produce PDTC did not show a growth enhancement with micromolar concentrations of zinc as seen in the wild type strain. The phenotype of the mutant strains was suppressed by the addition of PDTC. The outer membrane receptor and inner membrane permease components of PDTC utilization were necessary for relief of chelator (1,10-phenanthroline)-induced growth inhibition by Zn:PDTC. Iron uptake from 55Fe:PDTC was not affected by a 32-fold molar excess of Zn:PDTC. The data indicate that zinc present as Zn:PDTC can be utilized by strains possessing PDTC utilization functions but that transport is much less efficient than for Fe:PDTC. PMID:17066327

  3. A High-Content, Phenotypic Screen Identifies Fluorouridine as an Inhibitor of Pyoverdine Biosynthesis and Pseudomonas aeruginosa Virulence

    PubMed Central

    Kirienko, Daniel R.; Revtovich, Alexey V.

    2016-01-01

    ABSTRACT Pseudomonas aeruginosa is an opportunistic pathogen that causes severe health problems. Despite intensive investigation, many aspects of microbial virulence remain poorly understood. We used a high-throughput, high-content, whole-organism, phenotypic screen to identify small molecules that inhibit P. aeruginosa virulence in Caenorhabditis elegans. Approximately half of the hits were known antimicrobials. A large number of hits were nonantimicrobial bioactive compounds, including the cancer chemotherapeutic 5-fluorouracil. We determined that 5-fluorouracil both transiently inhibits bacterial growth and reduces pyoverdine biosynthesis. Pyoverdine is a siderophore that regulates the expression of several virulence determinants and is critical for pathogenesis in mammals. We show that 5-fluorouridine, a downstream metabolite of 5-fluorouracil, is responsible for inhibiting pyoverdine biosynthesis. We also show that 5-fluorouridine, in contrast to 5-fluorouracil, is a genuine antivirulence compound, with no bacteriostatic or bactericidal activity. To our knowledge, this is the first report utilizing a whole-organism screen to identify novel compounds with antivirulent properties effective against P. aeruginosa. IMPORTANCE Despite intense research effort from scientists and the advent of the molecular age of biomedical research, many of the mechanisms that underlie pathogenesis are still understood poorly, if at all. The opportunistic human pathogen Pseudomonas aeruginosa causes a variety of soft tissue infections and is responsible for over 50,000 hospital-acquired infections per year. In addition, P. aeruginosa exhibits a striking degree of innate and acquired antimicrobial resistance, complicating treatment. It is increasingly important to understand P. aeruginosa virulence. In an effort to gain this information in an unbiased fashion, we used a high-throughput phenotypic screen to identify small molecules that disrupted bacterial pathogenesis and

  4. A High-Content, Phenotypic Screen Identifies Fluorouridine as an Inhibitor of Pyoverdine Biosynthesis and Pseudomonas aeruginosa Virulence.

    PubMed

    Kirienko, Daniel R; Revtovich, Alexey V; Kirienko, Natalia V

    2016-01-01

    Pseudomonas aeruginosa is an opportunistic pathogen that causes severe health problems. Despite intensive investigation, many aspects of microbial virulence remain poorly understood. We used a high-throughput, high-content, whole-organism, phenotypic screen to identify small molecules that inhibit P. aeruginosa virulence in Caenorhabditis elegans. Approximately half of the hits were known antimicrobials. A large number of hits were nonantimicrobial bioactive compounds, including the cancer chemotherapeutic 5-fluorouracil. We determined that 5-fluorouracil both transiently inhibits bacterial growth and reduces pyoverdine biosynthesis. Pyoverdine is a siderophore that regulates the expression of several virulence determinants and is critical for pathogenesis in mammals. We show that 5-fluorouridine, a downstream metabolite of 5-fluorouracil, is responsible for inhibiting pyoverdine biosynthesis. We also show that 5-fluorouridine, in contrast to 5-fluorouracil, is a genuine antivirulence compound, with no bacteriostatic or bactericidal activity. To our knowledge, this is the first report utilizing a whole-organism screen to identify novel compounds with antivirulent properties effective against P. aeruginosa. IMPORTANCE Despite intense research effort from scientists and the advent of the molecular age of biomedical research, many of the mechanisms that underlie pathogenesis are still understood poorly, if at all. The opportunistic human pathogen Pseudomonas aeruginosa causes a variety of soft tissue infections and is responsible for over 50,000 hospital-acquired infections per year. In addition, P. aeruginosa exhibits a striking degree of innate and acquired antimicrobial resistance, complicating treatment. It is increasingly important to understand P. aeruginosa virulence. In an effort to gain this information in an unbiased fashion, we used a high-throughput phenotypic screen to identify small molecules that disrupted bacterial pathogenesis and increased host

  5. Computer aided gene mining for gingerol biosynthesis.

    PubMed

    James, Priyanka; Baby, Bincy; Charles, SonaSona; Nair, Lekshmysree Saraschandran; Nazeem, Puthiyaveetil Abdulla

    2015-01-01

    Inspite of the large body of genomic data obtained from the transcriptome of Zingiber officinale, very few studies have focused on the identification and characterization of miRNAs in gingerol biosynthesis. Zingiber officinale transcriptome was analyzed using EST dataset (38169 total) deposited in public domains. In this paper computational functional annotation of the available ESTs and identification of genes which play a significant role in gingerol biosynthesis are described. Zingiber officinale transcriptome was analyzed using EST dataset (38169 total) from ncbi. ESTs were clustered and assembled, resulting in 8624 contigs and 8821 singletons. Assembled dataset was then submitted to the EST functional annotation workflow including blast, gene ontology (go) analysis, and pathway enrichment by kyoto encyclopedia of genes and genomes (kegg) and interproscan. The unigene datasets were further exploited to identify simple sequence repeats that enable linkage mapping. A total of 409 simple sequence repeats were identified from the contigs. Furthermore we examined the existence of novel miRNAs from the ESTs in rhizome, root and leaf tissues. EST analysis revealed the presence of single hypothetical miRNA in rhizome tissue. The hypothetical miRNA is warranted to play an important role in controlling genes involved in gingerol biosynthesis and hence demands experimental validation. The assembly and associated information of transcriptome data provides a comprehensive functional and evolutionary characterization of genomics of Zingiber officinale. As an effort to make the genomic and transcriptomic data widely available to the public domain, the results were integrated into a web-based Ginger EST database which is freely accessible at http://www.kaubic.in/gingerest/. PMID:26229293

  6. The Biosynthesis of Capuramycin-type Antibiotics

    PubMed Central

    Cai, Wenlong; Goswami, Anwesha; Yang, Zhaoyong; Liu, Xiaodong; Green, Keith D.; Barnard-Britson, Sandra; Baba, Satoshi; Funabashi, Masanori; Nonaka, Koichi; Sunkara, Manjula; Morris, Andrew J.; Spork, Anatol P.; Ducho, Christian; Garneau-Tsodikova, Sylvie; Thorson, Jon S.; Van Lanen, Steven G.

    2015-01-01

    A-500359s, A-503083s, and A-102395 are capuramycin-type nucleoside antibiotics that were discovered using a screen to identify inhibitors of bacterial translocase I, an essential enzyme in peptidoglycan cell wall biosynthesis. Like the parent capuramycin, A-500359s and A-503083s consist of three structural components: a uridine-5′-carboxamide (CarU), a rare unsaturated hexuronic acid, and an aminocaprolactam, the last of which is substituted by an unusual arylamine-containing polyamide in A-102395. The biosynthetic gene clusters for A-500359s and A-503083s have been reported, and two genes encoding a putative non-heme Fe(II)-dependent α-ketoglutarate:UMP dioxygenase and an l-Thr:uridine-5′-aldehyde transaldolase were uncovered, suggesting that C–C bond formation during assembly of the high carbon (C6) sugar backbone of CarU proceeds from the precursors UMP and l-Thr to form 5′-C-glycyluridine (C7) as a biosynthetic intermediate. Here, isotopic enrichment studies with the producer of A-503083s were used to indeed establish l-Thr as the direct source of the carboxamide of CarU. With this knowledge, the A-102395 gene cluster was subsequently cloned and characterized. A genetic system in the A-102395-producing strain was developed, permitting the inactivation of several genes, including those encoding the dioxygenase (cpr19) and transaldolase (cpr25), which abolished the production of A-102395, thus confirming their role in biosynthesis. Heterologous production of recombinant Cpr19 and CapK, the transaldolase homolog involved in A-503083 biosynthesis, confirmed their expected function. Finally, a phosphotransferase (Cpr17) conferring self-resistance was functionally characterized. The results provide the opportunity to use comparative genomics along with in vivo and in vitro approaches to probe the biosynthetic mechanism of these intriguing structures. PMID:25855790

  7. Caste-Selective Pheromone Biosynthesis in Honeybees

    NASA Astrophysics Data System (ADS)

    Plettner, Erika; Slessor, Keith N.; Winston, Mark L.; Oliver, James E.

    1996-03-01

    Queen and worker honeybees (Apis mellifera L.) produce a caste-related blend of functionalized 8- and 10-carbon fatty acids in their mandibular glands. The biological functions of these compounds match the queen's reproductive and the worker's nonreproductive roles in the colony. Studies with deuterated substrates revealed that the biosynthesis of these acids begins with stearic acid, which is hydroxylated at the 17th or 18th position. The 18-carbon hydroxy acid chains are shortened, and the resulting 10-carbon hydroxy acids are oxidized in a caste-selective manner, thereby determining many of the functional differences between queens and workers.

  8. Developing New Antibiotics with Combinatorial Biosynthesis

    NASA Astrophysics Data System (ADS)

    Pohl, Nicola L.

    2000-11-01

    Polyketide synthases (PKSs), a class of enzymes found in soil bacteria that produce antibiotics such as erythromycin, string together acetate units using basic organic reactions. The manipulation of the sequence of these reactions at the genetic level has resulted in an alteration of the corresponding chemical structure of the antibiotic produced by the bacteria. This process, called combinatorial biosynthesis, allows the generation of many presently unknown complex structures that can be tested for antibacterial activity, thereby contributing to the race against antibiotic-resistant infectious bacteria.

  9. [Recent advances in lanthipeptide biosynthesis - A review].

    PubMed

    Mo, Tianlu; Xue, Lingui; Zhang, Qi

    2016-03-01

    Lanthipeptides are a growing class of ribosomally synthesized and posttranslationally modified peptide (RiPP) natural products. These compounds are widely distributed among taxonomically distant species, and their structures and biological activities are diverse, providing an important source for drug research and developement. In this review, we summarized the recent advances in the understanding of structure, classification, evolution and substrate-controlled biosynthetic mechanism of lanthipeptide, attempting to highlight the intriguing chemistry and enzymology in the biosynthesis of this growing family of natural products. PMID:27382781

  10. Biosynthesis of Gold Nanoparticles Using Pseudomonas Aeruginosa

    SciTech Connect

    Abd El-Aziz, M.; Badr, Y.; Mahmoud, M. A.

    2007-02-14

    Pseudomonas aeruginosa were used for extracellular biosynthesis of gold nanoparticles (Au NPs). Consequently, Au NPs were formed due to reduction of gold ion by bacterial cell supernatant of P. aeruginos ATCC 90271, P. aeruginos (2) and P. aeruginos (1). The UV-Vis. and fluorescence spectra of the bacterial as well as chemical prepared Au NPs were recorded. Transmission electron microscopy (TEM) micrograph showed the formation of well-dispersed gold nanoparticles in the range of 15-30 nm. The process of reduction being extracellular and may lead to the development of an easy bioprocess for synthesis of Au NPs.

  11. Carotenoid Biosynthesis in Arabidopsis: A Colorful Pathway

    PubMed Central

    Ruiz-Sola, M. Águila; Rodríguez-Concepción, Manuel

    2012-01-01

    Plant carotenoids are a family of pigments that participate in light harvesting and are essential for photoprotection against excess light. Furthermore, they act as precursors for the production of apocarotenoid hormones such as abscisic acid and strigolactones. In this review, we summarize the current knowledge on the genes and enzymes of the carotenoid biosynthetic pathway (which is now almost completely elucidated) and on the regulation of carotenoid biosynthesis at both transcriptional and post-transcriptional levels. We also discuss the relevance of Arabidopsis as a model system for the study of carotenogenesis and how metabolic engineering approaches in this plant have taught important lessons for carotenoid biotechnology. PMID:22582030

  12. Bioalteration of synthetic Fe(III)-, Fe(II)-bearing basaltic glasses and Fe-free glass in the presence of the heterotrophic bacteria strain Pseudomonas aeruginosa: Impact of siderophores

    NASA Astrophysics Data System (ADS)

    Perez, Anne; Rossano, Stéphanie; Trcera, Nicolas; Huguenot, David; Fourdrin, Chloé; Verney-Carron, Aurélie; van Hullebusch, Eric D.; Guyot, François

    2016-09-01

    This study aims to evaluate the role of micro-organisms and their siderophores in the first steps of the alteration processes of basaltic glasses in aqueous media. In this regard, three different types of glasses - with or without iron, in the reduced Fe(II) or oxidized Fe(III) states - were prepared on the basis of a simplified basaltic glass composition. Control and Pseudomonas aeruginosa inoculated experiments were performed in a buffered (pH 6.5) nutrient depleted medium to stimulate the production of the pyoverdine siderophore. Results show that the presence of P. aeruginosa has an effect on the dissolution kinetics of all glasses as most of the calculated elemental release rates are increased compared to sterile conditions. Reciprocally, the composition of the glass in contact with P. aeruginosa has an impact on the bacterial growth and siderophore production. As an essential nutrient for this microbial strain, Fe notably appears to play a central role during biotic experiments. Its presence in the glass stimulates the bacterial growth and minimizes the synthesis of pyoverdine. Moreover the initial Fe2+/Fe3+ ratio in the glasses modulates this synthesis, as pyoverdine is not detected at all in the system in contact with Fe(III)-bearing glass. Finally, the dissolution rates appear to be correlated to siderophore concentrations as they increase with respect to sterile experiments in the order Fe(III)-bearing glass < Fe(II)-bearing glass < Fe-free glass. This increase is attributed to complexation reactions between siderophores and Fe or Al for Fe(II)-bearing glass or Fe-free glass, respectively. The dissolution of an Fe-free glass is significantly improved in the presence of bacteria, as initial dissolution rates are increased by a factor of 3. This study attests to the essential role of siderophores in the P. aeruginosa-promoted dissolution processes of basaltic glasses as well as to the complex relationships between the nutritional potential of the glass and

  13. Ant trail pheromone biosynthesis is triggered by a neuropeptide hormone.

    PubMed

    Choi, Man-Yeon; Vander Meer, Robert K

    2012-01-01

    Our understanding of insect chemical communication including pheromone identification, synthesis, and their role in behavior has advanced tremendously over the last half-century. However, endocrine regulation of pheromone biosynthesis has progressed slowly due to the complexity of direct and/or indirect hormonal activation of the biosynthetic cascades resulting in insect pheromones. Over 20 years ago, a neurohormone, pheromone biosynthesis activating neuropeptide (PBAN) was identified that stimulated sex pheromone biosynthesis in a lepidopteran moth. Since then, the physiological role, target site, and signal transduction of PBAN has become well understood for sex pheromone biosynthesis in moths. Despite that PBAN-like peptides (∼200) have been identified from various insect Orders, their role in pheromone regulation had not expanded to the other insect groups except for Lepidoptera. Here, we report that trail pheromone biosynthesis in the Dufour's gland (DG) of the fire ant, Solenopsis invicta, is regulated by PBAN. RNAi knock down of PBAN gene (in subesophageal ganglia) or PBAN receptor gene (in DG) expression inhibited trail pheromone biosynthesis. Reduced trail pheromone was documented analytically and through a behavioral bioassay. Extension of PBAN's role in pheromone biosynthesis to a new target insect, mode of action, and behavioral function will renew research efforts on the involvement of PBAN in pheromone biosynthesis in Insecta. PMID:23226278

  14. Ant Trail Pheromone Biosynthesis Is Triggered by a Neuropeptide Hormone

    PubMed Central

    Choi, Man-Yeon; Vander Meer, Robert K.

    2012-01-01

    Our understanding of insect chemical communication including pheromone identification, synthesis, and their role in behavior has advanced tremendously over the last half-century. However, endocrine regulation of pheromone biosynthesis has progressed slowly due to the complexity of direct and/or indirect hormonal activation of the biosynthetic cascades resulting in insect pheromones. Over 20 years ago, a neurohormone, pheromone biosynthesis activating neuropeptide (PBAN) was identified that stimulated sex pheromone biosynthesis in a lepidopteran moth. Since then, the physiological role, target site, and signal transduction of PBAN has become well understood for sex pheromone biosynthesis in moths. Despite that PBAN-like peptides (∼200) have been identified from various insect Orders, their role in pheromone regulation had not expanded to the other insect groups except for Lepidoptera. Here, we report that trail pheromone biosynthesis in the Dufour's gland (DG) of the fire ant, Solenopsis invicta, is regulated by PBAN. RNAi knock down of PBAN gene (in subesophageal ganglia) or PBAN receptor gene (in DG) expression inhibited trail pheromone biosynthesis. Reduced trail pheromone was documented analytically and through a behavioral bioassay. Extension of PBAN's role in pheromone biosynthesis to a new target insect, mode of action, and behavioral function will renew research efforts on the involvement of PBAN in pheromone biosynthesis in Insecta. PMID:23226278

  15. Biosynthesis of nanoparticles using microbes- a review.

    PubMed

    Hulkoti, Nasreen I; Taranath, T C

    2014-09-01

    The biosynthesis of nanoparticles by microorganism is a green and eco-friendly technology. This review focuses on the use of consortium of diverse microorganisms belonging to both prokaryotes and eukaryotes for the synthesis of metallic nanoparticles viz. silver, gold, platinum, zirconium, palladium, iron, cadmium and metal oxides such as titanium oxide, zinc oxide, etc. These microorganisms include bacteria, actinomycetes, fungi and algae. The synthesis of nanoparticles may be intracellular or extracellular. The several workers have reported that NADH dependent nitrate reductase enzyme plays a vital role in the conversion of metallic ions to nanoparticles. The FTIR study reveals that diverse biomolecules viz. carboxyl group, primary and secondary amines, amide I, II, and III bands etc serve as a tool for bioreduction and capping agents there by offering stability to particles by preventing agglomeration and growth. The size and shape of the nanoparticles vary with the organism employed and conditions employed during the synthesis which included pH, temperature and substrate concentration. The microorganisms provide diverse environment for biosynthesis of nanoparticles. These particles are safe and eco-friendly with a lot of applications in medicine, agriculture, cosmetic industry, drug delivery and biochemical sensors. The challenges for redressal include optimal production and minimal time to obtain desired size and shape, to enhance the stability of nanoparticles and optimization of specific microorganisms for specific application. PMID:25001188

  16. Brassinosteroid biosynthesis and signalling in Petunia hybrida.

    PubMed

    Verhoef, Nathalie; Yokota, Takao; Shibata, Kyomi; de Boer, Gert-Jan; Gerats, Tom; Vandenbussche, Michiel; Koes, Ronald; Souer, Erik

    2013-05-01

    Brassinosteroids (BRs) are steroidal plant hormones that play an important role in the growth and development of plants. The biosynthesis of sterols and BRs as well as the signalling cascade they induce in plants have been elucidated largely through metabolic studies and the analysis of mutants in Arabidopsis and rice. Only fragmentary details about BR signalling in other plant species are known. Here a forward genetics strategy was used in Petunia hybrida, by which 19 families with phenotypic alterations typical for BR deficiency mutants were identified. In all mutants, the endogenous BR levels were severely reduced. In seven families, the tagged genes were revealed as the petunia BR biosynthesis genes CYP90A1 and CYP85A1 and the BR receptor gene BRI1. In addition, several homologues of key regulators of the BR signalling pathway were cloned from petunia based on homology with their Arabidopsis counterparts, including the BRI1 receptor, a member of the BES1/BZR1 transcription factor family (PhBEH2), and two GSK3-like kinases (PSK8 and PSK9). PhBEH2 was shown to interact with PSK8 and 14-3-3 proteins in yeast, revealing similar interactions to those during BR signalling in Arabidopsis. Interestingly, PhBEH2 also interacted with proteins implicated in other signalling pathways. This suggests that PhBEH2 might function as an important hub in the cross-talk between diverse signalling pathways. PMID:23599276

  17. Biosynthesis of archaeal membrane ether lipids

    PubMed Central

    Jain, Samta; Caforio, Antonella; Driessen, Arnold J. M.

    2014-01-01

    A vital function of the cell membrane in all living organism is to maintain the membrane permeability barrier and fluidity. The composition of the phospholipid bilayer is distinct in archaea when compared to bacteria and eukarya. In archaea, isoprenoid hydrocarbon side chains are linked via an ether bond to the sn-glycerol-1-phosphate backbone. In bacteria and eukarya on the other hand, fatty acid side chains are linked via an ester bond to the sn-glycerol-3-phosphate backbone. The polar head groups are globally shared in the three domains of life. The unique membrane lipids of archaea have been implicated not only in the survival and adaptation of the organisms to extreme environments but also to form the basis of the membrane composition of the last universal common ancestor (LUCA). In nature, a diverse range of archaeal lipids is found, the most common are the diether (or archaeol) and the tetraether (or caldarchaeol) lipids that form a monolayer. Variations in chain length, cyclization and other modifications lead to diversification of these lipids. The biosynthesis of these lipids is not yet well understood however progress in the last decade has led to a comprehensive understanding of the biosynthesis of archaeol. This review describes the current knowledge of the biosynthetic pathway of archaeal ether lipids; insights on the stability and robustness of archaeal lipid membranes; and evolutionary aspects of the lipid divide and the LUCA. It examines recent advances made in the field of pathway reconstruction in bacteria. PMID:25505460

  18. Fatty acid biosynthesis in pea root plastids

    SciTech Connect

    Stahl, R.J.; Sparace, S.A. )

    1989-04-01

    Fatty acid biosynthesis from (1-{sup 14}C)acetate was optimized in plastids isolated from primary root tips of 7-day-old germinating pea seeds. Fatty acid synthesis was maximum at approximately 80 nmoles/hr/mg protein in the presence of 200 {mu}M acetate, 0.5 mM each of NADH, NADPH and CoA, 6 mM each of ATP and MgCl{sub 2}, 1 mM each of the MnCl{sub 2} and glycerol-3-phosphate, 15 mM KHCO{sub 3}, and 0.1M Bis-tris-propane, pH 8.0 incubated at 35C. At the standard incubation temperature of 25C, fatty acid synthesis was linear from up to 6 hours with 80 to 100 {mu}g/mL plastid protein. ATP and CoA were absolute requirements, whereas KHCO{sub 3}, divalent cations and reduced nucleotides all improved activity by 80 to 85%. Mg{sup 2+} and NADH were the preferred cation and nucleotide, respectively. Dithiothreitol and detergents were generally inhibitory. The radioactive products of fatty acid biosynthesis were approximately 33% 16:0, 10% 18:0 and 56% 18:1 and generally did not vary with increasing concentrations of each cofactor.

  19. Essences in Metabolic Engineering of Lignan Biosynthesis

    PubMed Central

    Satake, Honoo; Koyama, Tomotsugu; Bahabadi, Sedigheh Esmaeilzadeh; Matsumoto, Erika; Ono, Eiichiro; Murata, Jun

    2015-01-01

    Lignans are structurally and functionally diverse phytochemicals biosynthesized in diverse plant species and have received wide attentions as leading compounds of novel drugs for tumor treatment and healthy diets to reduce of the risks of lifestyle-related non-communicable diseases. However, the lineage-specific distribution and the low-amount of production in natural plants, some of which are endangered species, hinder the efficient and stable production of beneficial lignans. Accordingly, the development of new procedures for lignan production is of keen interest. Recent marked advances in the molecular and functional characterization of lignan biosynthetic enzymes and endogenous and exogenous factors for lignan biosynthesis have suggested new methods for the metabolic engineering of lignan biosynthesis cascades leading to the efficient, sustainable, and stable lignan production in plants, including plant cell/organ cultures. Optimization of light conditions, utilization of a wide range of elicitor treatments, and construction of transiently gene-transfected or transgenic lignan-biosynthesizing plants are mainly being attempted. This review will present the basic and latest knowledge regarding metabolic engineering of lignans based on their biosynthetic pathways and biological activities, and the perspectives in lignan production via metabolic engineering. PMID:25946459

  20. Benzylisoquinoline alkaloid biosynthesis in opium poppy.

    PubMed

    Beaudoin, Guillaume A W; Facchini, Peter J

    2014-07-01

    Opium poppy (Papaver somniferum) is one of the world's oldest medicinal plants and remains the only commercial source for the narcotic analgesics morphine, codeine and semi-synthetic derivatives such as oxycodone and naltrexone. The plant also produces several other benzylisoquinoline alkaloids with potent pharmacological properties including the vasodilator papaverine, the cough suppressant and potential anticancer drug noscapine and the antimicrobial agent sanguinarine. Opium poppy has served as a model system to investigate the biosynthesis of benzylisoquinoline alkaloids in plants. The application of biochemical and functional genomics has resulted in a recent surge in the discovery of biosynthetic genes involved in the formation of major benzylisoquinoline alkaloids in opium poppy. The availability of extensive biochemical genetic tools and information pertaining to benzylisoquinoline alkaloid metabolism is facilitating the study of a wide range of phenomena including the structural biology of novel catalysts, the genomic organization of biosynthetic genes, the cellular and sub-cellular localization of biosynthetic enzymes and a variety of biotechnological applications. In this review, we highlight recent developments and summarize the frontiers of knowledge regarding the biochemistry, cellular biology and biotechnology of benzylisoquinoline alkaloid biosynthesis in opium poppy. PMID:24671624

  1. Monoterpene biosynthesis potential of plant subcellular compartments.

    PubMed

    Dong, Lemeng; Jongedijk, Esmer; Bouwmeester, Harro; Van Der Krol, Alexander

    2016-01-01

    Subcellular monoterpene biosynthesis capacity based on local geranyl diphosphate (GDP) availability or locally boosted GDP production was determined for plastids, cytosol and mitochondria. A geraniol synthase (GES) was targeted to plastids, cytosol, or mitochondria. Transient expression in Nicotiana benthamiana indicated local GDP availability for each compartment but resulted in different product levels. A GDP synthase from Picea abies (PaGDPS1) was shown to boost GDP production. PaGDPS1 was also targeted to plastids, cytosol or mitochondria and PaGDPS1 and GES were coexpressed in all possible combinations. Geraniol and geraniol-derived products were analyzed by GC-MS and LC-MS, respectively. GES product levels were highest for plastid-targeted GES, followed by mitochondrial- and then cytosolic-targeted GES. For each compartment local boosting of GDP biosynthesis increased GES product levels. GDP exchange between compartments is not equal: while no GDP is exchanged from the cytosol to the plastids, 100% of GDP in mitochondria can be exchanged to plastids, while only 7% of GDP from plastids is available for mitochondria. This suggests a direct exchange mechanism for GDP between plastids and mitochondria. Cytosolic PaGDPS1 competes with plastidial GES activity, suggesting an effective drain of isopentenyl diphosphate from the plastids to the cytosol. PMID:26356766

  2. Phytogenic biosynthesis and emission of methyl acetate.

    PubMed

    Jardine, Kolby; Wegener, Frederik; Abrell, Leif; van Haren, Joost; Werner, Christiane

    2014-02-01

    Acetylation of plant metabolites fundamentally changes their volatility, solubility and activity as semiochemicals. Here we present a new technique termed dynamic (13) C-pulse chasing to track the fate of C1-3 carbon atoms of pyruvate into the biosynthesis and emission of methyl acetate (MA) and CO2 . (13) C-labelling of MA and CO2 branch emissions respond within minutes to changes in (13) C-positionally labelled pyruvate solutions fed through the transpiration stream. Strong (13) C-labelling of MA emissions occurred only under pyruvate-2-(13) C and pyruvate-2,3-(13) C feeding, but not pyruvate-1-(13) C feeding. In contrast, strong (13) CO2 emissions were only observed under pyruvate-1-(13) C feeding. These results demonstrate that MA (and other volatile and non-volatile metabolites) derive from the C2,3 atoms of pyruvate while the C1 atom undergoes decarboxylation. The latter is a non-mitochondrial source of CO2 in the light generally not considered in studies of CO2 sources and sinks. Within a tropical rainforest mesocosm, we also observed atmospheric concentrations of MA up to 0.6 ppbv that tracked light and temperature conditions. Moreover, signals partially attributed to MA were observed in ambient air within and above a tropical rainforest in the Amazon. Our study highlights the potential importance of acetyl coenzyme A (CoA) biosynthesis as a source of acetate esters and CO2 to the atmosphere. PMID:23862653

  3. Engineering the MEP pathway enhanced ajmalicine biosynthesis.

    PubMed

    Chang, Kai; Qiu, Fei; Chen, Min; Zeng, Lingjiang; Liu, Xiaoqiang; Yang, Chunxian; Lan, Xiaozhong; Wang, Qiang; Liao, Zhihua

    2014-01-01

    The 2-C-methyl-D-erythritol-4-phosphate (MEP) pathway genes encoding DXR and MECS from Taxus species and STR from Catharanthus roseus were used to genetically modify the ajmalicine biosynthetic pathway in hairy root cultures of C. roseus. As expected, the STR-overexpressed root cultures showed twofold higher accumulation of ajmalicine than the control. It was important to discover that overexpression of the single DXR or MECS gene from the MEP pathway also remarkably enhanced ajmalicine biosynthesis in transgenic hairy root cultures, and this suggested that engineering the MEP pathway by overexpression of DXR or MECS promoted the metabolic flux into ajmalicine biosynthesis. The transgenic hairy root cultures with co-overexpression of DXR and STR or MECS and STR had higher levels of ajmalicine than those with overexpression of a single gene alone such as DXR, MECS, and STR. It could be concluded that transgenic hairy root cultures harboring both DXR/MECS and STR possessed an increased flux in the terpenoid indole alkaloid biosynthetic pathway that enhanced ajmalicine yield, which was more efficient than cultures harboring only one of the three genes. PMID:24237015

  4. Brassinosteroid biosynthesis and signalling in Petunia hybrida

    PubMed Central

    Verhoef, Nathalie; Yokota, Takao; Shibata, Kyomi; de Boer, Gert-Jan; Gerats, Tom; Vandenbussche, Michiel; Koes, Ronald; Souer, Erik

    2013-01-01

    Brassinosteroids (BRs) are steroidal plant hormones that play an important role in the growth and development of plants. The biosynthesis of sterols and BRs as well as the signalling cascade they induce in plants have been elucidated largely through metabolic studies and the analysis of mutants in Arabidopsis and rice. Only fragmentary details about BR signalling in other plant species are known. Here a forward genetics strategy was used in Petunia hybrida, by which 19 families with phenotypic alterations typical for BR deficiency mutants were identified. In all mutants, the endogenous BR levels were severely reduced. In seven families, the tagged genes were revealed as the petunia BR biosynthesis genes CYP90A1 and CYP85A1 and the BR receptor gene BRI1. In addition, several homologues of key regulators of the BR signalling pathway were cloned from petunia based on homology with their Arabidopsis counterparts, including the BRI1 receptor, a member of the BES1/BZR1 transcription factor family (PhBEH2), and two GSK3-like kinases (PSK8 and PSK9). PhBEH2 was shown to interact with PSK8 and 14-3-3 proteins in yeast, revealing similar interactions to those during BR signalling in Arabidopsis. Interestingly, PhBEH2 also interacted with proteins implicated in other signalling pathways. This suggests that PhBEH2 might function as an important hub in the cross-talk between diverse signalling pathways. PMID:23599276

  5. Essences in metabolic engineering of lignan biosynthesis.

    PubMed

    Satake, Honoo; Koyama, Tomotsugu; Bahabadi, Sedigheh Esmaeilzadeh; Matsumoto, Erika; Ono, Eiichiro; Murata, Jun

    2015-01-01

    Lignans are structurally and functionally diverse phytochemicals biosynthesized in diverse plant species and have received wide attentions as leading compounds of novel drugs for tumor treatment and healthy diets to reduce of the risks of lifestyle-related non-communicable diseases. However, the lineage-specific distribution and the low-amount of production in natural plants, some of which are endangered species, hinder the efficient and stable production of beneficial lignans. Accordingly, the development of new procedures for lignan production is of keen interest. Recent marked advances in the molecular and functional characterization of lignan biosynthetic enzymes and endogenous and exogenous factors for lignan biosynthesis have suggested new methods for the metabolic engineering of lignan biosynthesis cascades leading to the efficient, sustainable, and stable lignan production in plants, including plant cell/organ cultures. Optimization of light conditions, utilization of a wide range of elicitor treatments, and construction of transiently gene-transfected or transgenic lignan-biosynthesizing plants are mainly being attempted. This review will present the basic and latest knowledge regarding metabolic engineering of lignans based on their biosynthetic pathways and biological activities, and the perspectives in lignan production via metabolic engineering. PMID:25946459

  6. Molecular Regulation of Antibiotic Biosynthesis in Streptomyces

    PubMed Central

    Liu, Gang; Chandra, Govind; Niu, Guoqing

    2013-01-01

    SUMMARY Streptomycetes are the most abundant source of antibiotics. Typically, each species produces several antibiotics, with the profile being species specific. Streptomyces coelicolor, the model species, produces at least five different antibiotics. We review the regulation of antibiotic biosynthesis in S. coelicolor and other, nonmodel streptomycetes in the light of recent studies. The biosynthesis of each antibiotic is specified by a large gene cluster, usually including regulatory genes (cluster-situated regulators [CSRs]). These are the main point of connection with a plethora of generally conserved regulatory systems that monitor the organism's physiology, developmental state, population density, and environment to determine the onset and level of production of each antibiotic. Some CSRs may also be sensitive to the levels of different kinds of ligands, including products of the pathway itself, products of other antibiotic pathways in the same organism, and specialized regulatory small molecules such as gamma-butyrolactones. These interactions can result in self-reinforcing feed-forward circuitry and complex cross talk between pathways. The physiological signals and regulatory mechanisms may be of practical importance for the activation of the many cryptic secondary metabolic gene cluster pathways revealed by recent sequencing of numerous Streptomyces genomes. PMID:23471619

  7. First step of glycosylphosphatidylinositol (GPI) biosynthesis cross-talks with ergosterol biosynthesis and Ras signaling in Candida albicans.

    PubMed

    Yadav, Bhawna; Bhatnagar, Shilpi; Ahmad, Mohammad Faiz; Jain, Priyanka; Pratyusha, Vavilala A; Kumar, Pravin; Komath, Sneha Sudha

    2014-02-01

    Candida albicans is a leading cause of fungal infections worldwide. It has several glycosylphosphatidylinositol (GPI)-anchored virulence factors. Inhibiting GPI biosynthesis attenuates its virulence. Building on our previous work, we explore the interaction of GPI biosynthesis in C. albicans with ergosterol biosynthesis and hyphal morphogenesis. This study is also the first report of transcriptional co-regulation existing between two subunits of the multisubunit enzyme complex, GPI-N-acetylglucosaminyltransferase (GPI-GnT), involved in the first step of GPI anchor biosynthesis in eukaryotes. Using mutational analysis, we show that the accessory subunits, GPI2 and GPI19, of GPI-GnT exhibit opposite effects on ergosterol biosynthesis and Ras signaling (which determines hyphal morphogenesis). This is because the two subunits negatively regulate one another; GPI19 mutants show up-regulation of GPI2, whereas GPI2 mutants show up-regulation of GPI19. Two different models were examined as follows. First, the two GPI-GnT subunits independently interact with ergosterol biosynthesis and Ras signaling. Second, the two subunits mutually regulate one another and thereby regulate sterol levels and Ras signaling. Analysis of double mutants of these subunits indicates that GPI19 controls ergosterol biosynthesis through ERG11 levels, whereas GPI2 determines the filamentation by cross-talk with Ras1 signaling. Taken together, this suggests that the first step of GPI biosynthesis talks to and regulates two very important pathways in C. albicans. This could have implications for designing new antifungal strategies. PMID:24356967

  8. Role of the small RNA RyhB in the Fur regulon in mediating the capsular polysaccharide biosynthesis and iron acquisition systems in Klebsiella pneumoniae

    PubMed Central

    2012-01-01

    Background The capsular polysaccharide (CPS) and iron acquisition systems are important determinants of Klebsiella pneumoniae infections, and we have previously reported that the ferric uptake repressor (Fur) can play dual role in iron acquisition and CPS biosynthesis. In many bacteria, Fur negatively controls the transcription of the small non-coding RNA RyhB to modulate cellular functions and virulence. However, in K. pneumoniae, the role played by RyhB in the Fur regulon has not been characterised. This study investigated Fur regulation of ryhB transcription and the functional role of RyhB in K. pneumoniae. Results Deletion of fur from K. pneumoniae increased the transcription of ryhB; the electric mobility shift assay and the Fur-titration assay revealed that Fur could bind to the promoter region of ryhB, suggesting that Fur directly represses ryhB transcription. Additionally, in a Δfur strain with elevated CPS production, deletion of ryhB obviously reduced CPS production. The following promoter-reporter assay and quantitative real-time PCR of cps genes verified that RyhB activated orf1 and orf16 transcription to elevate CPS production. However, deletion of ryhB did not affect the mRNA levels of rcsA, rmpA, or rmpA2. These results imply that Fur represses the transcription of ryhB to mediate the biosynthesis of CPS, which is independent of RcsA, RmpA, and RmpA2. In addition, the Δfur strain’s high level of serum resistance was attenuated by the deletion of ryhB, indicating that RyhB plays a positive role in protecting the bacterium from serum killing. Finally, deletion of ryhB in Δfur reduced the expression of several genes corresponding to 3 iron acquisition systems in K. pneumoniae, and resulted in reduced siderophore production. Conclusions The regulation and functional role of RyhB in K. pneumoniae is characterized in this study. RyhB participates in Fur regulon to modulate the bacterial CPS biosynthesis and iron acquisition systems in K. pneumoniae

  9. Starch Biosynthesis in Developing Wheat Grain 1

    PubMed Central

    Keeling, Peter L.; Wood, John R.; Tyson, R. Huw; Bridges, Ian G.

    1988-01-01

    We have used 13C-labeled sugars and nuclear magnetic resonance (NMR) spectrometry to study the metabolic pathway of starch biosynthesis in developing wheat grain (Triticum aestivum cv Mardler). Our aim was to examine the extent of redistribution of 13C between carbons atoms 1 and 6 of [1-13C] or [6-13C]glucose (or fructose) incorporated into starch, and hence provide evidence for or against the involvement of triose phosphates in the metabolic pathway. Starch synthesis in the endosperm tissue was studied in two experimental systems. First, the 13C sugars were supplied to isolated endosperm tissue incubated in vitro, and second the 13C sugars were supplied in vivo to the intact plant. The 13C starch produced by the endosperm tissue of the grain was isolated and enzymically degraded to glucose using amyloglucosidase, and the distribution of 13C in all glucosyl carbons was quantified by 13C-NMR spectrometry. In all of the experiments, irrespective of the incubation time or incubation conditions, there was a similar pattern of partial (between 15 and 20%) redistribution of label between carbons 1 and 6 of glucose recovered from starch. There was no detectable increase over background 13C incidence in carbons 2 to 5. Within each experiment, the same pattern of partial redistribution of label was found in the glucosyl and fructosyl moieties of sucrose extracted from the tissue. Since it is unlikely that sucrose is present in the amyloplast, we suggest that the observed redistribution of label occurred in the cytosolic compartment of the endosperm cells and that both sucrose and starch are synthesized from a common pool of intermediates, such as hexose phosphate. We suggest that redistribution of label occurs via a cytosolic pathway cycle involving conversion of hexose phosphate to triose phosphate, interconversion of triose phosphate by triose phosphate isomerase, and resynthesis of hexose phosphate in the cytosol. A further round of triose phosphate interconversion in

  10. Auxin Biosynthesis: Are the Indole-3-Acetic Acid and Phenylacetic Acid Biosynthesis Pathways Mirror Images?

    PubMed

    Cook, Sam D; Nichols, David S; Smith, Jason; Chourey, Prem S; McAdam, Erin L; Quittenden, Laura; Ross, John J

    2016-06-01

    The biosynthesis of the main auxin in plants (indole-3-acetic acid [IAA]) has been elucidated recently and is thought to involve the sequential conversion of Trp to indole-3-pyruvic acid to IAA However, the pathway leading to a less well studied auxin, phenylacetic acid (PAA), remains unclear. Here, we present evidence from metabolism experiments that PAA is synthesized from the amino acid Phe, via phenylpyruvate. In pea (Pisum sativum), the reverse reaction, phenylpyruvate to Phe, is also demonstrated. However, despite similarities between the pathways leading to IAA and PAA, evidence from mutants in pea and maize (Zea mays) indicate that IAA biosynthetic enzymes are not the main enzymes for PAA biosynthesis. Instead, we identified a putative aromatic aminotransferase (PsArAT) from pea that may function in the PAA synthesis pathway. PMID:27208245

  11. Lanosterol biosynthesis in the prokaryote Methylococcus capsulatus: insight into the evolution of sterol biosynthesis.

    PubMed

    Lamb, David C; Jackson, Colin J; Warrilow, Andrew G S; Manning, Nigel J; Kelly, Diane E; Kelly, Steven L

    2007-08-01

    A putative operon containing homologues of essential eukaryotic sterol biosynthetic enzymes, squalene monooxygenase and oxidosqualene cyclase, has been identified in the genome of the prokaryote Methylococcus capsulatus. Expression of the squalene monooxygenase yielded a protein associated with the membrane fraction, while expression of oxidosqualene cyclase yielded a soluble protein, contrasting with the eukaryotic enzyme forms. Activity studies with purified squalene monooxygenase revealed a catalytic activity in epoxidation of 0.35 nmol oxidosqualene produced/min/nmol squalene monooxygenase, while oxidosqualene cyclase catalytic activity revealed cyclization of oxidosqualene to lanosterol with 0.6 nmol lanosterol produced/min/nmol oxidosqualene cyclase and no other products observed. The presence of prokaryotic sterol biosynthesis is still regarded as rare, and these are the first representatives of such prokaryotic enzymes to be studied, providing new insight into the evolution of sterol biosynthesis in general. PMID:17567593

  12. A Novel Muconic Acid Biosynthesis Approach by Shunting Tryptophan Biosynthesis via Anthranilate

    PubMed Central

    Sun, Xinxiao; Lin, Yuheng; Huang, Qin; Yuan, Qipeng

    2013-01-01

    Muconic acid is the synthetic precursor of adipic acid, and the latter is an important platform chemical that can be used for the production of nylon-6,6 and polyurethane. Currently, the production of adipic acid relies mainly on chemical processes utilizing petrochemicals, such as benzene, which are generally considered environmentally unfriendly and nonrenewable, as starting materials. Microbial synthesis from renewable carbon sources provides a promising alternative under the circumstance of petroleum depletion and environment deterioration. Here we devised a novel artificial pathway in Escherichia coli for the biosynthesis of muconic acid, in which anthranilate, the first intermediate in the tryptophan biosynthetic branch, was converted to catechol and muconic acid by anthranilate 1,2-dioxygenase (ADO) and catechol 1,2-dioxygenase (CDO), sequentially and respectively. First, screening for efficient ADO and CDO from different microbial species enabled the production of gram-per-liter level muconic acid from supplemented anthranilate in 5 h. To further achieve the biosynthesis of muconic acid from simple carbon sources, anthranilate overproducers were constructed by overexpressing the key enzymes in the shikimate pathway and blocking tryptophan biosynthesis. In addition, we found that introduction of a strengthened glutamine regeneration system by overexpressing glutamine synthase significantly improved anthranilate production. Finally, the engineered E. coli strain carrying the full pathway produced 389.96 ± 12.46 mg/liter muconic acid from simple carbon sources in shake flask experiments, a result which demonstrates scale-up potential for microbial production of muconic acid. PMID:23603682

  13. Biosynthesis and Heterologous Production of Epothilones

    NASA Astrophysics Data System (ADS)

    Müller, Rolf

    Although a variety of chemical syntheses for the epothilones and various derivatives have been described, modifying the backbone of those natural products remains a major challenge. One alternative to chemical alteration is the elucidation and subsequent manipulation of the biosynthetic pathway via genetic engineering in the producing organism. This type of approach is known as “combinatorial biosynthesis” and holds great promise, especially in conjunction with semi-synthesis methods to alter the structure of the natural product. In parallel, production can be optimized in the natural producer if the regulatory mechanisms governing the biosynthesis are understood. Alternatively, the entire gene cluster can be transferred into a heterologous host, more amenable both to genetic alteration and overexpression.

  14. Biosynthesis of cucurbitacins in Bryonia dioica seedlings.

    PubMed

    Cattel, L; Balliano, G; Caputo, O; Viola, F

    1981-04-01

    The biosynthesis of cucurbitacins during the seed germination of Bryonia dioica was studied by analysis of the cucurbitacin-triterpenoid fraction and by tracer experiments with acetate-[2- (14)C]. Isolation of 10alpha-cucurbita-5,24-dien-3beta-ol (9a), the simplest tetracyclic triterpene with a cucurbitane skeleton, supports the view that (9a) is the general precursor of cucurbitacins. Moreover, following the tracer experiments, cucurbitacin E (1a) was the first cucurbitacin formed, whereas the less oxygenated bryodulcosigenin (4a) was not detectable during germination of the plant. In the course of the present investigation, a new pentacyclic triterpene, isomultiflorenol (11a) (possible precursor of bryonolic acid (5a)), was also isolated. PMID:17401854

  15. A Molecular Description of Cellulose Biosynthesis

    PubMed Central

    McNamara, Joshua T.; Morgan, Jacob L.W.; Zimmer, Jochen

    2016-01-01

    Cellulose is the most abundant biopolymer on Earth, and certain organisms from bacteria to plants and animals synthesize cellulose as an extracellular polymer for various biological functions. Humans have used cellulose for millennia as a material and an energy source, and the advent of a lignocellulosic fuel industry will elevate it to the primary carbon source for the burgeoning renewable energy sector. Despite the biological and societal importance of cellulose, the molecular mechanism by which it is synthesized is now only beginning to emerge. On the basis of recent advances in structural and molecular biology on bacterial cellulose synthases, we review emerging concepts of how the enzymes polymerize glucose molecules, how the nascent polymer is transported across the plasma membrane, and how bacterial cellulose biosynthesis is regulated during biofilm formation. Additionally, we review evolutionary commonalities and differences between cellulose synthases that modulate the nature of the cellulose product formed. PMID:26034894

  16. Glycerolipid biosynthesis in isolated pea root plastids

    SciTech Connect

    Xue, Lingru; Sparace, S.A. )

    1990-05-01

    Plastids have been isolated from germinating pea (Pisum sativum L.) roots by differential centrifugation and purified on Percoll gradients. Marker enzymes (NADPH: cytochrome c reductase, fumarase and fatty acid synthesis) indicate that greater than 50% of the plastids are recovered essentially free from mitochondrial and endoplasmic reticulum contamination. Fatty acids synthesized from ({sup 14}C)acetate by Percoll-purified plastids are primarily 16:0, 16:1 and 18:1. ({sup 14}C)Acetate-labelled fatty acids and ({sup 14}C)glycerol-3-phosphate are both readily incorporated into glycerolipid. Approximately 12% of the total activity for glycerolipid biosynthesis from glycerol-3-phosphate is recovered in the purified plastid fraction. Glycerolipids synthesized from these precursors are primarily TAG, DAG, PE, PG, PC, PI and PA. Acyl-CoA's also accumulate when acetate is the precursor.

  17. Metabolic model for diversity-generating biosynthesis.

    PubMed

    Tianero, Ma Diarey; Pierce, Elizabeth; Raghuraman, Shrinivasan; Sardar, Debosmita; McIntosh, John A; Heemstra, John R; Schonrock, Zachary; Covington, Brett C; Maschek, J Alan; Cox, James E; Bachmann, Brian O; Olivera, Baldomero M; Ruffner, Duane E; Schmidt, Eric W

    2016-02-16

    A conventional metabolic pathway leads to a specific product. In stark contrast, there are diversity-generating metabolic pathways that naturally produce different chemicals, sometimes of great diversity. We demonstrate that for one such pathway, tru, each ensuing metabolic step is slower, in parallel with the increasing potential chemical divergence generated as the pathway proceeds. Intermediates are long lived and accumulate progressively, in contrast with conventional metabolic pathways, in which the first step is rate-limiting and metabolic intermediates are short-lived. Understanding these fundamental differences enables several different practical applications, such as combinatorial biosynthesis, some of which we demonstrate here. We propose that these principles may provide a unifying framework underlying diversity-generating metabolism in many different biosynthetic pathways. PMID:26831074

  18. Biosynthesis of amphetamine analogs in plants.

    PubMed

    Hagel, Jillian M; Krizevski, Raz; Marsolais, Frédéric; Lewinsohn, Efraim; Facchini, Peter J

    2012-07-01

    Amphetamine analogs are produced by plants in the genus Ephedra and by Catha edulis, and include the widely used decongestants and appetite suppressants pseudoephedrine and ephedrine. A combination of yeast (Candida utilis or Saccharomyces cerevisiae) fermentation and subsequent chemical modification is used for the commercial production of these compounds. The availability of certain plant biosynthetic genes would facilitate the engineering of yeast strains capable of de novo pseudoephedrine and ephedrine biosynthesis. Chemical synthesis has yielded amphetamine analogs with myriad functional group substitutions and diverse pharmacological properties. The isolation of enzymes with the serendipitous capacity to accept novel substrates could allow the production of substituted amphetamines in synthetic biosystems. Here, we review the biology, biochemistry and biotechnological potential of amphetamine analogs in plants. PMID:22502775

  19. Biosurfactant Mediated Biosynthesis of Selected Metallic Nanoparticles

    PubMed Central

    Płaza, Grażyna A.; Chojniak, Joanna; Banat, Ibrahim M.

    2014-01-01

    Developing a reliable experimental protocol for the synthesis of nanomaterials is one of the challenging topics in current nanotechnology particularly in the context of the recent drive to promote green technologies in their synthesis. The increasing need to develop clean, nontoxic and environmentally safe production processes for nanoparticles to reduce environmental impact, minimize waste and increase energy efficiency has become essential in this field. Consequently, recent studies on the use of microorganisms in the synthesis of selected nanoparticles are gaining increased interest as they represent an exciting area of research with considerable development potential. Microorganisms are known to be capable of synthesizing inorganic molecules that are deposited either intra- or extracellularly. This review presents a brief overview of current research on the use of biosurfactants in the biosynthesis of selected metallic nanoparticles and their potential importance. PMID:25110864

  20. Human genetic disorders of sphingolipid biosynthesis.

    PubMed

    Astudillo, Leonardo; Sabourdy, Frédérique; Therville, Nicole; Bode, Heiko; Ségui, Bruno; Andrieu-Abadie, Nathalie; Hornemann, Thorsten; Levade, Thierry

    2015-01-01

    Monogenic defects of sphingolipid biosynthesis have been recently identified in human patients. These enzyme deficiencies affect the synthesis of sphingolipid precursors, ceramides or complex glycosphingolipids. They are transmitted as autosomal recessive or dominant traits, and their resulting phenotypes often replicate the abnormalities seen in murine models deficient for the corresponding enzymes. In quite good agreement with the known critical roles of sphingolipids in cells from the nervous system and the epidermis, these genetic defects clinically manifest as neurological disorders, including paraplegia, epilepsy or peripheral neuropathies, or present with ichthyosis. The present review summarizes the genetic alterations, biochemical changes and clinical symptoms of this new group of inherited metabolic disorders. Hypotheses regarding the molecular pathophysiology and potential treatments of these diseases are also discussed. PMID:25141825

  1. Sargassum myriocystum mediated biosynthesis of gold nanoparticles

    NASA Astrophysics Data System (ADS)

    Stalin Dhas, T.; Ganesh Kumar, V.; Stanley Abraham, L.; Karthick, V.; Govindaraju, K.

    2012-12-01

    Functionalized metal nanoparticles are unique in nature and are being developed for its specificity in drug targeting. In the present study, aqueous extract of Sargassum myriocystum is used for the biosynthesis of gold nanoparticles (AuNPs) by the reduction of chloroauric acid. The formation of nanoparticles reaction was complete within 15 min at 76 °C. The size, shape and elemental analysis of AuNPs were carried out using UV-visible absorption spectroscopy, FT-IR, TEM, SEM-EDAX, and XRD analysis. The newly formed AuNPs are stable, well-defined, polydispersed (triangular and spherical) and crystalline with an average size of 15 nm. The biomolecule involved in stabilizing AuNPs was identified using GC-MS.

  2. Flavones: From Biosynthesis to Health Benefits

    PubMed Central

    Jiang, Nan; Doseff, Andrea I.; Grotewold, Erich

    2016-01-01

    Flavones correspond to a flavonoid subgroup that is widely distributed in the plants, and which can be synthesized by different pathways, depending on whether they contain C- or O-glycosylation and hydroxylated B-ring. Flavones are emerging as very important specialized metabolites involved in plant signaling and defense, as well as key ingredients of the human diet, with significant health benefits. Here, we appraise flavone formation in plants, emphasizing the emerging theme that biosynthesis pathway determines flavone chemistry. Additionally, we briefly review the biological activities of flavones, both from the perspective of the functions that they play in biotic and abiotic plant interactions, as well as their roles as nutraceutical components of the human and animal diet. PMID:27338492

  3. Metabolic model for diversity-generating biosynthesis

    PubMed Central

    Tianero, Ma. Diarey; Pierce, Elizabeth; Raghuraman, Shrinivasan; Sardar, Debosmita; McIntosh, John A.; Heemstra, John R.; Schonrock, Zachary; Covington, Brett C.; Maschek, J. Alan; Cox, James E.; Bachmann, Brian O.; Olivera, Baldomero M.; Ruffner, Duane E.; Schmidt, Eric W.

    2016-01-01

    A conventional metabolic pathway leads to a specific product. In stark contrast, there are diversity-generating metabolic pathways that naturally produce different chemicals, sometimes of great diversity. We demonstrate that for one such pathway, tru, each ensuing metabolic step is slower, in parallel with the increasing potential chemical divergence generated as the pathway proceeds. Intermediates are long lived and accumulate progressively, in contrast with conventional metabolic pathways, in which the first step is rate-limiting and metabolic intermediates are short-lived. Understanding these fundamental differences enables several different practical applications, such as combinatorial biosynthesis, some of which we demonstrate here. We propose that these principles may provide a unifying framework underlying diversity-generating metabolism in many different biosynthetic pathways. PMID:26831074

  4. mitochondrial pathway for biosynthesis of lipid mediators

    PubMed Central

    Tyurina, Yulia Y.; Poloyac, Samuel M.; Tyurin, Vladimir A.; Kapralov, Alexander A.; Jiang, Jianfei; Anthonymuthu, Tamil Selvan; Kapralova, Valentina I.; Vikulina, Anna S.; Jung, Mi-Yeon; Epperly, Michael W.; Mohammadyani, Dariush; Klein-Seetharaman, Judith; Jackson, Travis C.; Kochanek, Patrick M.; Pitt, Bruce R.; Greenberger, Joel S.; Vladimirov, Yury A.; Bayır, Hülya; Kagan, Valerian E.

    2014-01-01

    The central role of mitochondria in metabolic pathways and in cell death mechanisms requires sophisticated signaling systems. Essential in this signaling process is an array of lipid mediators derived from polyunsaturated fatty acids. However, the molecular machinery for the production of oxygenated polyunsaturated fatty acids is localized in the cytosol and their biosynthesis has not been identified in mitochondria. Here we report that a range of diversified polyunsaturated molecular species derived from a mitochondria-specific phospholipid, cardiolipin, are oxidized by the intermembrane space hemoprotein, cytochrome c. We show that an assortment of oxygenated cardiolipin species undergoes phospholipase A2-catalyzed hydrolysis thus generating multiple oxygenated fatty acids, including well known lipid mediators. This represents a new biosynthetic pathway for lipid mediators. We demonstrate that this pathway including oxidation of polyunsaturated cardiolipins and accumulation of their hydrolysis products – oxygenated linoleic, arachidonic acids and monolyso-cardiolipins – is activated in vivo after acute tissue injury. PMID:24848241

  5. Terpenoids and Their Biosynthesis in Cyanobacteria

    PubMed Central

    Pattanaik, Bagmi; Lindberg, Pia

    2015-01-01

    Terpenoids, or isoprenoids, are a family of compounds with great structural diversity which are essential for all living organisms. In cyanobacteria, they are synthesized from the methylerythritol-phosphate (MEP) pathway, using glyceraldehyde 3-phosphate and pyruvate produced by photosynthesis as substrates. The products of the MEP pathway are the isomeric five-carbon compounds isopentenyl diphosphate and dimethylallyl diphosphate, which in turn form the basic building blocks for formation of all terpenoids. Many terpenoid compounds have useful properties and are of interest in the fields of pharmaceuticals and nutrition, and even potentially as future biofuels. The MEP pathway, its function and regulation, and the subsequent formation of terpenoids have not been fully elucidated in cyanobacteria, despite its relevance for biotechnological applications. In this review, we summarize the present knowledge about cyanobacterial terpenoid biosynthesis, both regarding the native metabolism and regarding metabolic engineering of cyanobacteria for heterologous production of non-native terpenoids. PMID:25615610

  6. A molecular description of cellulose biosynthesis.

    PubMed

    McNamara, Joshua T; Morgan, Jacob L W; Zimmer, Jochen

    2015-01-01

    Cellulose is the most abundant biopolymer on Earth, and certain organisms from bacteria to plants and animals synthesize cellulose as an extracellular polymer for various biological functions. Humans have used cellulose for millennia as a material and an energy source, and the advent of a lignocellulosic fuel industry will elevate it to the primary carbon source for the burgeoning renewable energy sector. Despite the biological and societal importance of cellulose, the molecular mechanism by which it is synthesized is now only beginning to emerge. On the basis of recent advances in structural and molecular biology on bacterial cellulose synthases, we review emerging concepts of how the enzymes polymerize glucose molecules, how the nascent polymer is transported across the plasma membrane, and how bacterial cellulose biosynthesis is regulated during biofilm formation. Additionally, we review evolutionary commonalities and differences between cellulose synthases that modulate the nature of the cellulose product formed. PMID:26034894

  7. Biosynthesis of the phytoalexin pisatin. [Pisum sativum

    SciTech Connect

    Preisig, C.L.; Bell, J.N.; Matthews, D.E.; VanEtten, H.D. ); Sun, Yuejin; Hrazdina, G. )

    1990-11-01

    NADPH-dependent reduction of 2{prime},7-dihydroxy-4{prime},5{prime}-methylenedioxyisoflavone to the isoflavanone sophorol, a proposed intermediate step in pisatin biosynthesis, was detected in extracts of Pisum sativum. This isoflavone reductase activity was inducible by treatment of pea seedlings with CuCl{sub 2}. The timing of induction coincided with that of the 6a-hydroxymaackiain 3-O-methyltransferase, which catalyzes the terminal biosynthetic step. Neither enzyme was light inducible. Further NADPH-dependent metabolism of sophorol by extracts of CuCl{sub 2}-treated seedlings was also observed; three products were radiolabeled when ({sup 3}H)sophorol was the substrate, one of which is tentatively identified as maackiain.

  8. Flavones: From Biosynthesis to Health Benefits.

    PubMed

    Jiang, Nan; Doseff, Andrea I; Grotewold, Erich

    2016-01-01

    Flavones correspond to a flavonoid subgroup that is widely distributed in the plants, and which can be synthesized by different pathways, depending on whether they contain C- or O-glycosylation and hydroxylated B-ring. Flavones are emerging as very important specialized metabolites involved in plant signaling and defense, as well as key ingredients of the human diet, with significant health benefits. Here, we appraise flavone formation in plants, emphasizing the emerging theme that biosynthesis pathway determines flavone chemistry. Additionally, we briefly review the biological activities of flavones, both from the perspective of the functions that they play in biotic and abiotic plant interactions, as well as their roles as nutraceutical components of the human and animal diet. PMID:27338492

  9. Glucose enhances indolic glucosinolate biosynthesis without reducing primary sulfur assimilation

    PubMed Central

    Miao, Huiying; Cai, Congxi; Wei, Jia; Huang, Jirong; Chang, Jiaqi; Qian, Hongmei; Zhang, Xin; Zhao, Yanting; Sun, Bo; Wang, Bingliang; Wang, Qiaomei

    2016-01-01

    The effect of glucose as a signaling molecule on induction of aliphatic glucosinolate biosynthesis was reported in our former study. Here, we further investigated the regulatory mechanism of indolic glucosinolate biosynthesis by glucose in Arabidopsis. Glucose exerted a positive influence on indolic glucosinolate biosynthesis, which was demonstrated by induced accumulation of indolic glucosinolates and enhanced expression of related genes upon glucose treatment. Genetic analysis revealed that MYB34 and MYB51 were crucial in maintaining the basal indolic glucosinolate accumulation, with MYB34 being pivotal in response to glucose signaling. The increased accumulation of indolic glucosinolates and mRNA levels of MYB34, MYB51, and MYB122 caused by glucose were inhibited in the gin2-1 mutant, suggesting an important role of HXK1 in glucose-mediated induction of indolic glucosinolate biosynthesis. In contrast to what was known on the function of ABI5 in glucose-mediated aliphatic glucosinolate biosynthesis, ABI5 was not required for glucose-induced indolic glucosinolate accumulation. In addition, our results also indicated that glucose-induced glucosinolate accumulation was due to enhanced sulfur assimilation instead of directed sulfur partitioning into glucosinolate biosynthesis. Thus, our data provide new insights into molecular mechanisms underlying glucose-regulated glucosinolate biosynthesis. PMID:27549907

  10. Glucose enhances indolic glucosinolate biosynthesis without reducing primary sulfur assimilation.

    PubMed

    Miao, Huiying; Cai, Congxi; Wei, Jia; Huang, Jirong; Chang, Jiaqi; Qian, Hongmei; Zhang, Xin; Zhao, Yanting; Sun, Bo; Wang, Bingliang; Wang, Qiaomei

    2016-01-01

    The effect of glucose as a signaling molecule on induction of aliphatic glucosinolate biosynthesis was reported in our former study. Here, we further investigated the regulatory mechanism of indolic glucosinolate biosynthesis by glucose in Arabidopsis. Glucose exerted a positive influence on indolic glucosinolate biosynthesis, which was demonstrated by induced accumulation of indolic glucosinolates and enhanced expression of related genes upon glucose treatment. Genetic analysis revealed that MYB34 and MYB51 were crucial in maintaining the basal indolic glucosinolate accumulation, with MYB34 being pivotal in response to glucose signaling. The increased accumulation of indolic glucosinolates and mRNA levels of MYB34, MYB51, and MYB122 caused by glucose were inhibited in the gin2-1 mutant, suggesting an important role of HXK1 in glucose-mediated induction of indolic glucosinolate biosynthesis. In contrast to what was known on the function of ABI5 in glucose-mediated aliphatic glucosinolate biosynthesis, ABI5 was not required for glucose-induced indolic glucosinolate accumulation. In addition, our results also indicated that glucose-induced glucosinolate accumulation was due to enhanced sulfur assimilation instead of directed sulfur partitioning into glucosinolate biosynthesis. Thus, our data provide new insights into molecular mechanisms underlying glucose-regulated glucosinolate biosynthesis. PMID:27549907

  11. Disorders of carnitine biosynthesis and transport.

    PubMed

    El-Hattab, Ayman W; Scaglia, Fernando

    2015-11-01

    Carnitine is a hydrophilic quaternary amine that plays a number of essential roles in metabolism with the main function being the transport of long-chain fatty acids from the cytosol to the mitochondrial matrix for β-oxidation. Carnitine can be endogenously synthesized. However, only a small fraction of carnitine is obtained endogenously while the majority is obtained from diet, mainly animal products. Carnitine is not metabolized and is excreted in urine. Carnitine homeostasis is regulated by efficient renal reabsorption that maintains carnitine levels within the normal range despite variabilities in dietary intake. Diseases occurring due to primary defects in carnitine metabolism and homeostasis are comprised in two groups: disorders of carnitine biosynthesis and carnitine transport defect. While the hallmark of carnitine transport defect is profound carnitine depletion, disorders of carnitine biosynthesis do not cause carnitine deficiency due to the fact that both carnitine obtained from diet and efficient renal carnitine reabsorption can maintain normal carnitine levels with the absence of endogenously synthesized carnitine. Carnitine transport defect phenotype encompasses a broad clinical spectrum including metabolic decompensation in infancy, cardiomyopathy in childhood, fatigability in adulthood, or absence of symptoms. The phenotypes associated with the carnitine transport defect result from the unavailability of enough carnitine to perform its functions particularly in fatty acid β-oxidation. Carnitine biosynthetic defects have been recently described and the phenotypic consequences of these defects are still emerging. Although these defects do not result in carnitine deficiency, they still could be associated with pathological phenotypes due to excess or deficiency of intermediate metabolites in the carnitine biosynthetic pathway and potential carnitine deficiency in early stages of life when brain and other organs develop. In addition to these two

  12. PLANT VOLATILES. Biosynthesis of monoterpene scent compounds in roses.

    PubMed

    Magnard, Jean-Louis; Roccia, Aymeric; Caissard, Jean-Claude; Vergne, Philippe; Sun, Pulu; Hecquet, Romain; Dubois, Annick; Hibrand-Saint Oyant, Laurence; Jullien, Frédéric; Nicolè, Florence; Raymond, Olivier; Huguet, Stéphanie; Baltenweck, Raymonde; Meyer, Sophie; Claudel, Patricia; Jeauffre, Julien; Rohmer, Michel; Foucher, Fabrice; Hugueney, Philippe; Bendahmane, Mohammed; Baudino, Sylvie

    2015-07-01

    The scent of roses (Rosa x hybrida) is composed of hundreds of volatile molecules. Monoterpenes represent up to 70% percent of the scent content in some cultivars, such as the Papa Meilland rose. Monoterpene biosynthesis in plants relies on plastid-localized terpene synthases. Combining transcriptomic and genetic approaches, we show that the Nudix hydrolase RhNUDX1, localized in the cytoplasm, is part of a pathway for the biosynthesis of free monoterpene alcohols that contribute to fragrance in roses. The RhNUDX1 protein shows geranyl diphosphate diphosphohydrolase activity in vitro and supports geraniol biosynthesis in planta. PMID:26138978

  13. The Yersinia pestis Siderophore, Yersiniabactin, and the ZnuABC system both contribute to Zinc acquisition and the development of lethal septicemic plague in mice

    PubMed Central

    Bobrov, Alexander G.; Kirillina, Olga; Fetherston, Jacqueline D.; Miller, M. Clarke; Burlison, Joseph A.; Perry, Robert D.

    2014-01-01

    Summary Bacterial pathogens must overcome host sequestration of zinc (Zn2+), an essential micronutrient, during the infectious disease process. While the mechanisms to acquire chelated Zn2+ by bacteria are largely undefined, many pathogens rely upon the ZnuABC family of ABC transporters. Here we show that in Yersinia pestis, irp2, a gene encoding the synthetase (HMWP2) for the siderophore yersiniabactin (Ybt) is required for growth under Zn2+-deficient conditions in a strain lacking ZnuABC. Moreover, growth stimulation with exogenous, purified apo-Ybt provides evidence that Ybt may serve as a zincophore for Zn2+ acquisition. Studies with the Zn2+-dependent transcriptional reporter znuA∷lacZ indicate that the ability to synthesize Ybt affects the levels of intracellular Zn2+. However, the outer membrane receptor Psn and TonB as well as the inner membrane (IM) ABC transporter YbtPQ, that are required for Fe3+ acquisition by Ybt, are not needed for Ybt-dependent Zn2+ uptake. In contrast, the predicted IM protein YbtX, a member of the Major Facilitator Superfamily, was essential for Ybt-dependent Zn2+ uptake. Finally, we show that the ZnuABC system and the Ybt synthetase HMWP2, presumably by Ybt synthesis, both contribute to the development of a lethal infection in a septicemic plague mouse model. PMID:24979062

  14. The role of iron in Mycobacterium smegmatis biofilm formation: the exochelin siderophore is essential in limiting iron conditions for biofilm formation but not for planktonic growth

    PubMed Central

    Ojha, Anil; Hatfull, Graham F

    2007-01-01

    Many species of mycobacteria form structured biofilm communities at liquid–air interfaces and on solid surfaces. Full development of Mycobacterium smegmatis biofilms requires addition of supplemental iron above 1 μM ferrous sulphate, although addition of iron is not needed for planktonic growth. Microarray analysis of the M. smegmatis transcriptome shows that iron-responsive genes – especially those involved in siderophore synthesis and iron uptake – are strongly induced during biofilm formation reflecting a response to iron deprivation, even when 2 μM iron is present. The acquisition of iron under these conditions is specifically dependent on the exochelin synthesis and uptake pathways, and the strong defect of an iron–exochelin uptake mutant suggests a regulatory role of iron in the transition to biofilm growth. In contrast, although the expression of mycobactin and iron ABC transport operons is highly upregulated during biofilm formation, mutants in these systems form normal biofilms in low-iron (2 μM) conditions. A close correlation between iron availability and matrix-associated fatty acids implies a possible metabolic role in the late stages of biofilm maturation, in addition to the early regulatory role. M. smegmatis surface motility is similarly dependent on iron availability, requiring both supplemental iron and the exochelin pathway to acquire it. PMID:17854402

  15. The Bacterial Stress-Responsive Hsp90 Chaperone (HtpG) Is Required for the Production of the Genotoxin Colibactin and the Siderophore Yersiniabactin in Escherichia coli.

    PubMed

    Garcie, Christophe; Tronnet, Sophie; Garénaux, Amélie; McCarthy, Alex J; Brachmann, Alexander O; Pénary, Marie; Houle, Sébastien; Nougayrède, Jean-Philippe; Piel, Jörn; Taylor, Peter W; Dozois, Charles M; Genevaux, Pierre; Oswald, Eric; Martin, Patricia

    2016-09-15

    The genotoxin colibactin, synthesized by Escherichia coli, is a secondary metabolite belonging to the chemical family of hybrid polyketide/nonribosomal peptide compounds. It is produced by a complex biosynthetic assembly line encoded by the pks pathogenicity island. The presence of this large cluster of genes in the E. coli genome is invariably associated with the high-pathogenicity island, encoding the siderophore yersiniabactin, which belongs to the same chemical family as colibactin. The E. coli heat shock protein HtpG (Hsp90Ec) is the bacterial homolog of the eukaryotic molecular chaperone Hsp90, which is involved in the protection of cellular proteins against a variety of environmental stresses. In contrast to eukaryotic Hsp90, the functions and client proteins of Hsp90Ec are poorly known. Here, we demonstrated that production of colibactin and yersiniabactin is abolished in the absence of Hsp90Ec We further characterized an interplay between the Hsp90Ec molecular chaperone and the ClpQ protease involved in colibactin and yersiniabactin synthesis. Finally, we demonstrated that Hsp90Ec is required for the full in vivo virulence of extraintestinal pathogenic E. coli This is the first report highlighting the role of heat shock protein Hps90Ec in the production of two secondary metabolites involved in E. coli virulence. PMID:27412582

  16. Regulation of volatile benzenoid biosynthesis in petunia flowers.

    PubMed

    Schuurink, Robert C; Haring, Michel A; Clark, David G

    2006-01-01

    The petunia flower has served as a model for the study of several physiological processes including floral development, self-incompatibility, anthocyanin biosynthesis and ethylene signalling during senescence. More recently, Petunia hybrida 'Mitchell' has been used to understand the complex regulation of volatile benzenoid biosynthesis, which occurs predominantly in flower petal tissues. Benzenoid biosynthesis is temporally and circadian controlled and is tightly down-regulated by ethylene during floral senescence. Using targeted transcriptomics and gene knockouts, both biosynthetic genes and a transcription factor regulating benzenoid synthesis have been recently discovered and characterized. It appears that benzenoid production is regulated predominantly by transcriptional control of the shikimate pathway, benzenoid biosynthesis genes and S-adenosyl-methionine cycle genes. PMID:16226052

  17. Pyrethrin biosynthesis and its regulation in Chrysanthemum cinerariaefolium.

    PubMed

    Matsuda, Kazuhiko

    2012-01-01

    Pyrethrins are a natural insecticide biosynthesized by the plant pyrethrum [Chrysanthemum cinerariaefolium (Current species name: Tanacetum cinerariifolium)] of the family Asteraceae. Although pyrethrins have been used to control household pests for the past century, little is known about the mechanism of biosynthesis, contrasting with intensive research on their synthetic analogs, pyrethroids. The author studied pyrethrin biosynthesis in young seedlings of C. cinerariaefolium. The results of experiments using (13)C-labeled glucose as the biosynthesis precursor indicated that the acid and alcohol moieties are biosynthesized via the 2-C-methyl-D: -erythritol 4-phosphate (MEP) and oxylipin pathways, respectively. Further study on the effects of wound-induced signals in leaves showed that biosynthesis is enhanced in response to both volatile and nonvolatile signals. PMID:22006239

  18. Biosynthesis of putrescine in the prostate gland of the rat

    PubMed Central

    Pegg, A. E.; Williams-Ashman, H. G.

    1968-01-01

    In the rat ventral prostate gland the biosynthesis of putrescine, a precursor of spermidine and spermine, is shown to occur by the direct decarboxylation of l-ornithine. Some properties of a soluble pyridoxal phosphate-dependent l-ornithine decarboxylase are described. The findings are discussed in relation to other enzymic reactions involved in the biosynthesis of polyamines by the prostate gland. PMID:5667265

  19. A Functional Genomics Approach to Tanshinone Biosynthesis Provides Stereochemical Insights

    PubMed Central

    2009-01-01

    Tanshinones are abietane-type norditerpenoid quinone natural products that are the bioactive components of the Chinese medicinal herb Salvia miltiorrhiza Bunge. The initial results from a functional genomics-based investigation of tanshinone biosynthesis, specifically the functional identification of the relevant diterpene synthases from S. miltiorrhiza, are reported. The cyclohexa-1,4-diene arrangement of the distal ring poises the resulting miltiradiene for the ensuing aromatization and hydroxylation to ferruginol suggested for tanshinone biosynthesis. PMID:19905026

  20. A functional genomics approach to tanshinone biosynthesis provides stereochemical insights.

    PubMed

    Gao, Wei; Hillwig, Matthew L; Huang, Luqi; Cui, Guanghong; Wang, Xueyong; Kong, Jianqiang; Yang, Bin; Peters, Reuben J

    2009-11-19

    Tanshinones are abietane-type norditerpenoid quinone natural products that are the bioactive components of the Chinese medicinal herb Salvia miltiorrhiza Bunge. The initial results from a functional genomics-based investigation of tanshinone biosynthesis, specifically the functional identification of the relevant diterpene synthases from S. miltiorrhiza, are reported. The cyclohexa-1,4-diene arrangement of the distal ring poises the resulting miltiradiene for the ensuing aromatization and hydroxylation to ferruginol suggested for tanshinone biosynthesis. PMID:19905026

  1. Complexity Generation during Natural Product Biosynthesis using Redox Enzymes

    PubMed Central

    Wang, Peng; Gao, Xue; Tang, Yi

    2012-01-01

    Redox enzymes such as FAD-dependent and cytochrome P450 oxygenases play indispensible roles in generating structural complexity during natural product biosynthesis. In the pre-assembly steps, redox enzymes can convert garden variety primary metabolites into unique starter and extender building blocks. In the post-assembly tailoring steps, redox cascades can transform nascent scaffolds into structurally complex final products. In this review, we will discuss several recently characterized redox enzymes in the biosynthesis of polyketides and nonribosomal peptides. PMID:22564679

  2. Increased thromboxane biosynthesis in essential thrombocythemia.

    PubMed

    Rocca, B; Ciabattoni, G; Tartaglione, R; Cortelazzo, S; Barbui, T; Patrono, C; Landolfi, R

    1995-11-01

    In order to investigate the in vivo thromboxane (TX) biosynthesis in essential thrombocythemia (ET), we measured the urinary excretion of the major enzymatic metabolites of TXB2, 11-dehydro-TXB2 and 2,3-dinor-TXB2 in 40 ET patients as well as in 26 gender- and age-matched controls. Urinary 11-dehydro-TXB2 was significantly higher (p < 0.001) in thrombocythemic patients (4,063 +/- 3,408 pg/mg creatinine; mean +/- SD) than in controls (504 +/- 267 pg/mg creatinine), with 34 patients (85%) having 11-dehydro-TXB2 > 2 SD above the control mean. Patients with platelet number < 1,000 x 10 (9)/1 (n = 25) had significantly higher (p < 0.05) 11-dehydro-TXB2 excretion than patients with higher platelet count (4,765 +/- 3,870 pg/mg creatinine, n = 25, versus 2,279 +/- 1,874 pg/mg creatinine, n = 15). Average excretion values of patients aging > 55 was significantly higher than in the younger group (4,784 +/- 3,948 pg/mg creatinine, n = 24, versus 2,405 +/- 1,885 pg/mg creatinine, n = 16, p < 0.05). Low-dose aspirin (50 mg/d for 7 days) largely suppressed 11-dehydro-TXB2 excretion in 7 thrombocythemic patients, thus suggesting that platelets were the main source of enhanced TXA2 biosynthesis. The platelet count-corrected 11-dehydro-TXB2 excretion was positively correlated with age (r = 0.325, n = 40, p < 0.05) and inversely correlated with platelet count (r = -0.381, n = 40, p < 0.05). In addition 11 out of 13 (85%) patients having increased count-corrected 11-dehydro-TXB2 excretion, belonged to the subgroup with age > 55 and platelet count < 1,000 x 10(9)/1. We conclude that in essential thrombocythemia: 1) enhanced 11-dehydro-TXB2 excretion largely reflects platelet activation in vivo; 2) age as well as platelet count appear to influence the determinants of platelet activation in this setting, and can help in assessing the thrombotic risk and therapeutic strategy in individual patients. PMID:8607099

  3. Fenarimol, a Pyrimidine-Type Fungicide, Inhibits Brassinosteroid Biosynthesis

    PubMed Central

    Oh, Keimei; Matsumoto, Tadashi; Yamagami, Ayumi; Hoshi, Tomoki; Nakano, Takeshi; Yoshizawa, Yuko

    2015-01-01

    The plant steroid hormone brassinosteroids (BRs) are important signal mediators that regulate broad aspects of plant growth and development. With the discovery of brassinoazole (Brz), the first specific inhibitor of BR biosynthesis, several triazole-type BR biosynthesis inhibitors have been developed. In this article, we report that fenarimol (FM), a pyrimidine-type fungicide, exhibits potent inhibitory activity against BR biosynthesis. FM induces dwarfism and the open cotyledon phenotype of Arabidopsis seedlings in the dark. The IC50 value for FM to inhibit stem elongation of Arabidopsis seedlings grown in the dark was approximately 1.8 ± 0.2 μM. FM-induced dwarfism of Arabidopsis seedlings could be restored by brassinolide (BL) but not by gibberellin (GA). Assessment of the target site of FM in BR biosynthesis by feeding BR biosynthesis intermediates indicated that FM interferes with the side chain hydroxylation of BR biosynthesis from campestanol to teasterone. Determination of the binding affinity of FM to purified recombinant CYP90D1 indicated that FM induced a typical type II binding spectrum with a Kd value of approximately 0.79 μM. Quantitative real-time PCR analysis of the expression level of the BR responsive gene in Arabidopsis seedlings indicated that FM induces the BR deficiency in Arabidopsis. PMID:26230686

  4. Lipid Biosynthesis in Developing Mustard Seed

    PubMed Central

    Mukherjee, Kumar D.

    1983-01-01

    Cotyledons of developing mustard (Sinapis alba L.) seed have been found to synthesize lipids containing the common plant fatty acids and very long-chain monounsaturated (icosenoic, erucic, and tetracosenic) and saturated (icosanoic, docosanoic, and tetracosanoic) fatty acids from various radioactive precursors. The in vivo pattern of labeling of acyl lipids, either from fatty acids synthesized `endogenously' from radioactive acetate or malonate, or from radioactive fatty acids added `exogenously', indicates the involvement of the following pathways in the biosynthesis of triacylglycerols. Palmitic, stearic, and oleic acid, synthesized in the acyl carrier protein-track, are channeled to the Coenzyme A (CoA)-track and converted to triacylglycerols via the glycerol-3-phosphate pathway. Pools of stearoyl-CoA and oleoyl-CoA are elongated to very long-chain saturated and monounsaturated acyl-CoA, respectively. Most of the very long-chain saturated acyl-CoAs acylate preformed diacylglycerols. Very long-chain monounsaturated acyl-CoAs are converted to triacylglycerols, partly via phosphatidic acids and diacylglycerols, and partly by acylation of preformed diacylglycerols. PMID:16663345

  5. Cholesterol biosynthesis modulation regulates dengue viral replication.

    PubMed

    Rothwell, Christopher; Lebreton, Aude; Young Ng, Chuan; Lim, Joanne Y H; Liu, Wei; Vasudevan, Subhash; Labow, Mark; Gu, Feng; Gaither, L Alex

    2009-06-20

    We performed a focused siRNA screen in an A549 dengue type 2 New Guinea C subgenomic replicon cell line (Rluc-replicon) that contains a Renilla luciferase cassette. We found that siRNA mediated knock down of mevalonate diphospho decarboxylase (MVD) inhibited viral replication of the Rluc-replicon and DEN-2 NGC live virus replication in A549 cells. When the Rluc-replicon A459 cells were grown in delipidated media the replicon expression was suppressed and MVD knock down could further sensitize Renilla expression. Hymeglusin and zaragozic acid A could inhibit DEN-2 NGC live virus replication in K562 cells, while lovastatin could inhibit DEN-2 NGC live virus replication in human peripheral blood mononuclear cells. Renilla expression could be rescued in fluvastatin treated A549 Rluc-replicon cells after the addition of mevalonate, and partially restored with geranylgeranyl pyrophosphate, or farnesyl pyrophosphate. Our data suggest genetic and pharmacological modulation of cholesterol biosynthesis can regulate dengue virus replication. PMID:19419745

  6. A Biotin Biosynthesis Gene Restricted to Helicobacter

    PubMed Central

    Bi, Hongkai; Zhu, Lei; Jia, Jia; Cronan, John E.

    2016-01-01

    In most bacteria the last step in synthesis of the pimelate moiety of biotin is cleavage of the ester bond of pimeloyl-acyl carrier protein (ACP) methyl ester. The paradigm cleavage enzyme is Escherichia coli BioH which together with the BioC methyltransferase allows synthesis of the pimelate moiety by a modified fatty acid biosynthetic pathway. Analyses of the extant bacterial genomes showed that bioH is absent from many bioC-containing bacteria and is replaced by other genes. Helicobacter pylori lacks a gene encoding a homologue of the known pimeloyl-ACP methyl ester cleavage enzymes suggesting that it encodes a novel enzyme that cleaves this intermediate. We isolated the H. pylori gene encoding this enzyme, bioV, by complementation of an E. coli bioH deletion strain. Purified BioV cleaved the physiological substrate, pimeloyl-ACP methyl ester to pimeloyl-ACP by use of a catalytic triad, each member of which was essential for activity. The role of BioV in biotin biosynthesis was demonstrated using a reconstituted in vitro desthiobiotin synthesis system. BioV homologues seem the sole pimeloyl-ACP methyl ester esterase present in the Helicobacter species and their occurrence only in H. pylori and close relatives provide a target for development of drugs to specifically treat Helicobacter infections. PMID:26868423

  7. Explorations into the biosynthesis of bioscorine

    SciTech Connect

    Michelson, R.H.

    1988-01-01

    The biosynthesis of dioscorine in Dioscorea hispida has been studied by the feeding of putative precursors labelled at specific positions with {sup 2}H, {sup 3}H, and {sup 14}C. Administration of (3-{sup 14}C)3-hydroxy-3-methylglutaric acid to D. hispida by the wick method afforded dioscorine labelled preferentially at the C{sub 10} position implying that the biosynthetic pathway to the acetate-derived half of the dioscorine skeleton is going through this compound. Administration of ethyl (6-{sup 14}C)orsellinate to D. hispida by the wick method failed to give an appreciable incorporation into dioscroine thereby disproving an alternative mechanism describing the formation of the acetate-derived half of the dioscorine skeleton. Two attempts to simulate the alternative mechanism by oxidatively cleaving ethyl orsellinate also failed, further disfavoring this mechanism. Administration of (2,3){sup 13}C{sub 2}, {sup 14}C{sub 2}succinic acid, (3-{sup 14}C)aspartic acid and (7a-{sup 14}C)tryptophan by the leaf painting method gave very low incorporations into dioscorine making determination of the source of the nicotinic acid half of the dioscorine skeleton inconclusive. Administration of (6-{sup 2}H, {sup 3}H)nicotinic acid to D. hispida by the wick method afforded dioscorine exhibiting complete retention of {sup 3}H thereby disfavoring a mechanism involving a 3,6-dihydropyridine intermediate in the formation of the dioscorine skeleton.

  8. Mitochondrial Fusion Is Essential for Steroid Biosynthesis

    PubMed Central

    Cooke, Mariana; Soria, Gastón; Cornejo Maciel, Fabiana; Gottifredi, Vanesa; Podestá, Ernesto J.

    2012-01-01

    Although the contribution of mitochondrial dynamics (a balance in fusion/fission events and changes in mitochondria subcellular distribution) to key biological process has been reported, the contribution of changes in mitochondrial fusion to achieve efficient steroid production has never been explored. The mitochondria are central during steroid synthesis and different enzymes are localized between the mitochondria and the endoplasmic reticulum to produce the final steroid hormone, thus suggesting that mitochondrial fusion might be relevant for this process. In the present study, we showed that the hormonal stimulation triggers mitochondrial fusion into tubular-shaped structures and we demonstrated that mitochondrial fusion does not only correlate-with but also is an essential step of steroid production, being both events depend on PKA activity. We also demonstrated that the hormone-stimulated relocalization of ERK1/2 in the mitochondrion, a critical step during steroidogenesis, depends on mitochondrial fusion. Additionally, we showed that the SHP2 phosphatase, which is required for full steroidogenesis, simultaneously modulates mitochondrial fusion and ERK1/2 localization in the mitochondrion. Strikingly, we found that mitofusin 2 (Mfn2) expression, a central protein for mitochondrial fusion, is upregulated immediately after hormone stimulation. Moreover, Mfn2 knockdown is sufficient to impair steroid biosynthesis. Together, our findings unveil an essential role for mitochondrial fusion during steroidogenesis. These discoveries highlight the importance of organelles’ reorganization in specialized cells, prompting the exploration of the impact that organelle dynamics has on biological processes that include, but are not limited to, steroid synthesis. PMID:23029265

  9. Biosynthesis of plasmenylcholine in guinea pig heart

    SciTech Connect

    Wientzek, M.; Choy, P.C.

    1986-05-01

    In some mammalian hearts, up to 40% of the choline phosphoglyceride (CPG) exists as plasmenylcholine (1-alkenyl-2-acyl-glycero-3-phosphocholine). Although the majority of diacylphosphatidylcholine (PC) in mammalian hearts is synthesized from choline via the CDP-choline pathway, the formation of plasmenylcholine from choline was not known. In this study, they investigated the biosynthesis of plasmenyl-choline in the isolated guinea pig heart by perfusion with (/sup 3/H)choline. Labelled choline containing metabolites and labelled plasmenylcholine were isolated and determined at different perfusion time points. Significant amounts of labelling were found only in choline, phosphocholine, CDP-choline, plasmenyl-choline and PC. In addition, a precursor-product relationship was observed between the labelling of CDP-choline and plasmenylcholine. Such a relationship was not observed between choline and plasmenylcholine. Hence, they postulate that the incorporation of choline into plasmenylcholine is via the CDP-choline pathway and not via base exchange. The ability to condense 1-alkenyl-2-acyl-glycerol with CDP-choline was also demonstrated in vitro with guinea pig heart microsomes.

  10. Synthetic Biological Approaches to Natural Product Biosynthesis

    PubMed Central

    Winter, Jaclyn M; Tang, Yi

    2012-01-01

    Small molecules produced in Nature continue to be an inspiration for the development of new therapeutic agents. These natural products possess exquisite chemical diversity, which gives rise to their wide range of biological activities. In their host organism, natural products are assembled and modified by dedicated biosynthetic pathways that Nature has meticulously developed. Often times, the complex structures or chemical modifications instated by these pathways are difficult to replicate using traditional synthetic methods. An alternative approach for creating or enhancing the structural variation of natural products is through combinatorial biosynthesis. By rationally reprogramming and manipulating the biosynthetic machinery responsible for their production, unnatural metabolites that were otherwise inaccessible can be obtained. Additionally, new chemical structures can be synthesized or derivatized by developing the enzymes that carry out these complicated chemical reactions into biocatalysts. In this review, we will discuss a variety of combinatorial biosynthetic strategies, their technical challenges, and highlight some recent (since 2007) examples of rationally designed unnatural metabolites, as well as platforms that have been established for the production and modification of clinically important pharmaceutical compounds. PMID:22221832

  11. Regulation of Phosphatidylcholine Biosynthesis in Saccharomyces cerevisiae

    PubMed Central

    Waechter, Charles J.; Lester, Robert L.

    1971-01-01

    Evidence is presented which indicates that the biosynthesis of phosphatidylcholine by the methylation pathway in growing cultures of Saccharomyces cerevisiae is repressed by the presence of choline in the growth medium. This result, obtained previously for glucose-grown cells, was also observed for lactate-grown cells, of which half of the phosphatidylcholine is mitochondrial. A respiration-deficient mutant of the parent wild-type strain has been studied, and its inability to form functional mitochondria cannot be due to an impaired methylation pathway, as it has been shown to incorporate 14C-CH3-methionine into all of the methylated glycerophosphatides. The incorporation rate is depressed by the inclusion of 1 mm choline in the growth medium, suggesting a regulatory effect similar to that demonstrated for the wild-type strain. The effects of choline on the glycerophospholipid composition of lactate and glucose-grown cells is presented. The repressive effects of the two related bases, mono- and dimethylethanolamine, were examined, and reduced levels of 14C-CH3-methionine incorporation were found for cells grown in the presence of these bases. The effect of choline on the methylation rates is reversible and glucosegrown cells regain the nonrepressed level of methylation activity in 60 to 80 min after removal of choline from the growth medium. Images PMID:5547992

  12. The biosynthesis of sterols in higher plants

    PubMed Central

    Goad, L. J.; Goodwin, T. W.

    1966-01-01

    1. [2-14C]Mevalonate was incorporated into squalene and the major phytosterols of pea and maize leaves; it was also incorporated into compounds belonging to the 4,4-dimethyl and 4α-methyl steroid groups and which may be possible phytosterol intermediates. 2. l-[Me-14C]Methionine was incorporated into the major sterols and also into the 4,4-dimethyl and 4α-methyl steroid groups. No radioactivity was detected in squalene. 3. Under anaerobic conditions incorporation of [2-14C]-mevalonate into the non-saponifiable lipid of pea leaves was drastically decreased but radioactive squalene was accumulated. 4. Cycloartenol, 24-methylenecycloartanol, 24-methylenelophenol, 24-ethylidenelophenol, fucosterol, β-sitosterol, stigmasterol and campesterol have been identified by gas–liquid chromatography in pea leaves. 5. The significance of these results in connexion with phytosterol biosynthesis and the introduction of the alkyl group at C-24 into phytosterols is discussed. ImagesFig. 1. PMID:5964970

  13. Regulation of mammalian nucleotide metabolism and biosynthesis

    PubMed Central

    Lane, Andrew N.; Fan, Teresa W.-M.

    2015-01-01

    Nucleotides are required for a wide variety of biological processes and are constantly synthesized de novo in all cells. When cells proliferate, increased nucleotide synthesis is necessary for DNA replication and for RNA production to support protein synthesis at different stages of the cell cycle, during which these events are regulated at multiple levels. Therefore the synthesis of the precursor nucleotides is also strongly regulated at multiple levels. Nucleotide synthesis is an energy intensive process that uses multiple metabolic pathways across different cell compartments and several sources of carbon and nitrogen. The processes are regulated at the transcription level by a set of master transcription factors but also at the enzyme level by allosteric regulation and feedback inhibition. Here we review the cellular demands of nucleotide biosynthesis, their metabolic pathways and mechanisms of regulation during the cell cycle. The use of stable isotope tracers for delineating the biosynthetic routes of the multiple intersecting pathways and how these are quantitatively controlled under different conditions is also highlighted. Moreover, the importance of nucleotide synthesis for cell viability is discussed and how this may lead to potential new approaches to drug development in diseases such as cancer. PMID:25628363

  14. Lipid Flippases for Bacterial Peptidoglycan Biosynthesis

    PubMed Central

    Ruiz, Natividad

    2015-01-01

    The biosynthesis of cellular polysaccharides and glycoconjugates often involves lipid-linked intermediates that need to be translocated across membranes. Essential pathways such as N-glycosylation in eukaryotes and biogenesis of the peptidoglycan (PG) cell wall in bacteria share a common strategy where nucleotide-sugars are used to build a membrane-bound oligosaccharide precursor that is linked to a phosphorylated isoprenoid lipid. Once made, these lipid-linked intermediates must be translocated across a membrane so that they can serve as substrates in a different cellular compartment. How translocation occurs is poorly understood, although it clearly requires a transporter or flippase. Identification of these transporters is notoriously difficult, and, in particular, the identity of the flippase of lipid II, an intermediate required for PG biogenesis, has been the subject of much debate. Here, I will review the body of work that has recently fueled this controversy, centered on proposed flippase candidates FtsW, MurJ, and AmJ. PMID:26792999

  15. Retinoic acid: its biosynthesis and metabolism.

    PubMed

    Napoli, J L

    1999-01-01

    This article presents a model that integrates the functions of retinoid-binding proteins with retinoid metabolism. One of these proteins, the widely expressed (throughout retinoid target tissues and in all vertebrates) and highly conserved cellular retinol-binding protein (CRBP), sequesters retinol in an internal binding pocket that segregates it from the intracellular milieu. The CRBP-retinol complex appears to be the quantitatively major form of retinol in vivo, and may protect the promiscuous substrate from nonenzymatic degradation and/or non-specific enzymes. For example, at least seven types of dehydrogenases catalyze retinal synthesis from unbound retinol in vitro (NAD+ vs. NADP+ dependent, cytosolic vs. microsomal, short-chain dehydrogenases/reductases vs. medium-chain alcohol dehydrogenases). But only a fraction of these (some of the short-chain de-hydrogenases/reductases) have the fascinating additional ability of catalyzing retinal synthesis from CRBP-bound retinol as well. Similarly, CRBP and/or other retinoid-binding proteins function in the synthesis of retinal esters, the reduction of retinal generated from intestinal beta-carotene metabolism, and retinoic acid metabolism. The discussion details the evidence supporting an integrated model of retinoid-binding protein/metabolism. Also addressed are retinoid-androgen interactions and evidence incompatible with ethanol causing fetal alcohol syndrome by competing directly with retinol dehydrogenation to impair retinoic acid biosynthesis. PMID:10506831

  16. Biosynthesis of cadmium sulphide quantum semiconductor crystallites

    NASA Astrophysics Data System (ADS)

    Dameron, C. T.; Reese, R. N.; Mehra, R. K.; Kortan, A. R.; Carroll, P. J.; Steigerwald, M. L.; Brus, L. E.; Winge, D. R.

    1989-04-01

    NANOMETRE-SCALE semiconductor quantum crystallites exhibit size-dependent and discrete excited electronic states which occur at energies higher than the band gap of the corresponding bulk solid1-4. These crystallites are too small to have continuous energy bands, even though a bulk crystal structure is present. The onset of such quantum properties sets a fundamental limit to device miniaturization in microelectronics5. Structures with either one, two or all three dimensions on the nanometer scale are of particular interest in solid state physics6. We report here our discovery of the biosynthesis of quantum crystallites in yeasts Candida glabrata and Schizosaccharomyces pombe, cultured in the presence of cad-mium salts. Short chelating peptides of general structure (γ-Glu-Cys)n-Gly control the nucleation and growth of CdS crystallites to peptide-capped intracellular particles of diameter 20 Å. These quantum CdS crystallites are more monodisperse than CdS par-ticles synthesized chemically. X-ray data indicate that, at this small size, the CdS structure differs from that of bulk CdS and tends towards a six-coordinate rock-salt structure.

  17. Control of triacylglycerol biosynthesis in plants

    SciTech Connect

    Not Available

    1993-01-31

    Seeds of most species of the Umbelliferae (Apiaciae), Araliaceae, and Garryaceae families are characterized by their high content of the unusual C[sub 18] monounsaturated fatty acid petroselinic acid (18:l[Delta][sup 6cis]). Prior to a recent report of this lab, little was known of the biosynthetic origin of the cis[Delta][sup 6] double bond of petroselinic acid. Such knowledge may be of both biochemical and biotechnological significance. Because petroselinic acid is potentially the product of a novel desaturase, information regarding its synthesis may contribute to an understanding of fatty acid desaturation mechanisms in plants. Through chemical cleavage at its double bond, petroselinic acid can be used as a precursor of lauric acid (12:0), a component of detergents and surfactants, and adipic acid (6:0 dicarboxylic), the monomeric component of nylon 6,6. Therefore, the development of an agronomic source of an oil rich in petroselinic acid is of biotechnological interest. As such, studies of petroselinic acid biosynthesis may provide basic information required for any attempt to genetically engineer the production and accumulation of this fatty acid in an existing oilseed.

  18. Biosynthesis of the manumycin group antibiotics

    SciTech Connect

    Thiericke, R.; Zeeck, A. ); Nakagawa, Akira; Omura, Satoshi ); Herrold, R.E.; Wu, S.T.S. ); Beale, J.M.; Floss, H.G. )

    1990-05-09

    The biosynthesis of the manumycin group antibiotics manumycin (1) and asukamycin (2) was studied in Streptomyces parvulus Tue 64 and Streptomyces nodosus ssp. asukaensis ATCC 29,757 by using radioactive and stable isotope tracer techniques and high-field NMR spectroscopy. The results have demonstrated that the central, multifunctional mC{sub 7}N unit typical of this group of antibiotics, which serves as the starter unit for a short polyketide chain, is biosynthesized from a C{sub 4} Krebs cycle and a C{sub 3} triose phosphate pool intermediate by a new pathway, distinct from the shikimate, polyketide, or pentose phosphate routes leading to other mC{sub 7}N units in nature. The C{sub 5} unit in both 1 and 2 arises by a novel intramolecular cyclization of 5-aminolevulinic acid, and a cyclohexane ring and the adjacent carbon in 2 arise from the seven carbon atoms of shikimic acid. The side chains of both antibiotics represent typical polyketide-derived moieties, differing with respect to their combinations of starter and elongation units.

  19. Development of a PCR protocol for the detection of Aeromonas salmonicida in fish by amplification of the fstA (ferric siderophore receptor) gene.

    PubMed

    Beaz-Hidalgo, Roxana; Magi, Gian Enrico; Balboa, Sabela; Barja, Juan L; Romalde, Jesús L

    2008-04-30

    The aims of the study were to evaluate a new PCR protocol designed to detect Aeromonas salmonicida in fish tissues and to develop a non-destructive method for the diagnosis of furunculosis. A set of primers (Fer3, Fer4), flanking a fragment of the fstA gene (coding for the ferric-siderophore receptor) was designed, showing to be sensitive and specific. When compared to PCR methods previously reported, the new protocol recognized all the 69 A. salmonicida strains evaluated, with no cross-reactions with the other bacterial species analysed. Sensitivity assays were performed in fish tissues seeded with serial dilutions of pure cultures of A. salmonicida and mixed cultures of this bacterium with Vibrio anguillarum and Aeromonas hydrophila. Detection limits obtained were of 60 and 450 bacterial cells 100 mg(-1) of tissue, respectively. Mucus and blood were evaluated in order to develop a non-destructive tool to detect the pathogen. The detection limits in seeded mucus and blood samples were 2.5 x 10(2) and 1 x 10(5) bacterial cells mL(-1), respectively. When the method was used to detect A. salmonicida in asymptomatic wild salmon, four samples of mucus and six of blood were positive, corresponding to 6 out of the 31 fish examined, whereas only one of the samples resulted positive by culture methods. It is concluded that the PCR protocol evaluated is fast, specific and sensitive to detect A. salmonicida in infected and asymptomatic fish, and will be helpful for the control of the disease through the prompt detection of carriers within fish populations. PMID:18035507

  20. Soybean oil biosynthesis: role of diacylglycerol acyltransferases.

    PubMed

    Li, Runzhi; Hatanaka, Tomoko; Yu, Keshun; Wu, Yongmei; Fukushige, Hirotada; Hildebrand, David

    2013-03-01

    Diacylglycerol acyltransferase (DGAT) catalyzes the acyl-CoA-dependent acylation of sn-1,2-diacylglycerol to form seed oil triacylglycerol (TAG). To understand the features of genes encoding soybean (Glycine max) DGATs and possible roles in soybean seed oil synthesis and accumulation, two full-length cDNAs encoding type 1 diacylglycerol acyltransferases (GmDGAT1A and GmDGAT1B) were cloned from developing soybean seeds. These coding sequences share identities of 94 % and 95 % in protein and DNA sequences. The genomic architectures of GmDGAT1A and GmDGAT1B both contain 15 introns and 16 exons. Differences in the lengths of the first exon and most of the introns were found between GmDGAT1A and GmDGAT1B genomic sequences. Furthermore, detailed in silico analysis revealed a third predicted DGAT1, GmDGAT1C. GmDGAT1A and GmDGAT1B were found to have similar activity levels and substrate specificities. Oleoyl-CoA and sn-1,2-diacylglycerol were preferred substrates over vernoloyl-CoA and sn-1,2-divernoloylglycerol. Both transcripts are much more abundant in developing seeds than in other tissues including leaves, stem, roots, and flowers. Both soybean DGAT1A and DGAT1B are highly expressed at developing seed stages of maximal TAG accumulation with DGAT1B showing highest expression at somewhat later stages than DGAT1A. DGAT1A and DGAT1B show expression profiles consistent with important roles in soybean seed oil biosynthesis and accumulation. PMID:23322364

  1. Biosynthesis of the Aromatic Amino Acids.

    PubMed

    Pittard, James; Yang, Ji

    2008-09-01

    This chapter describes in detail the genes and proteins of Escherichia coli involved in the biosynthesis and transport of the three aromatic amino acids tyrosine, phenylalanine, and tryptophan. It provides a historical perspective on the elaboration of the various reactions of the common pathway converting erythrose-4-phosphate and phosphoenolpyruvate to chorismate and those of the three terminal pathways converting chorismate to phenylalanine, tyrosine, and tryptophan. The regulation of key reactions by feedback inhibition, attenuation, repression, and activation are also discussed. Two regulatory proteins, TrpR (108 amino acids) and TyrR (513 amino acids), play a major role in transcriptional regulation. The TrpR protein functions only as a dimer which, in the presence of tryptophan, represses the expression of trp operon plus four other genes (the TrpR regulon). The TyrR protein, which can function both as a dimer and as a hexamer, regulates the expression of nine genes constituting the TyrR regulon. TyrR can bind each of the three aromatic amino acids and ATP and under their influence can act as a repressor or activator of gene expression. The various domains of this protein involved in binding the aromatic amino acids and ATP, recognizing DNA binding sites, interacting with the alpha subunit of RNA polymerase, and changing from a monomer to a dimer or a hexamer are all described. There is also an analysis of the various strategies which allow TyrR in conjunction with particular amino acids to differentially affect the expression of individual genes of the TyrR regulon. PMID:26443741

  2. Dithiolopyrrolone Natural Products: Isolation, Synthesis and Biosynthesis

    PubMed Central

    Qin, Zhiwei; Huang, Sheng; Yu, Yi; Deng, Hai

    2013-01-01

    Dithiolopyrrolones are a class of antibiotics that possess the unique pyrrolinonodithiole (4H-[1,2] dithiolo [4,3-b] pyrrol-5-one) skeleton linked to two variable acyl groups. To date, there are approximately 30 naturally occurring dithiolopyrrolone compounds, including holomycin, thiolutin, and aureothricin, and more recently thiomarinols, a unique class of hybrid marine bacterial natural products containing a dithiolopyrrolone framework linked by an amide bridge with an 8-hydroxyoctanoyl chain linked to a monic acid. Generally, dithiolopyrrolone antibiotics have broad-spectrum antibacterial activity against various microorganisms, including Gram-positive and Gram-negative bacteria, and even parasites. Holomycin appeared to be active against rifamycin-resistant bacteria and also inhibit the growth of the clinical pathogen methicillin-resistant Staphylococcus aureus N315. Its mode of action is believed to inhibit RNA synthesis although the exact mechanism has yet to be established in vitro. A recent work demonstrated that the fish pathogen Yersinia ruckeri employs an RNA methyltransferase for self-resistance during the holomycin production. Moreover, some dithiolopyrrolone derivatives have demonstrated promising antitumor activities. The biosynthetic gene clusters of holomycin have recently been identified in S. clavuligerus and characterized biochemically and genetically. The biosynthetic gene cluster of thiomarinol was also identified from the marine bacterium Pseudoalteromonas sp. SANK 73390, which was uniquely encoded by two independent pathways for pseudomonic acid and pyrrothine in a novel plasmid. The aim of this review is to give an overview about the isolations, characterizations, synthesis, biosynthesis, bioactivities and mode of action of this unique family of dithiolopyrrolone natural products, focusing on the period from 1940s until now. PMID:24141227

  3. Coenzyme Q biosynthesis in health and disease.

    PubMed

    Acosta, Manuel Jesús; Vazquez Fonseca, Luis; Desbats, Maria Andrea; Cerqua, Cristina; Zordan, Roberta; Trevisson, Eva; Salviati, Leonardo

    2016-08-01

    Coenzyme Q (CoQ, or ubiquinone) is a remarkable lipid that plays an essential role in mitochondria as an electron shuttle between complexes I and II of the respiratory chain, and complex III. It is also a cofactor of other dehydrogenases, a modulator of the permeability transition pore and an essential antioxidant. CoQ is synthesized in mitochondria by a set of at least 12 proteins that form a multiprotein complex. The exact composition of this complex is still unclear. Most of the genes involved in CoQ biosynthesis (COQ genes) have been studied in yeast and have mammalian orthologues. Some of them encode enzymes involved in the modification of the quinone ring of CoQ, but for others the precise function is unknown. Two genes appear to have a regulatory role: COQ8 (and its human counterparts ADCK3 and ADCK4) encodes a putative kinase, while PTC7 encodes a phosphatase required for the activation of Coq7. Mutations in human COQ genes cause primary CoQ(10) deficiency, a clinically heterogeneous mitochondrial disorder with onset from birth to the seventh decade, and with clinical manifestation ranging from fatal multisystem disorders, to isolated encephalopathy or nephropathy. The pathogenesis of CoQ(10) deficiency involves deficient ATP production and excessive ROS formation, but possibly other aspects of CoQ(10) function are implicated. CoQ(10) deficiency is unique among mitochondrial disorders since an effective treatment is available. Many patients respond to oral CoQ(10) supplementation. Nevertheless, treatment is still problematic because of the low bioavailability of the compound, and novel pharmacological approaches are currently being investigated. This article is part of a Special Issue entitled 'EBEC 2016: 19th European Bioenergetics Conference, Riva del Garda, Italy, July 2-6, 2016', edited by Prof. Paolo Bernardi. PMID:27060254

  4. Recent advances in combinatorial biosynthesis for drug discovery

    PubMed Central

    Sun, Huihua; Liu, Zihe; Zhao, Huimin; Ang, Ee Lui

    2015-01-01

    Because of extraordinary structural diversity and broad biological activities, natural products have played a significant role in drug discovery. These therapeutically important secondary metabolites are assembled and modified by dedicated biosynthetic pathways in their host living organisms. Traditionally, chemists have attempted to synthesize natural product analogs that are important sources of new drugs. However, the extraordinary structural complexity of natural products sometimes makes it challenging for traditional chemical synthesis, which usually involves multiple steps, harsh conditions, toxic organic solvents, and byproduct wastes. In contrast, combinatorial biosynthesis exploits substrate promiscuity and employs engineered enzymes and pathways to produce novel “unnatural” natural products, substantially expanding the structural diversity of natural products with potential pharmaceutical value. Thus, combinatorial biosynthesis provides an environmentally friendly way to produce natural product analogs. Efficient expression of the combinatorial biosynthetic pathway in genetically tractable heterologous hosts can increase the titer of the compound, eventually resulting in less expensive drugs. In this review, we will discuss three major strategies for combinatorial biosynthesis: 1) precursor-directed biosynthesis; 2) enzyme-level modification, which includes swapping of the entire domains, modules and subunits, site-specific mutagenesis, and directed evolution; 3) pathway-level recombination. Recent examples of combinatorial biosynthesis employing these strategies will also be highlighted in this review. PMID:25709407

  5. Jasmonate-induced biosynthesis of andrographolide in Andrographis paniculata.

    PubMed

    Sharma, Shiv Narayan; Jha, Zenu; Sinha, Rakesh Kumar; Geda, Arvind Kumar

    2015-02-01

    Andrographolide is a prominent secondary metabolite found in Andrographis paniculata that exhibits enormous pharmacological effects. In spite of immense value, the normal biosynthesis of andrographolide results in low amount of the metabolite. To induce the biosynthesis of andrographolide, we attempted elicitor-induced activation of andrographolide biosynthesis in cell cultures of A. paniculata. This was carried out by using methyl jasmonate (MeJA) as an elicitor. Among the various concentrations of MeJA tested at different time periods, 5 µM MeJA yielded 5.25 times more andrographolide content after 24 h of treatment. The accumulation of andrographolide was correlated with the expression level of known regulatory genes (hmgs, hmgr, dxs, dxr, isph and ggps) of mevalonic acid (MVA) and 2-C-methyl-d-erythritol-4-phosphate (MEP) pathways. These results established the involvement of MeJA in andrographolide biosynthesis by inducing the transcription of its biosynthetic pathways genes. The coordination of isph, ggps and hmgs expression highly influenced the andrographolide biosynthesis. PMID:25104168

  6. Roles of lignin biosynthesis and regulatory genes in plant development.

    PubMed

    Yoon, Jinmi; Choi, Heebak; An, Gynheung

    2015-11-01

    Lignin is an important factor affecting agricultural traits, biofuel production, and the pulping industry. Most lignin biosynthesis genes and their regulatory genes are expressed mainly in the vascular bundles of stems and leaves, preferentially in tissues undergoing lignification. Other genes are poorly expressed during normal stages of development, but are strongly induced by abiotic or biotic stresses. Some are expressed in non-lignifying tissues such as the shoot apical meristem. Alterations in lignin levels affect plant development. Suppression of lignin biosynthesis genes causes abnormal phenotypes such as collapsed xylem, bending stems, and growth retardation. The loss of expression by genes that function early in the lignin biosynthesis pathway results in more severe developmental phenotypes when compared with plants that have mutations in later genes. Defective lignin deposition is also associated with phenotypes of seed shattering or brittle culm. MYB and NAC transcriptional factors function as switches, and some homeobox proteins negatively control lignin biosynthesis genes. Ectopic deposition caused by overexpression of lignin biosynthesis genes or master switch genes induces curly leaf formation and dwarfism. PMID:26297385

  7. Dissecting the role of glutathione biosynthesis in Plasmodium falciparum

    PubMed Central

    Patzewitz, Eva-Maria; Wong, Eleanor H; Müller, Sylke

    2012-01-01

    Glutathione (γ-glutamylcysteinyl-glycine, GSH) has vital functions as thiol redox buffer and cofactor of antioxidant and detoxification enzymes. Plasmodium falciparum possesses a functional GSH biosynthesis pathway and contains mM concentrations of the tripeptide. It was impossible to delete in P. falciparum the genes encoding γ-glutamylcysteine synthetase (γGCS) or glutathione synthetase (GS), the two enzymes synthesizing GSH, although both gene loci were not refractory to recombination. Our data show that the parasites cannot compensate for the loss of GSH biosynthesis via GSH uptake. This suggests an important if not essential function of GSH biosynthesis pathway for the parasites. Treatment with the irreversible inhibitor of γGCS L-buthionine sulfoximine (BSO) reduced intracellular GSH levels in P. falciparum and was lethal for their intra-erythrocytic development, corroborating the suggestion that GSH biosynthesis is important for parasite survival. Episomal expression of γgcs in P. falciparum increased tolerance to BSO attributable to increased levels of γGCS. Concomitantly expression of glutathione reductase was reduced leading to an increased GSH efflux. Together these data indicate that GSH levels are tightly regulated by a functional GSH biosynthesis and the reduction of GSSG. PMID:22151036

  8. Amino acid biosynthesis in the spirochete Leptospira: evidence for a novel pathway of isoleucine biosynthesis.

    PubMed

    Charon, N W; Johnson, R C; Peterson, D

    1974-01-01

    Radioactive carbon dioxide was incubated with growing cells of Leptospira interrogans serotypes semaranga and tarassovi, and the specific activities and distribution of the label within the cellular amino acids were determined. The origins of the carbon skeletons of all the acid-stable amino acids except isoleucine were found to be consistent with known biosynthetic pathways for these amino acids. Experiments using radioactive carbon dioxide and other tracers indicated that most of the isoleucine was synthesized by a pathway not involving threonine. The origin of the carbon skeleton of isoleucine consisted of two residues of pyruvate (carbons 2 and 3) and acetate of acetyl-coenzyme A by this pathway. Isotope competition studies indicated that the pathway was regulated by isoleucine. The results are discussed in relation to two proposed pathways of isoleucine biosynthesis involving citramalate as an intermediate. PMID:4808901

  9. Sesterterpene ophiobolin biosynthesis involving multiple gene clusters in Aspergillus ustus

    PubMed Central

    Chai, Hangzhen; Yin, Ru; Liu, Yongfeng; Meng, Huiying; Zhou, Xianqiang; Zhou, Guolin; Bi, Xupeng; Yang, Xue; Zhu, Tonghan; Zhu, Weiming; Deng, Zixin; Hong, Kui

    2016-01-01

    Terpenoids are the most diverse and abundant natural products among which sesterterpenes account for less than 2%, with very few reports on their biosynthesis. Ophiobolins are tricyclic 5–8–5 ring sesterterpenes with potential pharmaceutical application. Aspergillus ustus 094102 from mangrove rizhosphere produces ophiobolin and other terpenes. We obtained five gene cluster knockout mutants, with altered ophiobolin yield using genome sequencing and in silico analysis, combined with in vivo genetic manipulation. Involvement of the five gene clusters in ophiobolin synthesis was confirmed by investigation of the five key terpene synthesis relevant enzymes in each gene cluster, either by gene deletion and complementation or in vitro verification of protein function. The results demonstrate that ophiobolin skeleton biosynthesis involves five gene clusters, which are responsible for C15, C20, C25, and C30 terpenoid biosynthesis. PMID:27273151

  10. Sesterterpene ophiobolin biosynthesis involving multiple gene clusters in Aspergillus ustus.

    PubMed

    Chai, Hangzhen; Yin, Ru; Liu, Yongfeng; Meng, Huiying; Zhou, Xianqiang; Zhou, Guolin; Bi, Xupeng; Yang, Xue; Zhu, Tonghan; Zhu, Weiming; Deng, Zixin; Hong, Kui

    2016-01-01

    Terpenoids are the most diverse and abundant natural products among which sesterterpenes account for less than 2%, with very few reports on their biosynthesis. Ophiobolins are tricyclic 5-8-5 ring sesterterpenes with potential pharmaceutical application. Aspergillus ustus 094102 from mangrove rizhosphere produces ophiobolin and other terpenes. We obtained five gene cluster knockout mutants, with altered ophiobolin yield using genome sequencing and in silico analysis, combined with in vivo genetic manipulation. Involvement of the five gene clusters in ophiobolin synthesis was confirmed by investigation of the five key terpene synthesis relevant enzymes in each gene cluster, either by gene deletion and complementation or in vitro verification of protein function. The results demonstrate that ophiobolin skeleton biosynthesis involves five gene clusters, which are responsible for C15, C20, C25, and C30 terpenoid biosynthesis. PMID:27273151

  11. Improved Precursor Directed Biosynthesis in E. coli via Directed Evolution

    PubMed Central

    Lee, Ho Young; Harvey, Colin J.B.; Cane, David E.; Khosla, Chaitan

    2010-01-01

    Erythromycin and related macrolide antibiotics are widely used polyketide natural products. We have evolved an engineered biosynthetic pathway in Escherichia coli that yields erythromycin analogs from simple synthetic precursors. Multiple rounds of mutagenesis and screening led to the identification of new mutant strains with improved efficiency for precursor directed biosynthesis. Genetic and biochemical analysis suggested that the phenotypically relevant alterations in these mutant strains were localized exclusively to the host-vector system, and not to the polyketide synthase. We also demonstrate the utility of this improved system through engineered biosynthesis of a novel alkynyl erythromycin derivative with comparable antibacterial activity to its natural counterpart. In addition to reinforcing the power of directed evolution for engineering macrolide biosynthesis, our studies have identified a new lead substance for investigating structure-function relationships in the bacterial ribosome. PMID:21081955

  12. Inhibitors targeting on cell wall biosynthesis pathway of MRSA.

    PubMed

    Hao, Haihong; Cheng, Guyue; Dai, Menghong; Wu, Qinghua; Yuan, Zonghui

    2012-11-01

    Methicillin resistant Staphylococcus aureus (MRSA), widely known as a type of new superbug, has aroused world-wide concern. Cell wall biosynthesis pathway is an old but good target for the development of antibacterial agents. Peptidoglycan and wall teichoic acids (WTAs) biosynthesis are two main processes of the cell wall biosynthesis pathway (CWBP). Other than penicillin-binding proteins (PBPs), some key factors (Mur enzymes, lipid I or II precursor, etc.) in CWBP are becoming attractive molecule targets for the discovery of anti-MRSA compounds. A number of new compounds, with higher affinity for PBPs or with inhibitory activity on such molecule targets in CWBP of MRSA, have been in the pipeline recently. This review concludes recent research achievements and provides a complete picture of CWBP of MRSA, including the peptidoglycan and wall teichoic acids synthesis pathway. The potential inhibitors targeting on CWBP are subsequently presented to improve development of novel therapeutic strategies for MRSA. PMID:22898792

  13. Squalestatin Is an Inhibitor of Carotenoid Biosynthesis in Plasmodium falciparum

    PubMed Central

    Gabriel, Heloisa B.; Silva, Marcia F.; Kimura, Emília A.; Wunderlich, Gerhard

    2015-01-01

    The increasing resistance of malaria parasites to almost all available drugs calls for the characterization of novel targets and the identification of new compounds. Carotenoids are polyisoprenoids from plants, algae, and some bacteria, and they are biosynthesized by Plasmodium falciparum but not by mammalian cells. Biochemical and reverse genetics approaches were applied to demonstrate that phytoene synthase (PSY) is a key enzyme for carotenoid biosynthesis in P. falciparum and is essential for intraerythrocytic growth. The known PSY inhibitor squalestatin reduces biosynthesis of phytoene and kills parasites during the intraerythrocytic cycle. PSY-overexpressing parasites showed increased biosynthesis of phytoene and its derived product phytofluene and presented a squalestatin-resistant phenotype, suggesting that this enzyme is the primary target of action of this drug in the parasite. PMID:25779575

  14. Quinolizidine alkaloid biosynthesis: recent advances and future prospects

    PubMed Central

    Bunsupa, Somnuk; Yamazaki, Mami; Saito, Kazuki

    2012-01-01

    Lys-derived alkaloids, including piperidine, quinolizidine, indolizidine, and lycopodium alkaloids, are widely distributed throughout the plant kingdom. Several of these alkaloids have beneficial properties for humans and have been used in medicine. However, the molecular mechanisms underlying the biosynthesis of these alkaloids are not well understood. In the present article, we discuss recent advances in our understanding of Lys-derived alkaloids, especially the biochemistry, molecular biology, and biotechnology of quinolizidine alkaloid (QA) biosynthesis. We have also highlighted Lys decarboxylase (LDC), the enzyme that catalyzes the first committed step of QA biosynthesis and answers a longstanding question about the molecular entity of LDC activity in plants. Further prospects using current advanced technologies, such as next-generation sequencing, in medicinal plants have also been discussed. PMID:23112802

  15. Lovastatin Biosynthesis by Aspergillus terreus in a Chemically Defined Medium

    PubMed Central

    Hajjaj, Hassan; Niederberger, Peter; Duboc, Philippe

    2001-01-01

    Lovastatin is a secondary metabolite produced by Aspergillus terreus. A chemically defined medium was developed in order to investigate the influence of carbon and nitrogen sources on lovastatin biosynthesis. Among several organic and inorganic defined nitrogen sources metabolized by A. terreus, glutamate and histidine gave the highest lovastatin biosynthesis level. For cultures on glucose and glutamate, lovastatin synthesis initiated when glucose consumption levelled off. When A. terreus was grown on lactose, lovastatin production initiated in the presence of residual lactose. Experimental results showed that carbon source starvation is required in addition to relief of glucose repression, while glutamate did not repress biosynthesis. A threefold-higher specific productivity was found with the defined medium on glucose and glutamate, compared to growth on complex medium with glucose, peptonized milk, and yeast extract. PMID:11375168

  16. Identification of unique mechanisms for triterpene biosynthesis in Botryococcus braunii

    PubMed Central

    Niehaus, Tom D.; Okada, Shigeru; Devarenne, Timothy P.; Watt, David S.; Sviripa, Vitaliy; Chappell, Joe

    2011-01-01

    Botryococcene biosynthesis is thought to resemble that of squalene, a metabolite essential for sterol metabolism in all eukaryotes. Squalene arises from an initial condensation of two molecules of farnesyl diphosphate (FPP) to form presqualene diphosphate (PSPP), which then undergoes a reductive rearrangement to form squalene. In principle, botryococcene could arise from an alternative rearrangement of the presqualene intermediate. Because of these proposed similarities, we predicted that a botryococcene synthase would resemble squalene synthase and hence isolated squalene synthase-like genes from Botryococcus braunii race B. While B. braunii does harbor at least one typical squalene synthase, none of the other three squalene synthase-like (SSL) genes encodes for botryococcene biosynthesis directly. SSL-1 catalyzes the biosynthesis of PSPP and SSL-2 the biosynthesis of bisfarnesyl ether, while SSL-3 does not appear able to directly utilize FPP as a substrate. However, when combinations of the synthase-like enzymes were mixed together, in vivo and in vitro, robust botryococcene (SSL-1+SSL-3) or squalene biosynthesis (SSL1+SSL-2) was observed. These findings were unexpected because squalene synthase, an ancient and likely progenitor to the other Botryococcus triterpene synthases, catalyzes a two-step reaction within a single enzyme unit without intermediate release, yet in B. braunii, these activities appear to have separated and evolved interdependently for specialized triterpene oil production greater than 500 MYA. Coexpression of the SSL-1 and SSL-3 genes in different configurations, as independent genes, as gene fusions, or targeted to intracellular membranes, also demonstrate the potential for engineering even greater efficiencies of botryococcene biosynthesis. PMID:21746901

  17. Genetic regulations of the biosynthesis of microbial surfactants: an overview.

    PubMed

    Das, Palashpriya; Mukherjee, Soumen; Sen, Ramkrishna

    2008-01-01

    Microbial biosurfactants are surface active metabolites synthesized by microbes growing on a variety of substrates. In spite of having great potential for commercial, therapeutic and environmental applications, industrial level production has not been realized for their low yields and productivities. One vital factor determining their biosynthesis is the genetic makeup of the producer organisms. Studies on molecular genetics and biochemistry of the synthesis of several biosurfactants have revealed the operons, the enzymes and the metabolic pathways required for their extracellular production. Surfactin, a cyclic lipopeptide biosurfactant is a potent antimicrobial agent and is produced as a result of non-ribosomal biosynthesis catalyzed by a large multienzyme peptide synthetase complex called the surfactin synthetase. Pathways for the synthesis of other lipopeptides such as iturin, lichenysin and arthrofactin are also mediated by similar enzyme complexes. These non-ribosomal peptide synthetases (NRPSs) responsible for lipopeptide biosynthesis display a high degree of structural similarity among themselves even from distant microbial species. Plasmid-encoded- rhlA, B, R and I genes of rhl quorum sensing system are required for production of glycolipid biosurfactants by Pseudomonas species. Molecular genetics of biosynthesis of alasan and emulsan by Acinetobacter species and of the fungal biosurfactants such as mannosylerythritol lipids (MEL) and hydrophobins have been deciphered. However, limited genetic information is available about biosynthesis of other biosurfactants such as viscosin, amphisin and putisolvin produced by some strains of Pseudomonas species. Understanding of the genetic regulatory mechanisms would help to develop metabolically engineered hyper-producing strains with better product characteristics and acquired capability of utilizing cheap agro-industrial wastes as substrates. This article thus provides an overview of the role and importance of

  18. Physiological insights into all-trans-retinoic acid biosynthesis

    PubMed Central

    Napoli, Joseph L.

    2011-01-01

    All-trans-retinoic acid (atRA) provides essential support to diverse biological systems and physiological processes. Epithelial differentiation and its relationship to cancer and embryogenesis have typified intense areas of interest into atRA function. Recently, however, interest in atRA action in the nervous system, the immune system, energy balance and obesity has increased considerably, especially concerning postnatal function. atRA action depends on atRA biosynthesis: defects in retinoid-dependent processes increasingly relate to defects in atRA biogenesis. Considerable evidence indicates that physiological atRA biosynthesis occurs via a regulated process, consisting of a complex interaction of retinoid binding-proteins and retinoid recognizing enzymes. An accrual of biochemical, physiological and genetic data have identified specific functional outcomes for the retinol dehydrogenases, RDH1, RDH10, and DHRS9, as physiological catalysts of the first step in atRA biosynthesis, and for the retinal dehydrogenases RALDH1, RALDH2, and RALDH3, as catalysts of the second and irreversible step. Each of these enzymes associates with explicit biological processes mediated by atRA. Redundancy occurs, but seems limited. Cumulative data supports a model of interactions among these enzymes with retinoid binding-proteins, with feedback regulation and/or control by atRA via modulating gene expression of multiple participants. The ratio apo-CRBP1/holo-CRBP1 participates by influencing retinol flux into and out of storage as retinyl esters, thereby modulating substrate to support atRA biosynthesis. atRA biosynthesis requires presence of both an RDH and an RALDH: conversely, absence of one isozyme of either step does not indicate lack of atRA biosynthesis at the site. PMID:21621639

  19. Natural Products as Tools for Chemogenomic Analysis of Mycotoxin Biosynthesis and Fungal Stress-Response Systems

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Certain phenolics having antioxidative activity can inhibit aflatoxin biosynthesis by Aspergillus flavus, with no effect on fungal growth. Contrastingly, exposing A. flavus to oxidative stress, such as hydrogen peroxide, enhances aflatoxin biosynthesis. Use of gene-deletion mutants of Saccharomyces ...

  20. [Advances in the regulation of cephalosporin C biosynthesis - A review].

    PubMed

    Liu, Jiajia; Liu, Gang

    2016-03-01

    The beta-lactam antibiotic cephalosporin C is produced industrially by Acremonium chrysogenum. Its derivative 7-aminocephalosporanic acid (7-ACA) is the intermediate of most chemical modification cephalosporins that are the most frequently used antibiotics for the therapy of infectious diseases. Due to its importance, the biosynthetic pathway of cephalosporin C has been elucidated in Acremonium chrysogenum. To improve the yield of cephalosporin C and reduce the cost of production, recent studies have been focused on the sophisticated regulation of cephalosporin C biosynthesis. In this review, recent advances in cephalosporin C biosynthesis and regulation are summarized. PMID:27382789

  1. Final Report on Regulation of Guaiacyl and Syringyl Monolignol Biosynthesis

    SciTech Connect

    Vincent L. Chiang

    2006-03-09

    The focus of this research is to understand syringyl monolignol biosynthesis that leads to the formation of syringyl lignin, a type of lignin that can be easily removed during biomass conversion. We have achieved the three originally proposed goals for this project. (1) SAD and CAD genes (enzyme catalytic and kinetic properties) and their functional relevance to CAld5H/AldOMT pathway, (2) spatiotemporal expression patterns of Cald5H, AldOMT, SAD and CAD genes, and (3) functions of CAld5H, AldOMT, and SAD genes in vivo using transgenic aspen. Furthermore, we also found that microRNA might be involved in the upstream regulatory network of lignin biosynthesis and wood formation. The achievements are as below. (1) Based on biochemical and molecular studies, we discovered a novel syringyl-specific alcohol dehydrogenase (SAD) involved in monolignol biosynthesis in angiosperm trees. Through CAld5H/OMT/SAD mediation, syringyl monolignol biosynthesis branches out from guaiacyl pathway at coniferaldehyde; (2) The function of CAld5H gene in this syringyl monolignol biosynthesis pathway also was confirmed in vivo in transgenic Populus; (3) The proposed major monolignol biosynthesis pathways were further supported by the involving biochemical functions of CCR based on a detailed kinetic study; (4) Gene promoter activity analysis also supported the cell-type specific expression of SAD and CAD genes in xylem tissue, consistent with the cell-specific locations of SAD and CAD proteins and with the proposed pathways; (5) We have developed a novel small interfering RNA (siRNA)-mediated stable gene-silencing system in transgenic plants; (6) Using the siRNA and P. trichocarpa transformation/regeneration systems we are currently producing transgenic P. trichocarpa to investigate the interactive functions of CAD and SAD in regulating guaiacyl and syringyl lignin biosynthesis; (7) We have cloned for the first time from a tree species, P. trichocarpa, small regulatory RNAs termed micro

  2. Cyclopiazonic Acid Biosynthesis of Aspergillus flavus and Aspergillus oryzae

    PubMed Central

    Chang, Perng-Kuang; Ehrlich, Kenneth C.; Fujii, Isao

    2009-01-01

    Cyclopiazonic acid (CPA) is an indole-tetramic acid neurotoxin produced by some of the same strains of A. flavus that produce aflatoxins and by some Aspergillus oryzae strains. Despite its discovery 40 years ago, few reviews of its toxicity and biosynthesis have been reported. This review examines what is currently known about the toxicity of CPA to animals and humans, both by itself or in combination with other mycotoxins. The review also discusses CPA biosynthesis and the genetic diversity of CPA production in A. flavus/oryzae populations. PMID:22069533

  3. Engineered Biosynthesis of Natural Products in Heterologous Hosts

    PubMed Central

    Luo, Yunzi; Li, Bing-Zhi; Liu, Duo; Zhang, Lu; Chen, Yan; Jia, Bin; Zeng, Bo-Xuan; Zhao, Huimin; Yuan, Ying-Jin

    2015-01-01

    Natural products produced by microorganisms and plants are a major resource of antibacterial and anticancer drugs as well as industrially useful compounds. However, the native producers often suffer from low productivity and titers. Here we summarize the recent applications of heterologous biosynthesis for the production of several important classes of natural products such as terpenoids, flavonoids, alkaloids, and polyketides. In addition, we will discuss the new tools and strategies at multi-scale levels including gene, pathway, genome and community levels for highly efficient heterologous biosynthesis of natural products. PMID:25960127

  4. Biotin biosynthesis in Mycobacterium tuberculosis: physiology, biochemistry and molecular intervention.

    PubMed

    Salaemae, Wanisa; Azhar, Al; Booker, Grant W; Polyak, Steven W

    2011-09-01

    Biotin is an important micronutrient that serves as an essential enzyme cofactor. Bacteria obtain biotin either through de novo synthesis or by active uptake from exogenous sources. Mycobacteria are unusual amongst bacteria in that their primary source of biotin is through de novo synthesis. Here we review the importance of biotin biosynthesis in the lifecycle of Mycobacteria. Genetic screens designed to identify key metabolic processes have highlighted a role for the biotin biosynthesis in bacilli growth, infection and survival during the latency phase. These studies help to establish the biotin biosynthetic pathway as a potential drug target for new anti-tuberculosis agents. PMID:21976058

  5. Biosynthesis of oxygen and nitrogen-containing heterocycles in polyketides

    PubMed Central

    Hemmerling, Franziska

    2016-01-01

    Summary This review highlights the biosynthesis of heterocycles in polyketide natural products with a focus on oxygen and nitrogen-containing heterocycles with ring sizes between 3 and 6 atoms. Heterocycles are abundant structural elements of natural products from all classes and they often contribute significantly to their biological activity. Progress in recent years has led to a much better understanding of their biosynthesis. In this context, plenty of novel enzymology has been discovered, suggesting that these pathways are an attractive target for future studies. PMID:27559404

  6. Effects of a siderophore receptor and porin proteins-based vaccination on fecal shedding of Escherichia coli O157:H7 in experimentally inoculated cattle.

    PubMed

    Thornton, A B; Thomson, D U; Loneragan, G H; Fox, J T; Burkhardt, D T; Emery, D A; Nagaraja, T G

    2009-04-01

    The efficacy of a vaccine containing outer membrane siderophore receptor and porin (SRP) proteins for reducing fecal prevalence and shedding of Escherichia coli O157:H7 was evaluated in cattle inoculated with E. coli O157:H7. Thirty calves were randomly assigned to one of two groups, and on days 1 and 21 these calves were given subcutaneous injections of either a placebo (control) or the vaccine. Blood was collected weekly to monitor the serum anti-SRP antibody titers. Two weeks after the second vaccination, calves were orally inoculated with a mixture of five strains of nalidixic acid-resistant (NalR) E. coli O157:H7. Fecal samples and rectoanal mucosal swabs were collected daily for the first 5 days and then three times each week for the following 4 weeks to determine the presence and enumerate the fecal concentration of NalR E. coli O157:H7. At necropsy on day 35, gut contents and tissue swabs were collected to determine the presence and concentration of NalR E. coli O157:H7. Vaccinated cattle had significantly higher anti-SRP antibody titers than did control cattle, with a significant treatment x week interaction (P < 0.01). Vaccination of cattle with the SRP protein tended to decrease fecal concentration (1.9 versus 1.6 log CFU/g) of NalR E. coli O157:H7 (P = 0.10). The number of calves that were fecal culture positive for E. coli O157:H7 was lower (P = 0.05) in the vaccinated group than in the control group. The E. coli O157:H7 SRP vaccine tended to reduce fecal prevalence and concentration of E. coli O157:H7 in cattle orally inoculated with NalR E. coli 0157:H7 and may be a useful prehavest intervention strategy. Future research must be conducted on natural prevalence in feedlot operations to further evaluate the efficacy of this novel vaccine. PMID:19435240

  7. Biosynthesis of allene oxides in Physcomitrella patens

    PubMed Central

    2012-01-01

    Background The moss Physcomitrella patens contains C18- as well as C20-polyunsaturated fatty acids that can be metabolized by different enzymes to form oxylipins such as the cyclopentenone cis(+)-12-oxo phytodienoic acid. Mutants defective in the biosynthesis of cyclopentenones showed reduced fertility, aberrant sporophyte morphology and interrupted sporogenesis. The initial step in this biosynthetic route is the conversion of a fatty acid hydroperoxide to an allene oxide. This reaction is catalyzed by allene oxide synthase (AOS) belonging as hydroperoxide lyase (HPL) to the cytochrome P450 family Cyp74. In this study we characterized two AOS from P. patens, PpAOS1 and PpAOS2. Results Our results show that PpAOS1 is highly active with both C18 and C20-hydroperoxy-fatty acid substrates, whereas PpAOS2 is fully active only with C20-substrates, exhibiting trace activity (~1000-fold lower kcat/KM) with C18 substrates. Analysis of products of PpAOS1 and PpHPL further demonstrated that both enzymes have an inherent side activity mirroring the close inter-connection of AOS and HPL catalysis. By employing site directed mutagenesis we provide evidence that single amino acid residues in the active site are also determining the catalytic activity of a 9-/13-AOS – a finding that previously has only been reported for substrate specific 13-AOS. However, PpHPL cannot be converted into an AOS by exchanging the same determinant. Localization studies using YFP-labeled AOS showed that PpAOS2 is localized in the plastid while PpAOS1 may be found in the cytosol. Analysis of the wound-induced cis(+)-12-oxo phytodienoic acid accumulation in PpAOS1 and PpAOS2 single knock-out mutants showed that disruption of PpAOS1, in contrast to PpAOS2, results in a significantly decreased cis(+)-12-oxo phytodienoic acid formation. However, the knock-out mutants of neither PpAOS1 nor PpAOS2 showed reduced fertility, aberrant sporophyte morphology or interrupted sporogenesis. Conclusions Our study

  8. Biosynthesis and Molecular Genetics of Polyketides in Marine Dinoflagellates

    PubMed Central

    Kellmann, Ralf; Stüken, Anke; Orr, Russell J. S.; Svendsen, Helene M.; Jakobsen, Kjetill S.

    2010-01-01

    Marine dinoflagellates are the single most important group of algae that produce toxins, which have a global impact on human activities. The toxins are chemically diverse, and include macrolides, cyclic polyethers, spirolides and purine alkaloids. Whereas there is a multitude of studies describing the pharmacology of these toxins, there is limited or no knowledge regarding the biochemistry and molecular genetics involved in their biosynthesis. Recently, however, exciting advances have been made. Expressed sequence tag sequencing studies have revealed important insights into the transcriptomes of dinoflagellates, whereas other studies have implicated polyketide synthase genes in the biosynthesis of cyclic polyether toxins, and the molecular genetic basis for the biosynthesis of paralytic shellfish toxins has been elucidated in cyanobacteria. This review summarises the recent progress that has been made regarding the unusual genomes of dinoflagellates, the biosynthesis and molecular genetics of dinoflagellate toxins. In addition, the evolution of these metabolic pathways will be discussed, and an outlook for future research and possible applications is provided. PMID:20479965

  9. Two fatty acyl reductases involved in moth pheromone biosynthesis.

    PubMed

    Antony, Binu; Ding, Bao-Jian; Moto, Ken'Ichi; Aldosari, Saleh A; Aldawood, Abdulrahman S

    2016-01-01

    Fatty acyl reductases (FARs) constitute an evolutionarily conserved gene family found in all kingdoms of life. Members of the FAR gene family play diverse roles, including seed oil synthesis, insect pheromone biosynthesis, and mammalian wax biosynthesis. In insects, FAR genes dedicated to sex pheromone biosynthesis (pheromone-gland-specific fatty acyl reductase, pgFAR) form a unique clade that exhibits substantial modifications in gene structure and possesses unique specificity and selectivity for fatty acyl substrates. Highly selective and semi-selective 'single pgFARs' produce single and multicomponent pheromone signals in bombycid, pyralid, yponomeutid and noctuid moths. An intriguing question is how a 'single reductase' can direct the synthesis of several fatty alcohols of various chain lengths and isomeric forms. Here, we report two active pgFARs in the pheromone gland of Spodoptera, namely a semi-selective, C14:acyl-specific pgFAR and a highly selective, C16:acyl-specific pgFAR, and demonstrate that these pgFARs play a pivotal role in the formation of species-specific signals, a finding that is strongly supported by functional gene expression data. The study envisages a new area of research for disclosing evolutionary changes associated with C14- and C16-specific FARs in moth pheromone biosynthesis. PMID:27427355

  10. Shedding light on ovothiol biosynthesis in marine metazoans.

    PubMed

    Castellano, Immacolata; Migliaccio, Oriana; D'Aniello, Salvatore; Merlino, Antonello; Napolitano, Alessandra; Palumbo, Anna

    2016-01-01

    Ovothiol, isolated from marine invertebrate eggs, is considered one of the most powerful antioxidant with potential for drug development. However, its biological functions in marine organisms still represent a matter of debate. In sea urchins, the most accepted view is that ovothiol protects the eggs by the high oxidative burst at fertilization. In this work we address the role of ovothiol during sea urchin development to give new insights on ovothiol biosynthesis in metazoans. The gene involved in ovothiol biosynthesis OvoA was identified in Paracentrotus lividus genome (PlOvoA). PlOvoA embryo expression significantly increased at the pluteus stage and was up-regulated by metals at concentrations mimicking polluted sea-water and by cyclic toxic algal blooms, leading to ovothiol biosynthesis. In silico analyses of the PlOvoA upstream region revealed metal and stress responsive elements. Structural protein models highlighted conserved active site residues likely responsible for ovothiol biosynthesis. Phylogenetic analyses indicated that OvoA evolved in most marine metazoans and was lost in bony vertebrates during the transition from the aquatic to terrestrial environment. These results highlight the crucial role of OvoA in protecting embryos released in seawater from environmental cues, thus allowing the survival under different conditions. PMID:26916575

  11. HypC, the anthrone oxidase involved in aflatoxin biosynthesis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Based on gene disruption and enzyme activity, hypC, an open reading frame in the pksA (aflC)/nor-1 (aflD) intergenic region in the aflatoxin biosynthesis cluster, encodes a 17 kDa oxidase that catalyzes the conversion of norsolorinic acid anthrone to norsolorinic acid....

  12. Sampangine inhibits heme biosynthesis in both yeast and human

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The azaoxoaporphine alkaloid sampangine exhibits strong antiproliferation activity in various organisms. Previous studies suggested that it somehow affects heme metabolism and stimulates production of reactive oxygen species (ROS). In this study, we show that inhibition of heme biosynthesis is the p...

  13. Shedding light on ovothiol biosynthesis in marine metazoans

    PubMed Central

    Castellano, Immacolata; Migliaccio, Oriana; D’Aniello, Salvatore; Merlino, Antonello; Napolitano, Alessandra; Palumbo, Anna

    2016-01-01

    Ovothiol, isolated from marine invertebrate eggs, is considered one of the most powerful antioxidant with potential for drug development. However, its biological functions in marine organisms still represent a matter of debate. In sea urchins, the most accepted view is that ovothiol protects the eggs by the high oxidative burst at fertilization. In this work we address the role of ovothiol during sea urchin development to give new insights on ovothiol biosynthesis in metazoans. The gene involved in ovothiol biosynthesis OvoA was identified in Paracentrotus lividus genome (PlOvoA). PlOvoA embryo expression significantly increased at the pluteus stage and was up-regulated by metals at concentrations mimicking polluted sea-water and by cyclic toxic algal blooms, leading to ovothiol biosynthesis. In silico analyses of the PlOvoA upstream region revealed metal and stress responsive elements. Structural protein models highlighted conserved active site residues likely responsible for ovothiol biosynthesis. Phylogenetic analyses indicated that OvoA evolved in most marine metazoans and was lost in bony vertebrates during the transition from the aquatic to terrestrial environment. These results highlight the crucial role of OvoA in protecting embryos released in seawater from environmental cues, thus allowing the survival under different conditions. PMID:26916575

  14. Shedding light on ovothiol biosynthesis in marine metazoans

    NASA Astrophysics Data System (ADS)

    Castellano, Immacolata; Migliaccio, Oriana; D'Aniello, Salvatore; Merlino, Antonello; Napolitano, Alessandra; Palumbo, Anna

    2016-02-01

    Ovothiol, isolated from marine invertebrate eggs, is considered one of the most powerful antioxidant with potential for drug development. However, its biological functions in marine organisms still represent a matter of debate. In sea urchins, the most accepted view is that ovothiol protects the eggs by the high oxidative burst at fertilization. In this work we address the role of ovothiol during sea urchin development to give new insights on ovothiol biosynthesis in metazoans. The gene involved in ovothiol biosynthesis OvoA was identified in Paracentrotus lividus genome (PlOvoA). PlOvoA embryo expression significantly increased at the pluteus stage and was up-regulated by metals at concentrations mimicking polluted sea-water and by cyclic toxic algal blooms, leading to ovothiol biosynthesis. In silico analyses of the PlOvoA upstream region revealed metal and stress responsive elements. Structural protein models highlighted conserved active site residues likely responsible for ovothiol biosynthesis. Phylogenetic analyses indicated that OvoA evolved in most marine metazoans and was lost in bony vertebrates during the transition from the aquatic to terrestrial environment. These results highlight the crucial role of OvoA in protecting embryos released in seawater from environmental cues, thus allowing the survival under different conditions.

  15. Two fatty acyl reductases involved in moth pheromone biosynthesis

    PubMed Central

    Antony, Binu; Ding, Bao-Jian; Moto, Ken’Ichi; Aldosari, Saleh A.; Aldawood, Abdulrahman S.

    2016-01-01

    Fatty acyl reductases (FARs) constitute an evolutionarily conserved gene family found in all kingdoms of life. Members of the FAR gene family play diverse roles, including seed oil synthesis, insect pheromone biosynthesis, and mammalian wax biosynthesis. In insects, FAR genes dedicated to sex pheromone biosynthesis (pheromone-gland-specific fatty acyl reductase, pgFAR) form a unique clade that exhibits substantial modifications in gene structure and possesses unique specificity and selectivity for fatty acyl substrates. Highly selective and semi-selective ‘single pgFARs’ produce single and multicomponent pheromone signals in bombycid, pyralid, yponomeutid and noctuid moths. An intriguing question is how a ‘single reductase’ can direct the synthesis of several fatty alcohols of various chain lengths and isomeric forms. Here, we report two active pgFARs in the pheromone gland of Spodoptera, namely a semi-selective, C14:acyl-specific pgFAR and a highly selective, C16:acyl-specific pgFAR, and demonstrate that these pgFARs play a pivotal role in the formation of species-specific signals, a finding that is strongly supported by functional gene expression data. The study envisages a new area of research for disclosing evolutionary changes associated with C14- and C16-specific FARs in moth pheromone biosynthesis. PMID:27427355

  16. Cyclopiazonic Acid Biosynthesis of Aspergillus flavus and Aspergillus oryzae

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cyclopiazonic acid (CPA) is an indole-tetramic acid neurotoxin produced by some of the same strains of A. flavus that produce aflatoxins and by some Aspergillus oryzae strains. Despite its discovery 40 years ago, few reviews of its toxicity and biosynthesis have been reported. This review examines w...

  17. Genetic aspects of auxin biosynthesis and its regulation.

    PubMed

    Brumos, Javier; Alonso, Jose M; Stepanova, Anna N

    2014-05-01

    Auxin is an essential plant hormone that controls nearly every aspect of a plant's life, from embryo development to organ senescence. In the last decade the key genes involved in auxin transport, perception, signaling and response have been identified and characterized, but the elucidation of auxin biosynthesis has proven to be especially challenging. In plants, a significant amount of indole-3-acetic acid (IAA), the predominant biologically active form of auxin, is synthesized via a simple two-step route where indole-3-pyruvic acid (IPyA) produced from l-tryptophan by tryptophan aminotransferases (TAA1/TAR) is converted to IAA by the YUC family of flavin monooxygenases. The TAA1/TAR and YUC gene families constitute the first complete auxin biosynthetic pathway described in plants. Detailed characterization of these genes' expression patterns suggested a key role of local auxin biosynthesis in plant development. This has prompted an active search for the molecular mechanisms that regulate the spatiotemporal activity of the IPyA route. In addition to the TAA1/TAR and YUC-mediated auxin biosynthesis, several alternative routes of IAA production have been postulated to function in plants, but their biological significance is yet to be demonstrated. Herein, we take a genetic perspective to describe the current view of auxin biosynthesis and its regulation in plants, focusing primarily on Arabidopsis. PMID:24007561

  18. The magnesium chelation step in chlorophyll biosynthesis. Progress report 1993

    SciTech Connect

    Weinstein, J.D.

    1993-12-31

    Progress is reported on the identification and fractionation of Magnesium chealatase, an enzyme involved in addition of Mg to chlorophyll during the later`s biosynthesis. Progress is documented as a series of synopsis of published and unpublished papers by the author.

  19. Aspirin therapy and thromboxane biosynthesis in systemic lupus erythematosus.

    PubMed

    Avalos, Ingrid; Chung, Cecilia P; Oeser, Annette; Milne, Ginger L; Borntrager, Holly; Morrow, Jason D; Raggi, Paolo; Solus, Joseph; Stein, C Michael

    2007-01-01

    Incomplete suppression of thromboxane biosynthesis during aspirin therapy is associated with increased cardiovascular risk. Since systemic lupus erythematosus (SLE) is associated with platelet activation and increased cardiovascular mortality, we compared thromboxane and prostacyclin biosynthesis in patients with SLE and control subjects, and measured inhibition of thromboxane excretion in aspirin-treated subjects. We measured the urinary excretion of 11-dehydro thromboxane B( 2) (TXB(2)) and 2,3-dinor 6-ketoPGF(1alpha) (PGI-M), the stable metabolites of thromboxane A(2) and prostacyclin, respectively, in 74 patients with SLE and 70 controls. In subjects who were not receiving aspirin, TXB(2) excretion was higher in patients with SLE [0.40 ng/mg creatinine (0.26-0.64), median (interquartile range)] than controls [0.31 ng/mg creatinine (0.23-0.44)] (P = 0.04), and in these patients, TXB(2) excretion correlated with disease activity (rho = 0.28, P = 0.03) and tumor necrosis factor alpha (rho = 0.48, P < 0.001). Aspirin therapy resulted in significantly lower TXB(2) excretion in controls (P = 0.01), but not in patients with SLE (P = 0.10), compared with subjects not receiving aspirin. Prostacyclin biosynthesis did not differ among patients and controls, and was not affected by aspirin (P all >0.35). Thromboxane biosynthesis is increased in SLE and is associated with disease activity. Additionally, response to aspirin may be attenuated in some patients with SLE. PMID:18042592

  20. Exploiting nongenetic cell-to-cell variation for enhanced biosynthesis.

    PubMed

    Xiao, Yi; Bowen, Christopher H; Liu, Di; Zhang, Fuzhong

    2016-05-01

    Biosynthesis enables renewable production of manifold compounds, yet often biosynthetic performance must be improved for it to be economically feasible. Nongenetic, cell-to-cell variations in protein and metabolite concentrations are naturally inherent, suggesting the existence of both high- and low-performance variants in all cultures. Although having an intrinsic source of low performers might cause suboptimal ensemble biosynthesis, the existence of high performers suggests an avenue for performance enhancement. Here we develop in vivo population quality control (PopQC) to continuously select for high-performing, nongenetic variants. We apply PopQC to two biosynthetic pathways using two alternative design principles and demonstrate threefold enhanced production of both free fatty acid (FFA) and tyrosine. We confirm that PopQC improves ensemble biosynthesis by selecting for nongenetic high performers. Additionally, we use PopQC in fed-batch FFA production and achieve 21.5 g l(-1) titer and 0.5 g l(-1) h(-1) productivity. Given the ubiquity of nongenetic variation, PopQC should be applicable to a variety of metabolic pathways for enhanced biosynthesis. PMID:26999780

  1. Anaerobic biosynthesis of the lower ligand of vitamin B12

    PubMed Central

    Hazra, Amrita B.; Han, Andrew W.; Mehta, Angad P.; Mok, Kenny C.; Osadchiy, Vadim; Begley, Tadhg P.; Taga, Michiko E.

    2015-01-01

    Vitamin B12 (cobalamin) is required by humans and other organisms for diverse metabolic processes, although only a subset of prokaryotes is capable of synthesizing B12 and other cobamide cofactors. The complete aerobic and anaerobic pathways for the de novo biosynthesis of B12 are known, with the exception of the steps leading to the anaerobic biosynthesis of the lower ligand, 5,6-dimethylbenzimidazole (DMB). Here, we report the identification and characterization of the complete pathway for anaerobic DMB biosynthesis. This pathway, identified in the obligate anaerobic bacterium Eubacterium limosum, is composed of five previously uncharacterized genes, bzaABCDE, that together direct DMB production when expressed in anaerobically cultured Escherichia coli. Expression of different combinations of the bza genes revealed that 5-hydroxybenzimidazole, 5-methoxybenzimidazole, and 5-methoxy-6-methylbenzimidazole, all of which are lower ligands of cobamides produced by other organisms, are intermediates in the pathway. The bza gene content of several bacterial and archaeal genomes is consistent with experimentally determined structures of the benzimidazoles produced by these organisms, indicating that these genes can be used to predict cobamide structure. The identification of the bza genes thus represents the last remaining unknown component of the biosynthetic pathway for not only B12 itself, but also for three other cobamide lower ligands whose biosynthesis was previously unknown. Given the importance of cobamides in environmental, industrial, and human-associated microbial metabolism, the ability to predict cobamide structure may lead to an improved ability to understand and manipulate microbial metabolism. PMID:26246619

  2. Post-genome research on the biosynthesis of ergot alkaloids.

    PubMed

    Li, Shu-Ming; Unsöld, Inge A

    2006-10-01

    Genome sequencing provides new opportunities and challenges for identifying genes for the biosynthesis of secondary metabolites. A putative biosynthetic gene cluster of fumigaclavine C, an ergot alkaloid of the clavine type, was identified in the genome sequence of ASPERGILLUS FUMIGATUS by a bioinformatic approach. This cluster spans 22 kb of genomic DNA and comprises at least 11 open reading frames (ORFs). Seven of them are orthologous to genes from the biosynthetic gene cluster of ergot alkaloids in CLAVICEPS PURPUREA. Experimental evidence of the identified cluster was provided by heterologous expression and biochemical characterization of two ORFs, FgaPT1 and FgaPT2, in the cluster of A. FUMIGATUS, which show remarkable similarities to dimethylallyltryptophan synthase from C. PURPUREA and function as prenyltransferases. FgaPT2 converts L-tryptophan to dimethylallyltryptophan and thereby catalyzes the first step of ergot alkaloid biosynthesis, whilst FgaPT1 catalyzes the last step of the fumigaclavine C biosynthesis, i. e., the prenylation of fumigaclavine A at C-2 position of the indole nucleus. In addition to information obtained from the gene cluster of ergot alkaloids from C. PURPUREA, the identification of the biosynthetic gene cluster of fumigaclavine C in A. FUMIGATUS opens an alternative way to study the biosynthesis of ergot alkaloids in fungi. PMID:16902860

  3. On the phenotypic spectrum of serine biosynthesis defects.

    PubMed

    El-Hattab, Ayman W; Shaheen, Ranad; Hertecant, Jozef; Galadari, Hassan I; Albaqawi, Badi S; Nabil, Amira; Alkuraya, Fowzan S

    2016-05-01

    L-serine is a non-essential amino acid that is de novo synthesized via the enzymes phosphoglycerate dehydrogenase (PGDH), phosphoserine aminotransferase (PSAT), and phosphoserine phosphatase (PSP). Besides its role in protein synthesis, L-serine is a precursor of a number of important compounds. Serine biosynthesis defects result from deficiencies in PGDH, PSAT, or PSP and have a broad phenotypic spectrum ranging from Neu-Laxova syndrome, a lethal multiple congenital anomaly disease at the severe end to a childhood disease with intellectual disability at the mild end, with infantile growth deficiency, and severe neurological manifestations as an intermediate phenotype. In this report, we present three subjects with serine biosynthesis effects. The first was a stillbirth with Neu-Laxova syndrome and a homozygous mutation in PHGDH. The second was a neonate with growth deficiency, microcephaly, ichthyotic skin lesions, seizures, contractures, hypertonia, distinctive facial features, and a homozygous mutation in PSAT1. The third subject was an infant with growth deficiency, microcephaly, ichthyotic skin lesions, anemia, hypertonia, distinctive facial features, low serine and glycine in plasma and CSF, and a novel homozygous mutation in PHGDH gene. Herein, we also review previous reports of serine biosynthesis defects and mutations in the PHGDH, PSAT1, and PSPH genes, discuss the variability in the phenotypes associated with serine biosynthesis defects, and elaborate on the vital roles of serine and the potential consequences of its deficiency. PMID:26960553

  4. Redundant pathways of sunscreen biosynthesis in a cyanobacterium.

    PubMed

    Spence, Edward; Dunlap, Walter C; Shick, J Malcolm; Long, Paul F

    2012-03-01

    Route of the sun block: according to empirical evidence, sun-screening mycosporine-like amino acids (MAAs) in Eukarya originate from the shikimic acid pathway, whereas in cyanobacteria, biosynthesis of the MAA shinorine reportedly occurs through the pentose phosphate pathway. However, gene deletion shows that the cyanobacterium Anabaena variabilis ATCC 29143 does not biosynthesise shinorine exclusively by this route. PMID:22278966

  5. Brassinosteroids Are Master Regulators of Gibberellin Biosynthesis in Arabidopsis

    PubMed Central

    Unterholzner, Simon J.; Rozhon, Wilfried; Papacek, Michael; Ciomas, Jennifer; Lange, Theo; Kugler, Karl G.; Mayer, Klaus F.; Sieberer, Tobias; Poppenberger, Brigitte

    2015-01-01

    Plant growth and development are highly regulated processes that are coordinated by hormones including the brassinosteroids (BRs), a group of steroids with structural similarity to steroid hormones of mammals. Although it is well understood how BRs are produced and how their signals are transduced, BR targets, which directly confer the hormone’s growth-promoting effects, have remained largely elusive. Here, we show that BRs regulate the biosynthesis of gibberellins (GAs), another class of growth-promoting hormones, in Arabidopsis thaliana. We reveal that Arabidopsis mutants deficient in BR signaling are severely impaired in the production of bioactive GA, which is correlated with defective GA biosynthetic gene expression. Expression of the key GA biosynthesis gene GA20ox1 in the BR signaling mutant bri1-301 rescues many of its developmental defects. We provide evidence that supports a model in which the BR-regulated transcription factor BES1 binds to a regulatory element in promoters of GA biosynthesis genes in a BR-induced manner to control their expression. In summary, our study underscores a role of BRs as master regulators of GA biosynthesis and shows that this function is of major relevance for the growth and development of vascular plants. PMID:26243314

  6. Polyketides in Aspergillus terreus: biosynthesis pathway discovery and application.

    PubMed

    Yin, Ying; Cai, Menghao; Zhou, Xiangshan; Li, Zhiyong; Zhang, Yuanxing

    2016-09-01

    The knowledge of biosynthesis gene clusters, production improving methods, and bioactivity mechanisms is very important for the development of filamentous fungi metabolites. Metabolic engineering and heterologous expression methods can be applied to improve desired metabolite production, when their biosynthesis pathways have been revealed. And, stable supplement is a necessary basis of bioactivity mechanism discovery and following clinical trial. Aspergillus terreus is an outstanding producer of many bioactive agents, and a large part of them are polyketides. In this review, we took polyketides from A. terreus as examples, focusing on 13 polyketide synthase (PKS) genes in A. terreus NIH 2624 genome. The biosynthesis pathways of nine PKS genes have been reported, and their downstream metabolites are lovastatin, terreic acid, terrein, geodin, terretonin, citreoviridin, and asperfuranone, respectively. Among them, lovastatin is a well-known hypolipidemic agent. Terreic acid, terrein, citreoviridin, and asperfuranone show good bioactivities, especially anticancer activities. On the other hand, geodin and terretonin are mycotoxins. So, biosynthesis gene cluster information is important for the production or elimination of them. We also predicted three possible gene clusters that contain four PKS genes by homologous gene alignment with other Aspergillus strains. We think that this is an effective way to mine secondary metabolic gene clusters. PMID:27455860

  7. Radiochemical Assay of Juvenile Hormone Biosynthesis Rate in Ants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Juvenile hormone (JH) is an important insect hormone known to have many effects on development, reproduction,and behavior in both solitary and social insects. This protocol describes how to quantify in vitro biosynthesis rates from excised corpora allata (CA), the glands responsible for JH productio...

  8. YCZ-18 Is a New Brassinosteroid Biosynthesis Inhibitor

    PubMed Central

    Oh, Keimei; Matsumoto, Tadashi; Yamagami, Ayumi; Ogawa, Atushi; Yamada, Kazuhiro; Suzuki, Ryuichiro; Sawada, Takayuki; Fujioka, Shozo; Yoshizawa, Yuko; Nakano, Takeshi

    2015-01-01

    Plant hormone brassinosteroids (BRs) are a group of polyhydroxylated steroids that play critical roles in regulating broad aspects of plant growth and development. The structural diversity of BRs is generated by the action of several groups of P450s. Brassinazole is a specific inhibitor of C-22 hydroxylase (CYP90B1) in BR biosynthesis, and the application use of brassinazole has emerged as an effective way of complementing BR-deficient mutants to elucidate the functions of BRs. In this article, we report a new triazole-type BR biosynthesis inhibitor, YCZ-18. Quantitative analysis the endogenous levels of BRs in Arabidopsis indicated that YCZ-18 significantly decreased the BR contents in plant tissues. Assessment of the binding affinity of YCZ-18to purified recombinant CYP90D1 indicated that YCZ-18 induced a typical type II binding spectrum with a Kd value of approximately 0.79 μM. Analysis of the mechanisms underlying the dwarf phenotype associated with YCZ-18 treatment of Arabidopsis indicated that the chemically induced dwarf phenotype was caused by a failure of cell elongation. Moreover, dissecting the effect of YCZ-18 on the induction or down regulation of genes responsive to BRs indicated that YCZ-18 regulated the expression of genes responsible for BRs deficiency in Arabidopsis. These findings indicate that YCZ-18 is a potent BR biosynthesis inhibitor and has a new target site, C23-hydroxylation in BR biosynthesis. Application of YCZ-18 will be a good starting point for further elucidation of the detailed mechanism of BR biosynthesis and its regulation. PMID:25793645

  9. Basis for the control of purine biosynthesis by purine ribonucleotides.

    PubMed Central

    Itakura, M; Sabina, R L; Heald, P W; Holmes, E W

    1981-01-01

    An animal model was used to determine the basis for the increase in purine biosynthesis that results from hepatic depletion of purine nucleotides, such as seen in patients with type I glycogen storage disease or following fructose administration. Mice were injected intravenously with glucose or fructose, 2.5 mg/g of body weight, and the animals were killed at 0, 3, and 30 min following carbohydrate infusion. Fructose, but not glucose, administration led to a threefold increase in [14C]glycine incorporation into hepatic purine nucleotides documenting an increase in the rate of purine biosynthesis in the liver of fructose-treated animals. In the fructose, but not the glucose-treated animals, there was a reduction in the hepatic content of purine nucleotides that are inhibitory for amidophosphoribosyltransferase, the enzyme that catalyzes the first reaction unique to the pathway of purine biosynthesis. PP-ribose-P, an important metabolite in the control of purine biosynthesis, was increased 2,3-fold in liver following fructose, but not glucose administration. In conjunction with the decrease in inhibitory nucleotides and increase in PP-ribose-P 29% of amidophosphoribosyltransferase was shifted from the large inactive to the small active form of the enzyme. Results of these studies demonstrate that the end-products of the pathway, purine nucleotides, control the activity of the enzyme that catalyzes the first reaction leading to purine nucleotide synthesis either through a direct effect of purine nucleotides on the enzyme, through an indirect effect of the change in nucleotides on PP-ribose-P synthesis, or a combination of these effects. The resultant changes in amidophosphoribosyltransferase conformation and activity provide a basis for understanding the increase in purine biosynthesis that results from hepatic depletion of purine nucleotides. PMID:6162862

  10. Restoration of nucleic acid biosynthesis after clinical death and factors stimulating the process in vivo.

    PubMed

    Konikova, A S; Petukhova, L M; Pogossova, A V; Vinarskaya, A A; Nikulin, V I

    1975-01-01

    The biosynthesis of RNA and DNA falls almost to zero in 60 min after the death of rabbits from anoxia, in all the organs of the body. Rapid artificial cooling of the rabbits to 20 degrees C undertaken within 10 min after death preserved nucleic acid biosynthesis and permitted restoration of life 3-4 h after death, with recovery beginning in 60 min. During the reanimation the addition of ATP to the blood stimulated the restoration of RNA biosynthesis in the spinal cord to a considerable extent; the addition of cocarboxylase to the blood promoted cardiac RNA biosynthesis as well as cardiac and pancreatic DNA biosynthesis during recovery. PMID:1197938

  11. Recent advances in the elucidation of enzymatic function in natural product biosynthesis

    PubMed Central

    Tan, Gao-Yi; Deng, Zixin; Liu, Tiangang

    2016-01-01

    With the successful production of artemisinic acid in yeast, the promising potential of synthetic biology for natural product biosynthesis is now being realized. The recent total biosynthesis of opioids in microbes is considered to be another landmark in this field. The importance and significance of enzymes in natural product biosynthetic pathways have been re-emphasized by these advancements. Therefore, the characterization and elucidation of enzymatic function in natural product biosynthesis are undoubtedly fundamental for the development of new drugs and the heterologous biosynthesis of active natural products. Here, discoveries regarding enzymatic function in natural product biosynthesis over the past year are briefly reviewed. PMID:26989472

  12. Regulatory Cross-Talks and Cascades in Rice Hormone Biosynthesis Pathways Contribute to Stress Signaling

    PubMed Central

    Deb, Arindam; Grewal, Rumdeep K.; Kundu, Sudip

    2016-01-01

    Crosstalk among different hormone signaling pathways play an important role in modulating plant response to both biotic and abiotic stress. Hormone activity is controlled by its bio-availability, which is again influenced by its biosynthesis. Thus, independent hormone biosynthesis pathways must be regulated and co-ordinated to mount an integrated response. One of the possibilities is to use cis-regulatory elements to orchestrate expression of hormone biosynthesis genes. Analysis of CREs, associated with differentially expressed hormone biosynthesis related genes in rice leaf under Magnaporthe oryzae attack and drought stress enabled us to obtain insights about cross-talk among hormone biosynthesis pathways at the transcriptional level. We identified some master transcription regulators that co-ordinate different hormone biosynthesis pathways under stress. We found that Abscisic acid and Brassinosteroid regulate Cytokinin conjugation; conversely Brassinosteroid biosynthesis is affected by both Abscisic acid and Cytokinin. Jasmonic acid and Ethylene biosynthesis may be modulated by Abscisic acid through DREB transcription factors. Jasmonic acid or Salicylic acid biosynthesis pathways are co-regulated but they are unlikely to influence each others production directly. Thus, multiple hormones may modulate hormone biosynthesis pathways through a complex regulatory network, where biosynthesis of one hormone is affected by several other contributing hormones. PMID:27617021

  13. Regulatory Cross-Talks and Cascades in Rice Hormone Biosynthesis Pathways Contribute to Stress Signaling.

    PubMed

    Deb, Arindam; Grewal, Rumdeep K; Kundu, Sudip

    2016-01-01

    Crosstalk among different hormone signaling pathways play an important role in modulating plant response to both biotic and abiotic stress. Hormone activity is controlled by its bio-availability, which is again influenced by its biosynthesis. Thus, independent hormone biosynthesis pathways must be regulated and co-ordinated to mount an integrated response. One of the possibilities is to use cis-regulatory elements to orchestrate expression of hormone biosynthesis genes. Analysis of CREs, associated with differentially expressed hormone biosynthesis related genes in rice leaf under Magnaporthe oryzae attack and drought stress enabled us to obtain insights about cross-talk among hormone biosynthesis pathways at the transcriptional level. We identified some master transcription regulators that co-ordinate different hormone biosynthesis pathways under stress. We found that Abscisic acid and Brassinosteroid regulate Cytokinin conjugation; conversely Brassinosteroid biosynthesis is affected by both Abscisic acid and Cytokinin. Jasmonic acid and Ethylene biosynthesis may be modulated by Abscisic acid through DREB transcription factors. Jasmonic acid or Salicylic acid biosynthesis pathways are co-regulated but they are unlikely to influence each others production directly. Thus, multiple hormones may modulate hormone biosynthesis pathways through a complex regulatory network, where biosynthesis of one hormone is affected by several other contributing hormones. PMID:27617021

  14. Hydrogen sulfide mediates nicotine biosynthesis in tobacco (Nicotiana tabacum) under high temperature conditions.

    PubMed

    Chen, Xiaodong; Chen, Qian; Zhang, Xiaoming; Li, Ruijing; Jia, Yujie; Ef, Abd Allah; Jia, Aiqun; Hu, Liwei; Hu, Xiangyang

    2016-07-01

    Hydrogen sulfide (H2S) acts as a signal to induce many physiological processes in plants, but its role in controlling the biosynthesis of secondary metabolites is not well established. In this study, we found that high temperature (HT) treatment induced nicotine biosynthesis in tobacco (Nicotiana tabacum) and promoted the rapid accumulation of H2S. Furthermore, HT triggered the biosynthesis of jasmonic acid (JA), a plant hormone that promotes nicotine biosynthesis. Suppression of the H2S signal using chemical inhibitors or via RNAi suppression of l-cysteine desulphydrase (L-CD) in transgenic plants, compromised JA production and nicotine biosynthesis under HT treatments, and these inhibitory effects could be reversed by applying exogenous H2S. Based on these data, we propose that H2S is an important trigger of nicotine biosynthesis in tobacco under HT conditions, and that H2S acts upstream of JA signaling by modulating the transcription of genes associated with JA biosynthesis. PMID:27035256

  15. Farnesylation mediates brassinosteroid biosynthesis to regulate abscisic acid responses.

    PubMed

    N