Siderophore biosynthesis genes of Rhizobium sp. …

Most iron in the vertebrate hosts is bound to iron-binding proteins, including transferrin, or is sequestered intracellularly. In order for bacteria to grow and infect an animal host, they must first take up iron effectively. We thus attempted to ascertain whether the catecholate siderophore produced by A. salmonicida allows this important fish pathogen to utilize transferrin-bound iron. For this purpose, we tested the parent and the three mutants constructed in this study to determine their abilities to grow in CM9 with 30 μM apotransferrin, in which most of the iron is expected to be bound to the protein. We found that the parental strain RSP74.1 could grow at acceptable levels in the presence of holotransferrin. However, the growth of the asbG, asbC, and asbD mutants was impaired under these conditions (Fig. ). Interestingly, addition of 100 μM histamine to the asbG mutant partially enhanced growth in the presence of 30 μM transferrin. Altogether, these results indicate that the siderophore produced by A. salmonicida is required for iron assimilation from transferrin and show that A. salmonicida uses the catechol-type siderophore for removal of iron from transferrin rather than relying on a receptor for this iron-binding protein. Other pathogens capture transferrin-bound iron by means of siderophore-independent mechanisms and possess specific outer membrane receptors that recognize holotransferrin, as is the case in Haemophilus and Neisseria species ().

Siderophore biosynthesis genes ofRhizobiumsp. …

Genes involved in alcaligin siderophore biosynthesis are contained in an alcABCDE operon.

The regulatory logic of siderophore biosynthetic genes in bacteria ..

Genomic DNA from A. salmonicida ACR168.1 was partially digested with restriction enzyme Sau3AI and ligated into the SuperCos1 cosmid vector (Stratagene). Recombinant cosmids were packaged in vitro and transduced into E. coli XL1-Blue MR (Stratagene). The cosmid library was screened by performing colony PCR with pools of recombinant clones using primers targeted to genes encoding putative A. salmonicida siderophore biosynthesis genes encoded in plasmids pFMON24 and pFMON46 previously isolated using the FURTA (Table ).

Genes involved in siderophore biosynthesis ..

To determine the transcriptional organization of the siderophore biosynthesis gene cluster, A. salmonicida RSP74.1 was grown until exponential phase in low-iron CM9 (containing 40 μM EDDA), and total RNA was isolated using the RNAwiz isolation reagent (Ambion) by following the manufacturer's instructions. Reverse transcriptase PCR (RT-PCR) analyses were performed with 0.6 μg of RNA pretreated with RQ1 RNase-free DNase (Promega) by using the Moloney murine leukemia virus RT (Invitrogen). For operon mapping, a reverse transcription reaction was performed with primers RT-1 and RT-2, which are homologous to the 3′ ends of asbG and asbC, respectively (Fig. ; see Table S1 in the supplemental material). The resulting cDNAs were used as templates for PCR amplification with Taq polymerase (Bioline), using specific primer pairs for each gene (see Table S1 in the supplemental material). A negative control PCR was performed with total RNA without Moloney murine leukemia virus RT to confirm the lack of genomic DNA contamination in each reaction mixture, and genomic DNA was used as a positive control for PCR.

Production of dihydroxamate siderophore alcaligin by Alcaligines denitrificans subsp.
T1 - Elevated zinc induces siderophore biosynthesis genes and a zntA-like gene in Pseudomonas fluorescens

Biological function Bacteria and fungi

AB - 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.

26/04/2013 · Fungal siderophore biosynthesis is ..

and Armstrong, Sandra K., "Identification and characterization of iron-regulated Bordetella pertussis alcaligin siderophore biosynthesis genes.," Journal of Bacteriology 178, no.

These results indicated that AlcA is one of essential components for the alcaligin siderophore biosynthesis.

Siderophore scaffolds require flexibility and a certain ..

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.