Purification, spectroscopic analysis and biological activity of the macrocyclic dihydroxamate siderophore alcaligin produced by Bordetella pertussis and Bordetella bronchiseptica

Brickman, Timothy J.; Hansel, Jan Gerd; Miller, Marvin J.; Armstrong, Sandra K.

doi:10.1007/bf00144625

Cited by 51 publications

(76 citation statements)

References 25 publications

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“…The dihydroxamate molecule can thus be considered a cyclic dimer of two structural units consisting of 1-amino-4-(N-hydroxylamino)-2(S)-butanol and succinic acid. Consistent with this twofold symmetric structure, high-resolution proton and carbon nuclear magnetic resonance spectroscopy recently determined that the molecule displays a twofold symmetric conformation in aqueous solution, although evidence suggests that multiple C 2 -symmetric conformations of alcaligin coexist in aqueous and methanolic solution (10). Alcaligin shares structural features with the ferrioxamine family of siderophores (30), which are produced by actinomycetes as well as certain enterobacterial species such as H. alvei (55).…”

Section: Discussionmentioning

confidence: 95%

“…Evidence for biological activity of the purified siderophores was gathered with growth stimulation and 55 Fealcaligin transport assays of Bordetella spp. (10). Uptake rates and saturability of iron uptake observed in 55 Fe transport assays were indicative of a high-affinity ferric alcaligin transport system and provided the first direct evidence, beyond simple growth stimulation assays, of alcaligin-mediated Bordetella iron transport.…”

mentioning

confidence: 79%

“…Siderophore assays were performed in triplicate in multiple experimental trials. Purified alcaligin used in control siderophore activity assays and spectroscopic analysis was isolated from B. bronchiseptica cultures by the simplified large-scale method described by Brickman and coworkers (10). Determination of hydroxamates was by the method of Csaky (15) with hydroxylamine hydrochloride as standard.…”

Section: Methodsmentioning

confidence: 99%

“…Recent purification and spectroscopic analysis of Bordetella siderophores (10,33) found that the iron chelators produced by both B. pertussis and B. bronchiseptica were identical to the potent macrocyclic dihydroxamate siderophore alcaligin, 1,8(S), 11,18(S)-tetrahydroxy-1,6,11,16-tetraazacycloeicosane-2,5,12, 15-tetrone (molecular formula, C 16 H 28 N 4 O 8 ; molecular weight, 404), previously isolated from the taxonomically related bacterial species Alcaligenes denitrificans subsp. xylosoxydans (35,36).…”

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confidence: 99%

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The ornithine decarboxylase gene odc is required for alcaligin siderophore biosynthesis in Bordetella spp.: putrescine is a precursor of alcaligin

Brickman

Armstrong

1996

J Bacteriol

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Chromosomal insertions defining Bordetella bronchiseptica siderophore phenotypic complementation group III mutants BRM3 and BRM5 were found to reside approximately 200 to 300 bp apart by restriction mapping of cloned genomic regions associated with the insertion markers. DNA hybridization analysis using B. bronchiseptica genomic DNA sequences flanking the cloned BRM3 insertion marker identified homologous Bordetella pertussis UT25 cosmids that complemented the siderophore biosynthesis defect of the group III B. bronchiseptica mutants. Subcloning and complementation analysis localized the complementing activity to a 2.8-kb B. pertussis genomic DNA region. Nucleotide sequencing identified an open reading frame predicted to encode a polypeptide exhibiting strong similarity at the primary amino acid level with several pyridoxal phosphate-dependent amino acid decarboxylases. Alcaligin production was fully restored to group III mutants by supplementation of iron-depleted culture media with putrescine (1,4-diaminobutane), consistent with defects in an ornithine decarboxylase activity required for alcaligin siderophore biosynthesis. Concordantly, the alcaligin biosynthesis defect of BRM3 was functionally complemented by the heterologous Escherichia coli speC gene encoding an ornithine decarboxylase activity. Enzyme assays confirmed that group III B. bronchiseptica siderophore-deficient mutants lack an ornithine decarboxylase activity required for the biosynthesis of alcaligin. Siderophore production by an analogous mutant of B. pertussis constructed by allelic exchange was undetectable. We propose the designation odc for the gene defined by these mutations that abrogate alcaligin siderophore production. Putrescine is an essential precursor of alcaligin in Bordetella spp.Nutritional iron limitation, mediated primarily by specific host iron-binding glycoproteins, is a front-line host defense against disease-causing infectious agents. Strategies aimed at defeating host iron restriction may involve the action of lowmolecular-mass, high-affinity, ferric iron-specific chelators of microbial origin, termed siderophores, that are synthesized coordinately with their cognate surface receptors and transport machinery in response to iron starvation (28). The role of siderophores in microbial pathogenesis is well established (29,53,54).Bordetella pertussis, the causative agent of human whooping cough or pertussis, and Bordetella bronchiseptica, the agent of swine atrophic rhinitis and kennel cough in dogs, are mucosal pathogens that colonize the upper respiratory tracts of their mammalian hosts. In the first reported molecular genetic studies of Bordetella iron acquisition systems, Armstrong and Clements (3) described the isolation of B. bronchiseptica mutants deficient in siderophore activity following transposon mutagenesis. DNA hybridization analysis using DNA probe sequences flanking the transposon insertions established the existence of homologs of B. bronchiseptica siderophore genes in B. pertussis. Reciprocal cross-feeding ex...

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Section: Discussionmentioning

confidence: 95%

mentioning

confidence: 79%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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The ornithine decarboxylase gene odc is required for alcaligin siderophore biosynthesis in Bordetella spp.: putrescine is a precursor of alcaligin

Brickman

Armstrong

1996

J Bacteriol

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“…Armstrong and Clements isolated and characterized B. bronchiseptica transposon-induced siderophore-deficient mutants; DNA hybridization studies using sequences flanking those transposon insertions confirmed the existence of homologs of B. bronchiseptica siderophore system genes in B. pertussis (6). The siderophores of B. pertussis and B. bronchiseptica have been characterized structurally, and each has been identified as the macrocyclic dihydroxamate siderophore known as alcaligin (11,37), previously isolated from the taxonomically related bacterial species Alcaligenes denitrificans subsp. xylosoxydans (41,42).…”

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confidence: 94%

Identification and characterization of iron-regulated Bordetella pertussis alcaligin siderophore biosynthesis genes

et al. 1996

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Bordetella bronchiseptica mutants BRM1, BRM6, and BRM9 fail to produce the native dihydroxamate siderophore alcaligin. A 4.5-kb BamHI-SmaI Bordetella pertussis genomic DNA fragment carried multiple genes required to restore alcaligin production to these siderophore-deficient mutants. Phenotypic complementation analysis using subclones of the 4.5-kb genomic region demonstrated that the closely linked BRM1 and BRM9 mutations were genetically separable from the BRM6 mutation, and both insertions exerted strong polar effects on expression of the downstream gene defined by the BRM6 mutation, suggesting a polycistronic transcriptional organization of these alcaligin biosynthesis genes. Subcloning and complementation experiments localized the putative Bordetella promoter to a 0.7-kb BamHI-SphI subregion of the cloned genomic DNA fragment. Nucleotide sequencing, phenotypic analysis of mutants, and protein expression by the 4.5-kb DNA fragment in Escherichia coli suggested the presence of three alcaligin system genes, namely, alcA, alcB, and alcC. The deduced protein products of alcA, alcB, and alcC have significant primary amino acid sequence similarities with known microbial siderophore biosynthesis enzymes. Primer extension analysis mapped the transcriptional start site of the putative alcaligin biosynthesis operon containing alcABC to a promoter region overlapping a proposed Fur repressor-binding site and demonstrated iron regulation at the transcriptional level.Iron is a fundamental nutritional requirement for virtually all cells, and its assimilation is considered essential for invading pathogenic bacteria to establish infection in the iron-limiting environment of the host (13, 56). Additionally, iron serves as an environmental modulator of the production of certain virulence factors in a number of bacteria (14,24,31,32,46). Despite host iron sequestration, mediated primarily by the glycoprotein family of iron-binding transferrins, pathogens multiply successfully in vivo because they express efficient ironscavenging systems in response to decreased iron availability. These iron retrieval systems utilize two general strategies: one involving high-affinity iron-chelating soluble siderophores (30,40) and the other using siderophore-independent cell surface receptor mechanisms allowing iron uptake directly from host sources such as transferrin, lactoferrin, and heme compounds (7,35,38,54).Bordetella pertussis, the causative agent of human whooping cough or pertussis, and Bordetella bronchiseptica, the agent of swine atrophic rhinitis and kennel cough in dogs, are bacterial pathogens that infect the respiratory epithelial mucosae of their hosts. Early reports described the production of putative siderophores by both B. pertussis and B. bronchiseptica in response to iron deficiency (1, 23). Armstrong and Clements isolated and characterized B. bronchiseptica transposon-induced siderophore-deficient mutants; DNA hybridization studies using sequences flanking those transposon insertions confirmed the existence of homologs o...

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Ironed Out: A Bacterial Siderophore Primer

Lakes

Rees

Abergel

2019

Encyclopedia of Inorganic and Bioinorganic Chemistry

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Nearly all living organisms require iron, the fourth most abundant element in the earth's crust, for survival. However, despite its abundance, ferric ion has extremely low solubility in aerobic environments at physiological pH (10 −18 M), and the Fe II /Fe III redox couple leads the production of destructive oxygen radicals via Fenton chemistry. To solubilize and regulate iron in biological systems, many organisms rely on proteins and small molecules to chelate iron and make it bioavailable. This tedious management of the distribution of iron has led to an arms race between mammals and microorganisms, each with the purpose to ensure their own supply of iron at the expense of the other. In order to acquire iron, microorganisms such as bacteria and fungi have developed small‐molecule chelators called siderophores that bind environmental iron and are actively reincorporated into the cell via specific receptors. When a bacterial pathogen invades a mammalian host, its ability to steal iron from the host's iron stores is determinant in the success of its pathogenicity. This article seeks to provide a high‐level perspective on the structural variety and coordination properties of siderophores, as well as on the biological relevance of siderophore‐mediated iron acquisition for bacterial survival. The promise of using or altering these iron acquisition pathways for medicinal and theranostic applications is also discussed.

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Purification, spectroscopic analysis and biological activity of the macrocyclic dihydroxamate siderophore alcaligin produced by Bordetella pertussis and Bordetella bronchiseptica

Cited by 51 publications

References 25 publications

The ornithine decarboxylase gene odc is required for alcaligin siderophore biosynthesis in Bordetella spp.: putrescine is a precursor of alcaligin

The ornithine decarboxylase gene odc is required for alcaligin siderophore biosynthesis in Bordetella spp.: putrescine is a precursor of alcaligin

Identification and characterization of iron-regulated Bordetella pertussis alcaligin siderophore biosynthesis genes

Ironed Out: A Bacterial Siderophore Primer

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