• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of jbacterPermissionsJournals.ASM.orgJournalJB ArticleJournal InfoAuthorsReviewers
J Bacteriol. Aug 1996; 178(16): 4877–4884.
PMCID: PMC178270

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

Abstract

Bordetella bronchiseptica mutants BRM1, BRM6, and BRM9 fail to produce the native dihydroxamate siderophore alcaligin. A 4.5-kb BamHI-Smal 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.

Full Text

The Full Text of this article is available as a PDF (375K).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Agiato LA, Dyer DW. Siderophore production and membrane alterations by Bordetella pertussis in response to iron starvation. Infect Immun. 1992 Jan;60(1):117–123. [PMC free article] [PubMed]
  • Alexeyev MF. Three kanamycin resistance gene cassettes with different polylinkers. Biotechniques. 1995 Jan;18(1):52–56. [PubMed]
  • Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. [PubMed]
  • Aricó B, Miller JF, Roy C, Stibitz S, Monack D, Falkow S, Gross R, Rappuoli R. Sequences required for expression of Bordetella pertussis virulence factors share homology with prokaryotic signal transduction proteins. Proc Natl Acad Sci U S A. 1989 Sep;86(17):6671–6675. [PMC free article] [PubMed]
  • Armstrong SK, Clements MO. Isolation and characterization of Bordetella bronchiseptica mutants deficient in siderophore activity. J Bacteriol. 1993 Feb;175(4):1144–1152. [PMC free article] [PubMed]
  • Blanton KJ, Biswas GD, Tsai J, Adams J, Dyer DW, Davis SM, Koch GG, Sen PK, Sparling PF. Genetic evidence that Neisseria gonorrhoeae produces specific receptors for transferrin and lactoferrin. J Bacteriol. 1990 Sep;172(9):5225–5235. [PMC free article] [PubMed]
  • Brickman TJ, Armstrong SK. Bordetella pertussis fur gene restores iron repressibility of siderophore and protein expression to deregulated Bordetella bronchiseptica mutants. J Bacteriol. 1995 Jan;177(1):268–270. [PMC free article] [PubMed]
  • Brickman TJ, Armstrong SK. The ornithine decarboxylase gene odc is required for alcaligin siderophore biosynthesis in Bordetella spp.: putrescine is a precursor of alcaligin. J Bacteriol. 1996 Jan;178(1):54–60. [PMC free article] [PubMed]
  • Brickman TJ, Hansel JG, Miller MJ, Armstrong SK. Purification, spectroscopic analysis and biological activity of the macrocyclic dihydroxamate siderophore alcaligin produced by Bordetella pertussis and Bordetella bronchiseptica. Biometals. 1996 Apr;9(2):191–203. [PubMed]
  • Brown DR, Parker CD. Cloning of the filamentous hemagglutinin of Bordetella pertussis and its expression in Escherichia coli. Infect Immun. 1987 Jan;55(1):154–161. [PMC free article] [PubMed]
  • Bullen JJ. The significance of iron in infection. Rev Infect Dis. 1981 Nov-Dec;3(6):1127–1138. [PubMed]
  • Calderwood SB, Mekalanos JJ. Iron regulation of Shiga-like toxin expression in Escherichia coli is mediated by the fur locus. J Bacteriol. 1987 Oct;169(10):4759–4764. [PMC free article] [PubMed]
  • Chomczynski P, Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. [PubMed]
  • de Lorenzo V, Bindereif A, Paw BH, Neilands JB. Aerobactin biosynthesis and transport genes of plasmid ColV-K30 in Escherichia coli K-12. J Bacteriol. 1986 Feb;165(2):570–578. [PMC free article] [PubMed]
  • de Lorenzo V, Wee S, Herrero M, Neilands JB. Operator sequences of the aerobactin operon of plasmid ColV-K30 binding the ferric uptake regulation (fur) repressor. J Bacteriol. 1987 Jun;169(6):2624–2630. [PMC free article] [PubMed]
  • DeShazer D, Wood GE, Friedman RL. Boiling eliminates artifact banding when sequencing double-stranded templates. Biotechniques. 1994 Aug;17(2):288–290. [PubMed]
  • Figurski DH, Helinski DR. Replication of an origin-containing derivative of plasmid RK2 dependent on a plasmid function provided in trans. Proc Natl Acad Sci U S A. 1979 Apr;76(4):1648–1652. [PMC free article] [PubMed]
  • Giardina PC, Foster LA, Musser JM, Akerley BJ, Miller JF, Dyer DW. bvg Repression of alcaligin synthesis in Bordetella bronchiseptica is associated with phylogenetic lineage. J Bacteriol. 1995 Nov;177(21):6058–6063. [PMC free article] [PubMed]
  • Giardina PC, Foster LA, Toth SI, Roe BA, Dyer DW. Identification of alcA, a Bordetella bronchiseptica gene necessary for alcaligin production. Gene. 1995 Dec 29;167(1-2):133–136. [PubMed]
  • Gorringe AR, Woods G, Robinson A. Growth and siderophore production by Bordetella pertussis under iron-restricted conditions. FEMS Microbiol Lett. 1990 Jan 1;54(1-3):101–105. [PubMed]
  • Grant CC, Vasil ML. Analysis of transcription of the exotoxin A gene of Pseudomonas aeruginosa. J Bacteriol. 1986 Dec;168(3):1112–1119. [PMC free article] [PubMed]
  • Hawley DK, McClure WR. Compilation and analysis of Escherichia coli promoter DNA sequences. Nucleic Acids Res. 1983 Apr 25;11(8):2237–2255. [PMC free article] [PubMed]
  • Hein J. Unified approach to alignment and phylogenies. Methods Enzymol. 1990;183:626–645. [PubMed]
  • Herrero M, de Lorenzo V, Neilands JB. Nucleotide sequence of the iucD gene of the pColV-K30 aerobactin operon and topology of its product studied with phoA and lacZ gene fusions. J Bacteriol. 1988 Jan;170(1):56–64. [PMC free article] [PubMed]
  • Keen NT, Tamaki S, Kobayashi D, Trollinger D. Improved broad-host-range plasmids for DNA cloning in gram-negative bacteria. Gene. 1988 Oct 15;70(1):191–197. [PubMed]
  • Kovach ME, Phillips RW, Elzer PH, Roop RM, 2nd, Peterson KM. pBBR1MCS: a broad-host-range cloning vector. Biotechniques. 1994 May;16(5):800–802. [PubMed]
  • Litwin CM, Calderwood SB. Role of iron in regulation of virulence genes. Clin Microbiol Rev. 1993 Apr;6(2):137–149. [PMC free article] [PubMed]
  • Lory S. Effect of iron on accumulation of exotoxin A-specific mRNA in Pseudomonas aeruginosa. J Bacteriol. 1986 Dec;168(3):1451–1456. [PMC free article] [PubMed]
  • Mei B, Budde AD, Leong SA. sid1, a gene initiating siderophore biosynthesis in Ustilago maydis: molecular characterization, regulation by iron, and role in phytopathogenicity. Proc Natl Acad Sci U S A. 1993 Feb 1;90(3):903–907. [PMC free article] [PubMed]
  • Mietzner TA, Morse SA. The role of iron-binding proteins in the survival of pathogenic bacteria. Annu Rev Nutr. 1994;14:471–493. [PubMed]
  • Moore CH, Foster LA, Gerbig DG, Jr, Dyer DW, Gibson BW. Identification of alcaligin as the siderophore produced by Bordetella pertussis and B. bronchiseptica. J Bacteriol. 1995 Feb;177(4):1116–1118. [PMC free article] [PubMed]
  • Morton DJ, Williams P. Siderophore-independent acquisition of transferrin-bound iron by Haemophilus influenzae type b. J Gen Microbiol. 1990 May;136(5):927–933. [PubMed]
  • Musser JM, Hewlett EL, Peppler MS, Selander RK. Genetic diversity and relationships in populations of Bordetella spp. J Bacteriol. 1986 Apr;166(1):230–237. [PMC free article] [PubMed]
  • Neilands JB. Siderophores: structure and function of microbial iron transport compounds. J Biol Chem. 1995 Nov 10;270(45):26723–26726. [PubMed]
  • Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. [PMC free article] [PubMed]
  • Scarlato V, Prugnola A, Aricò B, Rappuoli R. The bvg-dependent promoters show similar behaviour in different Bordetella species and share sequence homologies. Mol Microbiol. 1991 Oct;5(10):2493–2498. [PubMed]
  • Schmitt MP, Holmes RK. Iron-dependent regulation of diphtheria toxin and siderophore expression by the cloned Corynebacterium diphtheriae repressor gene dtxR in C. diphtheriae C7 strains. Infect Immun. 1991 Jun;59(6):1899–1904. [PMC free article] [PubMed]
  • Schneider DR, Parker CD. Effect of pyridines on phenotypic properties of Bordetella pertussis. Infect Immun. 1982 Nov;38(2):548–553. [PMC free article] [PubMed]
  • Schwyn B, Neilands JB. Universal chemical assay for the detection and determination of siderophores. Anal Biochem. 1987 Jan;160(1):47–56. [PubMed]
  • Stainer DW, Scholte MJ. A simple chemically defined medium for the production of phase I Bordetella pertussis. J Gen Microbiol. 1970 Oct;63(2):211–220. [PubMed]
  • Stibitz S. Use of conditionally counterselectable suicide vectors for allelic exchange. Methods Enzymol. 1994;235:458–465. [PubMed]
  • Stojiljkovic I, Bäumler AJ, Hantke K. Fur regulon in gram-negative bacteria. Identification and characterization of new iron-regulated Escherichia coli genes by a fur titration assay. J Mol Biol. 1994 Feb 18;236(2):531–545. [PubMed]
  • Tabor S, Richardson CC. A bacteriophage T7 RNA polymerase/promoter system for controlled exclusive expression of specific genes. Proc Natl Acad Sci U S A. 1985 Feb;82(4):1074–1078. [PMC free article] [PubMed]
  • Tsai J, Dyer DW, Sparling PF. Loss of transferrin receptor activity in Neisseria meningitidis correlates with inability to use transferrin as an iron source. Infect Immun. 1988 Dec;56(12):3132–3138. [PMC free article] [PubMed]
  • Visca P, Ciervo A, Orsi N. Cloning and nucleotide sequence of the pvdA gene encoding the pyoverdin biosynthetic enzyme L-ornithine N5-oxygenase in Pseudomonas aeruginosa. J Bacteriol. 1994 Feb;176(4):1128–1140. [PMC free article] [PubMed]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...