All Title Author
Keywords Abstract

PLOS ONE  2012 

Purification and Structural Characterization of Siderophore (Corynebactin) from Corynebacterium diphtheriae

DOI: 10.1371/journal.pone.0034591

Full-Text   Cite this paper   Add to My Lib

Abstract:

During infection, Corynebacterium diphtheriae must compete with host iron-sequestering mechanisms for iron. C. diphtheriae can acquire iron by a siderophore-dependent iron-uptake pathway, by uptake and degradation of heme, or both. Previous studies showed that production of siderophore (corynebactin) by C. diphtheriae is repressed under high-iron growth conditions by the iron-activated diphtheria toxin repressor (DtxR) and that partially purified corynebactin fails to react in chemical assays for catecholate or hydroxamate compounds. In this study, we purified corynebactin from supernatants of low-iron cultures of the siderophore-overproducing, DtxR-negative mutant strain C. diphtheriae C7(β) ΔdtxR by sequential anion-exchange chromatography on AG1-X2 and Source 15Q resins, followed by reverse-phase high-performance liquid chromatography (RP-HPLC) on Zorbax C8 resin. The Chrome Azurol S (CAS) chemical assay for siderophores was used to detect and measure corynebactin during purification, and the biological activity of purified corynebactin was shown by its ability to promote growth and iron uptake in siderophore-deficient mutant strains of C. diphtheriae under iron-limiting conditions. Mass spectrometry and NMR analysis demonstrated that corynebactin has a novel structure, consisting of a central lysine residue linked through its α- and ε- amino groups by amide bonds to the terminal carboxyl groups of two different citrate residues. Corynebactin from C. diphtheriae is structurally related to staphyloferrin A from Staphylococcus aureus and rhizoferrin from Rhizopus microsporus in which d-ornithine or 1,4-diaminobutane, respectively, replaces the central lysine residue that is present in corynebactin.

References

[1]  Albrecht-Gary AM, Crumbliss AL (1998) Coordination chemistry of siderophores: thermodynamics and kinetics of iron chelation and release. Met Ions Biol Syst 35: 239–327.
[2]  Boukhalfa H, Crumbliss AL (2002) Chemical aspects of siderophore mediated iron transport. Biometals 15: 325–339.
[3]  Stintzi A, Raymond KN, editors. (2002) Siderophore chemistry. New York: Marcel Dekker, Inc. pp. 273–320.
[4]  Winkelmann G (1991) Handbook of microbial iron chelates. Boca Raton, FL: CRC Press.
[5]  Brickman TJ, McIntosh MA (1992) Overexpression and purification of ferric enterobactin esterase from Escherichia coli. Demonstration of enzymatic hydrolysis of enterobactin and its iron complex. J Biol Chem 267: 12350–12355.
[6]  Bell CF (1977) Metal Chelation Principles and Applications. Oxford: Clarendon Press.
[7]  Winkelmann G (2002) Microbial siderophore-mediated transport. Biochem Soc Trans 30: 691–696.
[8]  Budzikiewicz H (2005) Bacterial Citrate Siderophores. Mini-Reviews in Organic Chemistry 2: 119–124.
[9]  Smith MJ, Shoolery JN, Schwyn B, Holden I, Neilands JB (1985) Rhizobactin, a structurally novel siderophore from Rhizobium meliloti. J Am Chem Soc 107: 1739–1743.
[10]  Munzinger M, Taraz K, Budzikiewicz H, Drechsel H, Heymann P, et al. (1999) S,S-rhizoferrin (enantio-rhizoferrin)-a siderophore of Ralstonia (Pseudomonas) pickettii DSM6297-the optical antipode of R-rhizoferrin isolated from fungi. Biometals 12: 189–193.
[11]  Thieken A, Winkelmann G (1992) Rhizoferrin: a complexone type siderophore of the Mucorales and entomophthorales (Zygomycetes). FEMS Microbiol Lett 73: 37–41.
[12]  Konetschny-Rapp S, Jung G, Meiwes J, Zahner H (1990) Staphyloferrin A: a structurally new siderophore from staphylococci. Eur J Biochem 191: 65–74.
[13]  Meiwes J, Fiedler HP, Haag H, Zahner H, Konetschny-Rapp S, et al. (1990) Isolation and characterization of staphyloferrin A, a compound with siderophore activity from Staphylococcus hyicus DSM 20459. FEMS Microbiol Lett 55: 201–205.
[14]  Russell LM, Cryz SJ Jr, Holmes RK (1984) Genetic and biochemical evidence for a siderophore-dependent iron transport system in Corynebacterium diphtheriae. Infect Immun 45: 143–149.
[15]  Russell LM, Holmes RK (1983) Initial characterization of the ferric iron transport system of Corynebacterium diphtheriae. J Bacteriol 155: 1439–1442.
[16]  Russell LM, Holmes RK (1985) Highly toxinogenic but avirulent Park-Williams 8 strain of Corynebacterium diphtheriae does not produce siderophore. Infect Immun 47: 575–578.
[17]  Park WH, Williams AW (1896) The production of diphtheria toxin. J Exp Med 1: 164–185.
[18]  Kunkle CA, Schmitt MP (2005) Analysis of a DtxR-regulated iron transport and siderophore biosynthesis gene cluster in Corynebacterium diphtheriae. J Bacteriol 187: 422–433.
[19]  Budzikiewicz H, Bossenkamp A, Taraz K, Pandey A, Meyer JM (1997) Corynebactin, a cyclic catecholate siderophore from Corynebacterium glutamicum ATCC 14067 (Brevibacterium sp. DSM 20411). Z Naturforsch C biosci 52: 551–554.
[20]  May JJ, Wendrich TM, Marahiel MA (2001) The dhb operon of Bacillus subtilis encodes the biosynthetic template for the catecholic siderophore 2,3-dihydroxybenzoate-glycine-threonine trimeric ester bacillibactin. J Biol Chem 276: 7209–7217.
[21]  Dertz EA, Stintzi A, Raymond KN (2006) Siderophore-mediated iron transport in Bacillus subtilis and Corynebacterium glutamicum. J Biol Inorg Chem 11: 1087–1097.
[22]  Ikeda M, Nakagawa S (2003) The Corynebacterium glutamicum genome: features and impacts on biotechnological processes. Appl Microbiol Biotechnol 62: 99–109.
[23]  Kalinowski J, Bathe B, Bartels D, Bischoff N, Bott M, et al. (2003) The complete Corynebacterium glutamicum ATCC 13032 genome sequence and its impact on the production of -aspartate-derived amino acids and vitamins. J Biotechnol 104: 5–25.
[24]  Oram DM, Jacobson AD, Holmes RK (2006) Transcription of the contiguous sigB, dtxR, and galE genes in Corynebacterium diphtheriae: evidence for multiple transcripts and regulation by environmental factors. J Bacteriol 188: 2959–2973.
[25]  Drechsel H, Winkelmann G (2005) The configuration of the chiral carbon atoms in staphyloferrin A and analysis of the transport properties in Staphylococcus aureus. Biometals 18: 75–81.
[26]  Drechsel H, Thieken A, Reissbrodt R, Jung G, Winkelmann G (1993) Alpha-keto acids are novel siderophores in the genera Proteus, Providencia, and Morganella and are produced by amino acid deaminases. J Bacteriol 175: 2727–2733.
[27]  Raymond KN, Dertz EA, Kim SS (2003) Enterobactin: an archetype for microbial iron transport. Proc Natl Acad Sci U S A 100: 3584–3588.
[28]  Miller DA, Luo L, Hillson N, Keating TA, Walsh CT (2002) Yersiniabactin synthetase: a four-protein assembly line producing the nonribosomal peptide/polyketide hybrid siderophore of Yersinia pestis. Chem Biol 9: 333–344.
[29]  Harris WR, Carrano CJ, Raymond KN (1979) Coordination Chemistry of Microbial Iron Transport Compounds. Isolation, Characterization, and Formation Constants of Ferric Aerobactin. J Am Chem Soc 101: 2722–2727.
[30]  Lynch D, O'Brien J, Welch T, Clarke P, Cuiv PO, et al. (2001) Genetic organization of the region encoding regulation, biosynthesis, and transport of rhizobactin 1021, a siderophore produced by Sinorhizobium meliloti. J Bacteriol 183: 2576–2585.
[31]  Franza T, Mahe B, Expert D (2005) Erwinia chrysanthemi requires a second iron transport route dependent of the siderophore achromobactin for extracellular growth and plant infection. Mol Microbiol 55: 261–275.
[32]  Beasley FC, Vines ED, Grigg JC, Zheng Q, Liu S, et al. (2009) Characterization of staphyloferrin A biosynthetic and transport mutants in Staphylococcus aureus. Mol Microbiol 72: 947–963.
[33]  Cheung J, Beasley FC, Liu S, Lajoie GA, Heinrichs DE (2009) Molecular characterization of staphyloferrin B biosynthesis in Staphylococcus aureus. Mol Microbiol 74: 594–608.
[34]  Challis GL (2005) A widely distributed bacterial pathway for siderophore biosynthesis independent of nonribosomal peptide synthetases. Chembiochem 6: 601–611.
[35]  Cotton JL, Tao J, Balibar CJ (2009) Identification and characterization of the Staphylococcus aureus gene cluster coding for staphyloferrin A. Biochemistry 48: 1025–1035.
[36]  Muller K, Matzanke BF, Schunemann V, Trautwein AX, Hantke K (1998) FhuF, an iron-regulated protein of Escherichia coli with a new type of [2Fe-2S] center. Eur J Biochem 258: 1001–1008.
[37]  Freeman VJ (1951) Studies on the virulence of bacteriophage-infected strains of Corynebacterium diphtheriae. J Bacteriol 61: 675–688.
[38]  Zajdowicz SL, Jones-Carson J, Vazquez-Torres A, Jobling MG, Gill RE, et al. (2011) Alanine racemase mutants of Burkholderia pseudomallei and Burkholderia mallei and use of alanine racemase as a non-antibiotic-based selectable marker. PLoS One 6: e21523.
[39]  Schafer A, Tauch A, Jager W, Kalinowski J, Thierbach G, et al. (1994) Small mobilizable multi-purpose cloning vectors derived from the Escherichia coli plasmids pK18 and pK19: selection of defined deletions in the chromosome of Corynebacterium glutamicum. Gene 145: 69–73.
[40]  Markley JL, Bax A, Arata Y, Hilbers CW, Kaptein R, et al. (1998) Recommendations for the presentation of NMR structures of proteins and nucleic acids. IUPAC-IUBMB-IUPAB Inter-Union Task Group on the Standardization of Data Bases of Brotein and Nucleic Acid Structures Determined by NMR Spectroscopy. J Biomol NMR 12: 1–23.
[41]  Piantini U, Sorensen OW, Ernst RR (1982) Multiple Quantum filters for Elucidating NMR Coupling Networks. J Am Chem Soc 104: 6800–6801.
[42]  Rance M, Sorensen OW, Bodenhausen G, Wagner G, Ernst RR, et al. (1983) Improved spectral resolution in cosy 1H NMR spectra of proteins via double quantum filtering. Biochem Biophys Res Commun 117: 479–485.
[43]  Aue WP, Bartholdi E, Ernst RR (1976) 2-Dimensional Spectroscopy-Application to Nuclear Magnetic-Resonance. Journal of Chemical Physics 64: 2229–2246.
[44]  Bodenhausen G, Ruben DJ (1980) Natural Abundance N-15 NMR by Enhanced Heteronuclear Spectroscopy. Chemical Physics Letters 69: 185–189.
[45]  Muller L (1979) Sensitivity Enhanced Detection of Weak Nuclei Using Heteronuclear Multiple Quantum Coherence. Journal of the American Chemical Society 101: 4481–4484.
[46]  Marion D, Ikura M, Tschudin R, Bax A (1989) Rapid Recording of 2D NMR-Spectra without Phase Cycling - Application to the Study of Hydrogen-Exchange in Proteins. Journal of Magnetic Resonance 85: 393–399.
[47]  Delaglio F, Grzesiek S, Vuister GW, Zhu G, Pfiefer J, et al. (1995) NMRPipe: a multidimensional spectral processing system based on UNIX pipes. J Biomol NMR 6: 277–293.
[48]  Goddard TD, Kneller DG (1997) Sparky 3. University of California San Francisco. Available: http://www.cgl.ucsf.edu/home/sparky/. Accessed 2012 Mar 19.
[49]  Arnow L (1937) Colorimetric determination of the componenets of 3,4-dihydroxyphenylalanine-tyrosine mixtures. J Biol Chem 118: 531–537.
[50]  Csaky TZ (1948) On the estimation of bound hydroxylamine in biological materials. Acta Chem Scand 2: 450–454.

Full-Text

comments powered by Disqus