Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1990 May;172(5):2650–2657. doi: 10.1128/jb.172.5.2650-2657.1990

Kinetics of iron acquisition from ferric siderophores by Paracoccus denitrificans.

R J Bergeron 1, W R Weimar 1
PMCID: PMC208909  PMID: 2185228

Abstract

The kinetics of iron accumulation by iron-starved Paracoccus denitrificans during the first 2 min of exposure to 55Fe-labeled ferric siderophore chelates is described. Iron is acquired from the ferric chelate of the natural siderophore L-parabactin in a process exhibiting biphastic kinetics by Lineweaver-Burk analysis. The kinetic data for 1 microM less than [Fe L-parabactin] less than 10 microM fit a regression line which suggests a low-affinity system (Km = 3.9 +/- 1.2 microM, Vmax = 494 pg-atoms of 55Fe min-1 mg of protein-1), whereas the data for 0.1 microM less than or equal to [Fe L-parabactin] less than or equal to 1 microM fit another line consistent with a high-affinity system (Km = 0.24 +/- 0.06 microM, Vmax = 108 pg-atoms of 55Fe min-1 mg of protein-1). The Km of the high-affinity uptake is comparable to the binding affinity we had previously reported for the purified ferric L-parabactin receptor protein in the outer membrane. In marked contrast, ferric D-parabactin data fit a single regression line corresponding to a simple Michaelis-Menten process with comparatively low affinity (Km = 3.1 +/- 0.9 microM, Vmax = 125 pg-atoms of 55Fe min-1 mg of protein-1). Other catecholamide siderophores with an intact oxazoline ring derived from L-threonine (L-homoparabactin, L-agrobactin, and L-vibriobactin) also exhibit biphasic kinetics with a high-affinity component similar to ferric L-parabactin. Circular dichroism confirmed that these ferric chelates, like ferric L-parabactin, exist as the lambda enantiomers. The A forms ferric parabactin (ferrin D- and L-parabactin A), in which the oxazoline ring is hydrolyzed to the open-chain threonyl structure, exhibit linear kinetics with a comparatively high Km (1.4 +/- 0.3 microM) and high Vmax (324 pg-atoms of 55Fe min-1 of protein-1). Furthermore, the marked stereospecificity seen between ferric D- and L-parabactins is absent; i.e., iron acquisition from ferric parabactin A is non stereospecific. The mechanistic implications of these findings in relation to a stereospecific high-affinity binding followed by a nonstereospecific postreceptor processing is discussed.

Full text

PDF
2657

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Arceneaux J. E., Davis W. B., Downer D. N., Haydon A. H., Byers B. R. Fate of labeled hydroxamates during iron transport from hydroxamate-ion chelates. J Bacteriol. 1973 Sep;115(3):919–927. doi: 10.1128/jb.115.3.919-927.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bergeron R. J., Dionis J. B., Elliott G. T., Kline S. J. Mechanism and stereospecificity of the parabactin-mediated iron-transport system in Paracoccus denitrificans. J Biol Chem. 1985 Jul 5;260(13):7936–7944. [PubMed] [Google Scholar]
  3. Bergeron R. J., Weimar W. R., Dionis J. B. Demonstration of ferric L-parabactin-binding activity in the outer membrane of Paracoccus denitrificans. J Bacteriol. 1988 Aug;170(8):3711–3717. doi: 10.1128/jb.170.8.3711-3717.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Cleland W. W. The statistical analysis of enzyme kinetic data. Adv Enzymol Relat Areas Mol Biol. 1967;29:1–32. doi: 10.1002/9780470122747.ch1. [DOI] [PubMed] [Google Scholar]
  5. Corbin J. L., Bulen W. A. The isolation and identification of 2,3-dihydroxybenzoic acid and 2-N,6-N-di-92,3-dihydroxybenzoyl)-L-lysine formed by iron-deficient Azotobacter vinelandii. Biochemistry. 1969 Mar;8(3):757–762. doi: 10.1021/bi00831a002. [DOI] [PubMed] [Google Scholar]
  6. Cox C. D., Rinehart K. L., Jr, Moore M. L., Cook J. C., Jr Pyochelin: novel structure of an iron-chelating growth promoter for Pseudomonas aeruginosa. Proc Natl Acad Sci U S A. 1981 Jul;78(7):4256–4260. doi: 10.1073/pnas.78.7.4256. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Ecker D. J., Emery T. Iron uptake from ferrichrome A and iron citrate in Ustilago sphaerogena. J Bacteriol. 1983 Aug;155(2):616–622. doi: 10.1128/jb.155.2.616-622.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Erecińska M., Deutsch C. J., Davis J. S. Energy coupling to K+ transport in Paracoccus denitrificans. J Biol Chem. 1981 Jan 10;256(1):278–284. [PubMed] [Google Scholar]
  9. Frost G. E., Rosenberg H. The inducible citrate-dependent iron transport system in Escherichia coli K12. Biochim Biophys Acta. 1973 Nov 30;330(1):90–101. doi: 10.1016/0005-2736(73)90287-3. [DOI] [PubMed] [Google Scholar]
  10. Griffiths G. L., Sigel S. P., Payne S. M., Neilands J. B. Vibriobactin, a siderophore from Vibrio cholerae. J Biol Chem. 1984 Jan 10;259(1):383–385. [PubMed] [Google Scholar]
  11. Holman G. D., Busza A. L., Pierce E. J., Rees W. D. Evidence for negative cooperativity in human erythrocyte sugar transport. Biochim Biophys Acta. 1981 Dec 21;649(3):503–514. doi: 10.1016/0005-2736(81)90153-x. [DOI] [PubMed] [Google Scholar]
  12. Huschka H., Naegeli H. U., Leuenberger-Ryf H., Keller-Schierlein W., Winkelmann G. Evidence for a common siderophore transport system but different siderophore receptors in Neurospora crassa. J Bacteriol. 1985 May;162(2):715–721. doi: 10.1128/jb.162.2.715-721.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Isied S. S., Kuo G., Raymond K. N. Coordination isomers of biological iron transport compounds. V. The preparation and chirality of the chromium(III) enterobactin complex and model tris(catechol)chromium(III) analogues. J Am Chem Soc. 1976 Mar 31;98(7):1763–1767. doi: 10.1021/ja00423a021. [DOI] [PubMed] [Google Scholar]
  14. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  15. Lammers P. J., Sanders-Loehr J. Active transport of ferric schizokinen in Anabaena sp. J Bacteriol. 1982 Jul;151(1):288–294. doi: 10.1128/jb.151.1.288-294.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Lodge J. S., Gaines C. G., Arceneaux J. E., Byers B. R. Ferrisiderophore reductase activity in Agrobacterium tumefaciens. J Bacteriol. 1982 Feb;149(2):771–774. doi: 10.1128/jb.149.2.771-774.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Lodge J. S., Gaines C. G., Arceneaux J. E., Byers B. R. Non-hydrolytic release of iron from ferrienterobactin analogs by extracts of Bacillus subtilis. Biochem Biophys Res Commun. 1980 Dec 31;97(4):1291–1295. doi: 10.1016/s0006-291x(80)80006-4. [DOI] [PubMed] [Google Scholar]
  18. Markwell M. A., Haas S. M., Bieber L. L., Tolbert N. E. A modification of the Lowry procedure to simplify protein determination in membrane and lipoprotein samples. Anal Biochem. 1978 Jun 15;87(1):206–210. doi: 10.1016/0003-2697(78)90586-9. [DOI] [PubMed] [Google Scholar]
  19. Müller G., Raymond K. N. Specificity and mechanism of ferrioxamine-mediated iron transport in Streptomyces pilosus. J Bacteriol. 1984 Oct;160(1):304–312. doi: 10.1128/jb.160.1.304-312.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Negrin R. S., Neilands J. B. Ferrichrome transport in inner membrane vesicles of Escherichia coli K12. J Biol Chem. 1978 Apr 10;253(7):2339–2342. [PubMed] [Google Scholar]
  21. Ong S. A., Peterson T., Neilands J. B. Agrobactin, a siderophore from Agrobacterium tumefaciens. J Biol Chem. 1979 Mar 25;254(6):1860–1865. [PubMed] [Google Scholar]
  22. Ratledge C., Patel P. V., Mundy J. Iron transport in Mycobacterium smegmatis: the location of mycobactin by electron microscopy. J Gen Microbiol. 1982 Jul;128(7):1559–1565. doi: 10.1099/00221287-128-7-1559. [DOI] [PubMed] [Google Scholar]
  23. Rosenberg H. Transport of iron into bacterial cells. Methods Enzymol. 1979;56:388–394. doi: 10.1016/0076-6879(79)56036-4. [DOI] [PubMed] [Google Scholar]
  24. Snow G. A. Mycobactins: iron-chelating growth factors from mycobacteria. Bacteriol Rev. 1970 Jun;34(2):99–125. doi: 10.1128/br.34.2.99-125.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Tait G. H. The identification and biosynthesis of siderochromes formed by Micrococcus denitrificans. Biochem J. 1975 Jan;146(1):191–204. doi: 10.1042/bj1460191. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Wang C. C., Newton A. An additional step in the transport of iron defined by the tonB locus of Escherichia coli. J Biol Chem. 1971 Apr 10;246(7):2147–2151. [PubMed] [Google Scholar]
  27. Winkelmann G. Metabolic products of microorganisms. 132. Uptake of iron by Neurospora crassa. 3. Iron transport studies with ferrichrome-type compounds. Arch Mikrobiol. 1974 Jun 7;98(1):39–50. [PubMed] [Google Scholar]

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

RESOURCES