Molecular and General Genetics MGG

, Volume 217, Issue 2–3, pp 233–239 | Cite as

Iron-hydroxamate transport intoEscherichia coli K12: Localization of FhuD in the periplasm and of FhuB in the cytoplasmic membrane

  • Wolfgang Köster
  • Volkmar Braun


ThefhuB, fhuC andfhuD genes encode proteins which catalyze transport of iron(III)-hydroxamate compounds from the periplasm into the cytoplasm ofEscherichia coli. ThefhuB, C, D genes were cloned downstream of a strong phage T7 promoter and transcribed by T7 RNA polymerase. The overexpressed FhuD protein appeared in two forms of 31 and 28 kDa and was released upon conversion of vegetative cells into spheroplasts, suggesting synthesis of FhuD as a precursor and export into the periplasm. The very hydrophobic FhuB protein was found in the cytoplasmic membrane. These properties, together with the previously found homologies in the FhuC protein to ATP-binding proteins, display the characteristics of a periplasmic binding protein dependent transport system across the cytoplasmic membrane. The molecular weight of FhuB and the sequence offhuC, as previously published by us, was confirmed. FhuB exhibited double the size of most hydrophobic proteins of such systems and showed homology between the amino- and carboxy-terminal halves of the protein, indicating duplication of an original gene and subsequent fusion of the two DNA fragments.

Key words

Escherichia coli Iron-hydroxamate transport fhuB, C, D genes 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Ames GFL (1986) Bacterial periplasmic transport systems: structure, mechanism, and evolution. Annu Rev Biochem 55:397–425PubMedCrossRefGoogle Scholar
  2. Braun V, Burkhardt R, Schneider R, Zimmermann L (1982) Chromosomal genes for ColV plasmid — determined iron(III)-aerobactin transport inEscherichia coli. J Bacteriol 151:553–559PubMedGoogle Scholar
  3. Braun V, Gross R, Köster W, Zimmermann L (1983) Plasmid and chromosomal mutants in the iron(III)-aerobactin transport system ofEscherichia coli. Mol Gen Genet 192:131–139PubMedCrossRefGoogle Scholar
  4. Burkhardt R, Braun V (1987) Nucleotide sequence offhuC andfhuD genes involved in iron(III)-hydroxamate transport: domains in FhuC homologous to ATP-binding proteins. Mol Gen Genet 209:49–55PubMedCrossRefGoogle Scholar
  5. Coulton JW, Mason P, Allatt DD (1987)fhuC andfhuD genes for iron(III)-ferrichrome transport intoEscherichia coli K-12. J Bacteriol 169:3844–3849PubMedGoogle Scholar
  6. Fecker L, Braun V (1983) Cloning and expression of thefhu genes involved in iron(III)-hydroxamate uptake ofEscherichia coli. J Bacteriol 156:1301–1314PubMedGoogle Scholar
  7. Friedrich MJ, De Veaux LC, Kadner RJ (1986) Nucleotide sequence of thebtuCED genes involved in vitamin B12 transport inEscherichia coli and homology with components of periplasmic-binding-protein-dependent transport systems. J Bacteriol 167:928–934PubMedGoogle Scholar
  8. Hantke K (1981) Regulation of ferric iron transport inEscherichia coli K12: isolation of a constitutive mutant. Mol Gen Genet 182:288–292PubMedCrossRefGoogle Scholar
  9. Hantke K (1983) Identifikation of an iron uptake system specific for coprogen and rhodotorulic acid inEscherichia coli K-12. Mol Gen Genet 191:301–306PubMedCrossRefGoogle Scholar
  10. Hantke K, Braun V (1978) Functional interaction of thetonA/tonB receptor system in Escherichia coli. J Bacteriol 135:190–197PubMedGoogle Scholar
  11. Higgins CF, Gallagher MP, Mimmack ML, Pearce SR (1988) A family of closely related ATP-binding subunits from prokaryotic and eukaryotic cells. BioEssays 8:111–116PubMedCrossRefGoogle Scholar
  12. Hoffmann H, Fischer E, Kraut H, Braun V (1986) Preparation of the FhuA (TonA) receptor protein from cell envelopes of an overproducing strain ofEscherichia coli K-12. J Bacteriol 166:404–411PubMedGoogle Scholar
  13. Köster W, Braun V (1986) Iron hydroxamate transport ofEscherichia coli: nucleotide sequence of thefhuB gene and identification of the protein. Mol Gen Genet 204:435–442PubMedCrossRefGoogle Scholar
  14. Lugtenberg B, Meijers J, Peters R, van der Hock P, van Alphen L (1975) Electrophoretic resolution of the major outer membrane proteins ofEscherichia coli K-12 into four bands. FEBS Lett 58:254–258PubMedCrossRefGoogle Scholar
  15. Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning: a laboratory manual. Cold Spring Habor Laboratory Press, New YorkGoogle Scholar
  16. Miller JH (1972) Experiments in molecular genetics. Cold Spring Harbor Laboratory Press, New YorkGoogle Scholar
  17. Pierce JR, Earhart CF (1986)Escherichia coli K-12 envelope proteins specifically required for ferrienterobactin uptake. J Bacteriol 166:930–936PubMedGoogle Scholar
  18. Postle K, Skare JT (1988)Escherichia coli TonB protein is exported from the cytoplasm without proteolytic cleavage of its amino terminus. J Biol Chem 263: 11000–11007PubMedGoogle Scholar
  19. Pressler U, Staudenmaier H, Zimmermann L, Braun V (1988) Genetics of the iron dicitrate transport system ofEscherichia coli. J Bacteriol 170:2716–2724PubMedGoogle Scholar
  20. Prody C, Neilands JB (1984) Genetic and biochemical characterization of theEscherichia coli K-12fhuB mutation. J Bacteriol 157:874–880PubMedGoogle Scholar
  21. Randall LL, Hardy SJS (1986) Correlation of competence for export with lack of tertiary structure of the mature species: a study in vivo of maltose-binding protein inE. coli. Cell 46:921–928PubMedCrossRefGoogle Scholar
  22. Reitmeier RAF, Bragg PD (1974) Purification and characterization of a heat modifiable protein from the outer membrane ofEscherichia coli. FEBS Lett 41:195–198CrossRefGoogle Scholar
  23. Reynolds PR, Muttur GP, Bradbeer C (1980) Transport of vitamin B12 inEscherichia coli. Some observations on the role of the gene products of BtuC and TonB. J Biol Chem 255:4313–4319PubMedGoogle Scholar
  24. Schnaitman CA (1971) Solubilization of the cytoplasmic membrane ofEscherichia coli by Triton X-100. J Bacteriol 108:545–552PubMedGoogle Scholar
  25. Staudenmaier H, Van Hove B, Yaraghi Z, Braun V (1989) Nucleotide sequence of thefecBCDE genes and location of the proteins suggest a periplasmic binding protein-dependent transport mechanism for iron(III)-dicitrate inEscherichia coli. J Bacteriol 171, in pressGoogle Scholar
  26. Tabor S, Richardson CC (1985) A bacteriophage T7 RNA polymerase/promoter system for controlled exclusive expression of specific genes. Proc Natl Acad Sci USA 82:1074–1078PubMedCrossRefGoogle Scholar
  27. West SEH, Sparling PF (1987) Aerobactin utilization byNeisseria gonorrhoeae and cloning of a genomic DNA fragment that complementsEscherichia coli fhuB mutations. J Bacteriol 169:3414–3421PubMedGoogle Scholar
  28. Witholt B, Boekhout M, Brock M, Dingma J, van Herrikhuizen H, de Leij L (1976) An efficient and reproducible procedure for the formation of spheroplasts from variously grownEscherichia coli. Anal Biochem 74:160–170PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 1989

Authors and Affiliations

  • Wolfgang Köster
    • 1
  • Volkmar Braun
    • 1
  1. 1.Mikrobiologie IIUniversität TübingenTübingenFederal Republic of Germany

Personalised recommendations