This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Killmann, H Articles by Braun, V Search for Related Content PubMed PubMed Citation Articles by Killmann, H Articles by Braun, V

research-article

An aspartate deletion mutation defines a binding site of the multifunctional FhuA outer membrane receptor of Escherichia coli K-12.

Mikrobiologie II, Universität Tübingen, Germany.

ABSTRACT

The FhuA protein of the outer membrane serves as a receptor for phages T5, T1, and phi 80, for colicin M, for the antibiotic albomycin, and for ferrichrome and related siderophores. To identify protein regions important for the multiple FhuA activities, fhuA genes of spontaneous chromosomal mutants which expressed wild-type amounts of the FhuA protein were sequenced. A mutant which was partially T5 sensitive but impaired in all other functions was missing aspartate residue 348 of the mature protein as a result of a three-base deletion. This aspartate residue is part of the hydrophilic sequence Asp-Asp-Glu-Lys. Replacement by site-specific mutagenesis of each of the Asp residues by Tyr, of Glu by Val, and of Lys by Met reduced FhuA activity but less than the Asp deletion did. Ferrichrome inhibited binding of phage phi 80 and of colicin M to these mutants in an allele-specific manner. A completely resistant derivative of the Asp deletion mutant contained, in addition, a leucine-to-proline substitution at position 106 and eight changed bases, converting at positions 576 to 578 an Arg-Pro-Leu sequence to Ala-Arg-Cys. The latter mutations and the Leu-to-Pro replacement alone did not alter sensitivity to the phages but reduced sensitivity to colicin M and albomycin 10- to 1,000-fold. The proline replacements probably disturb FhuA conformation and, in concert with the Asp deletion, inactivate FhuA completely. It is concluded that the Asp deletion site defines a region of FhuA which directly participates in binding of all FhuA ligands. Growth promotion studies on iron-limited media revealed that certain siderophores of the hydroxamate type, such as butylferrichrome, ferrichrysin, and ferrirubin, are taken up not only via FhuA but also via the FhuE outer membrane receptor protein.

This article has been cited by other articles:

• Endriss, F., Braun, V. (2004). Loop Deletions Indicate Regions Important for FhuA Transport and Receptor Functions in Escherichia coli. J. Bacteriol. 186: 4818-4823 [Abstract] [Full Text]  
• Braun, M., Endriss, F., Killmann, H., Braun, V. (2003). In Vivo Reconstitution of the FhuA Transport Protein of Escherichia coli K-12. J. Bacteriol. 185: 5508-5518 [Abstract] [Full Text]  
• Endriss, F., Braun, M., Killmann, H., Braun, V. (2003). Mutant Analysis of the Escherichia coli FhuA Protein Reveals Sites of FhuA Activity. J. Bacteriol. 185: 4683-4692 [Abstract] [Full Text]  
• Killmann, H., Herrmann, C., Torun, A., Jung, G., Braun, V. (2002). TonB of Escherichia coli activates FhuA through interaction with the {beta}-barrel. Microbiology 148: 3497-3509 [Abstract] [Full Text]  
• Howard, S. P., Herrmann, C., Stratilo, C. W., Braun, V. (2001). In Vivo Synthesis of the Periplasmic Domain of TonB Inhibits Transport through the FecA and FhuA Iron Siderophore Transporters of Escherichia coli. J. Bacteriol. 183: 5885-5895 [Abstract] [Full Text]  
• Killmann, H., Braun, M., Herrmann, C., Braun, V. (2001). FhuA Barrel-Cork Hybrids Are Active Transporters and Receptors. J. Bacteriol. 183: 3476-3487 [Abstract] [Full Text]  
• Yeoman, K. H., Wisniewski-Dye, F., Timony, C., Stevens, J. B., deLuca, N. G., Downie, J. A., Johnston, A. W. B. (2000). Analysis of the Rhizobium leguminosarum siderophore-uptake gene fhuA: differential expression in free-living bacteria and nitrogen-fixing bacteroids and distribution of an fhuA pseudogene in different strains. Microbiology 146: 829-837 [Abstract] [Full Text]  
• Ferguson, A. D., Hofmann, E., Coulton, J. W., Diederichs, K., Welte, W. (1998). Siderophore-Mediated Iron Transport: Crystal Structure of FhuA with Bound Lipopolysaccharide. Science 282: 2215-2220 [Abstract] [Full Text]  
• Kilburn, L., Poole, K., Meyer, J.-M., Neshat, S. (1998). Insertion Mutagenesis of the Ferric Pyoverdine Receptor FpvA of Pseudomonas aeruginosa: Identification of Permissive Sites and a Region Important for Ligand Binding. J. Bacteriol. 180: 6753-6756 [Abstract] [Full Text]  
• Berlyn, M. K. B. (1998). Linkage Map of Escherichia coli K-12, Edition 10: The Traditional Map. Microbiol. Mol. Biol. Rev. 62: 814-984 [Abstract] [Full Text]  
• Killmann, H., Herrmann, C., Wolff, H., Braun, V. (1998). Identification of a New Site for Ferrichrome Transport by Comparison of the FhuA Proteins of Escherichia coli, Salmonella paratyphi B, Salmonella typhimurium, and Pantoea agglomerans. J. Bacteriol. 180: 3845-3852 [Abstract] [Full Text]  
• Bös, C., Lorenzen, D., Braun, V. (1998). Specific In Vivo Labeling of Cell Surface-Exposed Protein Loops: Reactive Cysteines in the Predicted Gating Loop Mark a Ferrichrome Binding Site and a Ligand-Induced Conformational Change of the Escherichia coli FhuA Protein. J. Bacteriol. 180: 605-613 [Abstract] [Full Text]  
• Plancon, L., Chami, M., Letellier, L. (1997). Reconstitution of FhuA, an Escherichia coli Outer Membrane Protein, into Liposomes. BINDING OF PHAGE T5 TO FhuA TRIGGERS THE TRANSFER OF DNA INTO THE PROTEOLIPOSOMES. J. Biol. Chem. 272: 16868-16872 [Abstract] [Full Text]