title: Active transport of an antibiotic rifamycin derivative by the outer-membrane protein FhuA.
abstract: BACKGROUND
  FhuA, an integral membrane protein of Escherichia coli, actively transports ferrichrome and the structurally related antibiotic albomycin across the outer membrane. The transport is coupled to the proton motive force, which energizes FhuA through the inner-membrane protein TonB. FhuA also transports the semisynthetic rifamycin derivative CGP 4832, although the chemical structure of this antibiotic differs markedly from that of ferric hydroxamates.
  
  
  RESULTS
  X-ray crystallography revealed that rifamycin CGP 4832 occupies the same ligand binding site as ferrichrome and albomycin, thus demonstrating a surprising lack of selectivity. However, the binding of rifamycin CGP 4832 is deviant from the complexes of FhuA with hydroxamate-type ligands in that it does not result in the unwinding of the switch helix but only in its destabilization, as reflected by increased B factors. Unwinding of the switch helix is proposed to be required for efficient binding of TonB to FhuA and for coupling the proton motive force of the cytoplasmic membrane with energy-dependent ligand transport. The transport data from cells expressing mutant FhuA proteins indicated conserved structural and mechanistic requirements for the transport of both types of compounds.
  
  
  CONCLUSIONS
  We conclude that the binding of rifamycin CGP 4832 destabilizes the switch helix and promotes the formation of a transport-competent FhuA-TonB complex, albeit with lower efficiency than ferrichrome. Active transport of this rifamycin derivative explains the 200-fold increase in potency as compared to rifamycin, which is not a FhuA-specific ligand and permeates across the cell envelope by passive diffusion only.
authors.affiliation: Fakultät für Biologie, Universität Konstanz, D-78457, Konstanz, Germany.
authors.forename: A D
authors.initials: AD
authors.lastname: Ferguson
authors.forename: J
authors.initials: J
authors.lastname: Ködding
authors.forename: G
authors.initials: G
authors.lastname: Walker
authors.forename: C
authors.initials: C
authors.lastname: Bös
authors.forename: J W
authors.initials: JW
authors.lastname: Coulton
authors.forename: K
authors.initials: K
authors.lastname: Diederichs
authors.forename: V
authors.initials: V
authors.lastname: Braun
authors.forename: W
authors.initials: W
authors.lastname: Welte
mesh_terms: D000495:Allosteric Site; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry; D001665:Binding Sites; D001692:Biological Transport; D001693:Biological Transport, Active; D002462:Cell Membrane / Q000378:metabolism; D018360:Crystallography, X-Ray; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000737:chemistry; D029968:Escherichia coli Proteins / Q000737:chemistry*; D005291:Ferrichrome / Q000737:chemistry; D008024:Ligands; D008565:Membrane Proteins / Q000737:chemistry; D008958:Models, Molecular; D009154:Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011991:Receptors, Virus / Q000737:chemistry*; D012294:Rifamycins / Q000737:chemistry* / Q000494:pharmacology*; D013050:Spectrometry, Fluorescence; D013997:Time Factors; D014364:Tryptophan / Q000737:chemistry
chemical_list: D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C050900:FhuA protein, E coli; D008024:Ligands; D008565:Membrane Proteins; D011991:Receptors, Virus; D012294:Rifamycins; C057616:tonB protein, Bacteria; C489181:tonB protein, E coli; D005291:Ferrichrome; D014364:Tryptophan