1887

Abstract

FhuA is a multifunctional protein in the outer membrane of that actively transports Fe–ferrichrome and the antibiotics albomycin and rifamycin CGP 4832, and serves as a receptor for the unrelated phages T5, T1, ϕ80 and UC-1, colicin M and microcin J25. The energy source for active transport is the proton-motive force of the cytoplasmic membrane, which is required for all FhuA functions except infection by phage T5, and is thought to be mediated to the outer-membrane receptor FhuA by the TonB protein. The crystal structure of FhuA consists of a β-barrel that is closed by a globular domain. The proximal region carries the TonB box (residues 7–11), for which genetic evidence exists that it interacts with the region around residue 160 of TonB. However, deletion of the TonB box along with the globular domain results in a protein, FhuAΔ5–160, that still displays TonB-dependent active ferrichrome transport across the outer membrane and confers sensitivity to the FhuA ligands. In this study synthetic nonapeptides identical in sequence to amino acids 150–158, 151–159, 152–160, 153–161 and 158–166 of TonB were shown to reduce ferrichrome transport of cells via wild-type FhuA and the corkless derivative FhuAΔ5–160, which suggests that this TonB region is involved in the interaction of TonB with the β-barrel of FhuA. TonB missense mutants reduced the activity of FhuA and FhuAΔ5–160. TonB proteins of different activated FhuA and FhuAΔ5–160 to a similar degree. TonB of displayed low activity in an mutant. Sequencing of the gene of revealed differences from TonB in the region around residue 160 of the deduced protein; these differences might contribute to the lower activity of the TonB protein when coupled to the FhuA protein. The data support the theory that the β-barrel receives the energy from the cytoplasmic membrane via TonB and responds to the energy input and thus represents the transporting domain of FhuA.

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2002-11-01
2019-10-22
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