Transmembrane topology of the two FhuB domains representing the hydrophobic components of bacterial ABC transporters involved in the uptake of siderophores, haem and vitamin B12.

Transport of siderophores of the hydroxamate type across the Escherichia coli cytoplasmic membrane depends on a periplasmic binding-protein-dependent (PBT) system. This uptake system consists of the binding protein FhuD, the membrane-associated putative ATP-hydrolase FhuC and the integral membrane protein FhuB. The two halves of FhuB [FhuB(N) and FhuB(C)], both essential for transport, are similar with respect to structure and function. Regions were identified in FhuB(N) and FhuB(C) which are presumably involved in the interaction of the two FhuB halves with each other or with other components of the uptake system, or with the different substrates. To determine the topology of the membrane-embedded polypeptide chain, FhuB'-'beta-lactamase protein fusions were analysed. The experimental data suggest that each half of the FhuB is able to fold autonomously into the lipid bilayer, which is a prerequisite for the assembly of both halves into a transport-competent formation. The hydrophobic components from PBT systems involved in the uptake of siderophores, haem and vitamin B12 define a subclass of polytopic integral membrane proteins. The topology of these 'siderophore family' proteins differs from that of the equivalent components of other PBT systems in that each polypeptide (and each half of FhuB) consists of 10 membrane-spanning regions, with the N- and C-termini located in the cytoplasm. The conserved region at a distance of about 90 amino acids from the C-terminus, typical of all hydrophobic PBT proteins, is also oriented to the cytoplasm. However, in the 'siderophore family' proteins this putative ATPase interaction loop is followed by four instead of two transmembrane spans.