Conserved amino acids in the N- and C-terminal domains of integral membrane transporter FhuB define sites important for intra- and intermolecular interactions

Transport of iron(III) hydroxamates across the inner membrane of Escherichia coli is mediated by a periplasmic binding protein-dependent transport (PBT) mechanism. FhuB, the integral membrane component of the system, is composed of covalently linked halves (FhuB[N] and FhuB[C]) which still function when present as two distinct polypeptide chains. Our analysis of two uptake-deficient FhuB derivatives provides evidence for a mechanistically novel type of functional complementation: 'domain displacement' in the cytoplasmic membrane. Amino acid residues 60 and 426 in the FhuB polypeptide chain may define key positions that are important for FhuB[N]-FhuB[C] interaction. Furthermore, FhuB derivatives, altered in either one of their conserved regions - typical of PBT related integral membrane proteins - displayed a dominant negative effect on ferric hydroxamate transport. The experimental data suggest that the two functionally equivalent conserved regions in FhuB[N] and fhuB[C] are primarily involved in the interaction with another component of the transport system, probably FhuC.