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

Abstract

Transport of iron(III) hydroxamates across the inner membrane of Escherichia coli is mediated by a peri-plasmic 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 cyto-plasmic 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.