, 1999). Membrane topology of Chr3N and Chr3C is antiparallel. The C-terminal end of Chr3N is located in the cytoplasm, whereas the C terminus of Chr3C lies in the periplasm (Fig. 1b and d). Jiménez-Mejía et al. (2006) reported a 13-TMS topology for P. aeruginosa ChrA protein, a member of the long-chain CHR family of the CHR superfamily. The two homologous halves of ChrA,

formed by six TMSs each, displayed antiparallel membrane topology between them. It was proposed that this structure arose from the duplication of an equally oriented six-TMS ancestral protein domain followed by insertion of a central TMS (TMS7); this insertion might have caused the repeated domains to adopt the opposite orientation in a native parallel structure (Jiménez-Mejía check details et al., 2006). Topologic inversion of halves of membrane proteins has been widely reported and is considered a common evolutionary process for these polypeptides (Ichihara et al., 2004;

Rapp et al., 2006). It was proposed that membrane proteins with two antiparallel domains arose from ancestral monodomain proteins with dual topology (Rapp et al., 2006), that is, proteins that may insert into the membrane in either orientation (a ‘flip-flopping’ protein; Bowie, 2006). This dual topology ancestor may form homodimers displaying opposite orientation in the membrane. Gene duplication followed by sequence divergence would result in heterodimeric proteins with subunits of fixed but opposite orientation. Experimental evidence supporting this evolutionary Selleck Gefitinib pathway has been obtained from the analysis of proteins of the small multidrug resistance (SMR) family (reviewed in Bay et al., 2008). Antiparallel

arrangement of E. coli homodimeric EmrE transporter has been widely reported (see Chen et al., 2007), although a parallel structure oxyclozanide has also been claimed (Steiner-Mordoch et al., 2008). Another SMR family member, the EbrAB protein pair, has also been assigned antiparallel membrane topology (Kikukawa et al., 2007). Closely homologous proteins RnfA and RnfE from E. coli (Saaf et al., 1999) and NqrD and NqrE from Vibrio cholerae (Duffy & Barquera, 2006), both pairs being NADH-oxidoreductases constituted by six-TMS monomers, showed a completely opposite membrane topology. Members of several 10-TMS transporter families are also constituted by 2 five-TMS repeat units arranged in opposite membrane orientations (Saier, 2003; Lolkema et al., 2005). Aquaporins (Murata et al., 2000), ClC chloride channels (Dutzler et al., 2002), AmtB ammonia transporters (Khademi et al., 2004), and members of the DUF606 family of bacterial transporters (Lolkema et al., 2008) are all additional examples of proteins composed of two repeated halves with opposite membrane orientations. Indeed, the antiparallel domain organization is observed more frequently in the 3D structures of membrane proteins than the parallel domain organization (Lolkema et al., 2008).