Structure of the periplasmic adaptor protein from a major facilitator superfamily (MFS) multidrug efflux pump

•Periplasmic adaptors are key to MFS-dependent tripartite pump efflux.•We present the structure of Aquifex aeolicus EmrA, an MFS pump adaptor.•The adaptor has an extended 127Å long α-helical coiled-coil and disordered β-barrel loop.•The inner membrane transporter interacting membrane proximal domain is absent in EmrA.•This extends the modular view of adaptors, mediating diverse pump assembly. Periplasmic adaptor proteins are key components of bacterial tripartite efflux pumps. The 2.85Å resolution structure of an MFS (major facilitator superfamily) pump adaptor, Aquifex aeolicus EmrA, shows linearly arranged α-helical coiled-coil, lipoyl, and β-barrel domains, but lacks the fourth membrane-proximal domain shown in other pumps to interact with the inner membrane transporter. The adaptor α-hairpin, which binds outer membrane TolC, is exceptionally long at 127Å, and the β-barrel contains a conserved disordered loop. The structure extends the view of adaptors as flexible, modular components that mediate diverse pump assembly, and suggests that in MFS tripartite pumps a hexamer of adaptors could provide a periplasmic seal.