Structures of MmpL complexes reveal the assembly and mechanism of this family of transporters

Abstract

We co-expressed the MmpL5 transporter and MmpS5 adaptor proteins in Mycobacterium smegmatis and defined their structures from these detergent-solubilized crude membranes. Data generated from these samples allowed us to simultaneously solve three distinct classes of membrane protein complexes to high resolutions. We observed that MmpL5 presents as a monomer in complex with the cytosolic meromycolate extension acyl carrier protein M (AcpM) in a molar ratio of 1:1, where these AcpM-MmpL5 complexes closely pack together to generate regular two-dimensional arrays. We also identified MmpL5 as a trimer that interacts with MmpS5 and AcpM in a molar ratio of 3:3:3 to assemble the tripartite complex AcpM-MmpL5-MmpS5 that spans both the inner and outer membranes of the mycobacterium. In addition, we discovered that MmpL5 and AcpM are able to form the trimeric AcpM-MmpL5 complex in a molar ratio of 3:3. The structural data reveal that the full-length MmpL5 trimer is capable of spanning the entire mycobacterial cell envelope to transport substrates. However, this assembly requires the presence of MmpS5 to stabilize secondary structural features of the MmpL5 periplasmic subdomains.