Molecular basis for substrate transport of Mycobacterium tuberculosis ABC importer DppABCD.

The type I adenosine 5'-triphosphate (ATP)-binding cassette (ABC) transporter DppABCD is believed to be responsible for the import of exogenous heme as an iron source into the cytoplasm of the human pathogen Mycobacterium tuberculosis (Mtb). Additionally, this system is also known to be involved in the acquisition of tri- or tetra-peptides. Here, we report the cryo-electron microscopy structures of the dual-function Mtb DppABCD transporter in three forms, namely, the apo, substrate-bound, and ATP-bound states. The apo structure reveals an unexpected and previously uncharacterized assembly mode for ABC importers, where the lipoprotein DppA, a cluster C substrate-binding protein (SBP), stands upright on the translocator DppBCD primarily through its hinge region and N-lobe. These structural data, along with biochemical studies, reveal the assembly of DppABCD complex and the detailed mechanism of DppABCD-mediated transport. Together, these findings provide a molecular roadmap for understanding the transport mechanism of a cluster C SBP and its translocator.

Cryo-EM structures for the Mycobacterium tuberculosis iron-loaded siderophore transporter IrtAB.

The adenosine 5'-triphosphate (ATP)-binding cassette (ABC) transporter, IrtAB, plays a vital role in the replication and viability of Mycobacterium tuberculosis (Mtb), where its function is to import iron-loaded siderophores. Unusually, it adopts the canonical type IV exporter fold. Herein, we report the structure of unliganded Mtb IrtAB and its structure in complex with ATP, ADP, or ATP analogue (AMP-PNP) at resolutions ranging from 2.8 to 3.5 Å. The structure of IrtAB bound ATP-Mg2+ shows a "head-to-tail" dimer of nucleotide-binding domains (NBDs), a closed amphipathic cavity within the transmembrane domains (TMDs), and a metal ion liganded to three histidine residues of IrtA in the cavity. Cryo-electron microscopy (Cryo-EM) structures and ATP hydrolysis assays show that the NBD of IrtA has a higher affinity for nucleotides and increased ATPase activity compared with IrtB. Moreover, the metal ion located in the TM region of IrtA is critical for the stabilization of the conformation of IrtAB during the transport cycle. This study provides a structural basis to explain the ATP-driven conformational changes that occur in IrtAB.