TY - JOUR TI - Glutathione binding to the plant AtAtm3 transporter and implications for the conformational coupling of ABC transporters AU - Fan, Chengcheng AU - Rees, Douglas C A2 - Drew, David A2 - Swartz, Kenton J A2 - Drew, David A2 - Seeger, Markus A VL - 11 PY - 2022 DA - 2022/03/25 SP - e76140 C1 - eLife 2022;11:e76140 DO - 10.7554/eLife.76140 UR - https://doi.org/10.7554/eLife.76140 AB - The ATP binding cassette (ABC) transporter of mitochondria (Atm) from Arabidopsis thaliana (AtAtm3) has been implicated in the maturation of cytosolic iron-sulfur proteins and heavy metal detoxification, plausibly by exporting glutathione derivatives. Using single-particle cryo-electron microscopy, we have determined four structures of AtAtm3 in three different conformational states: two inward-facing conformations (with and without bound oxidized glutathione [GSSG]), together with closed and outward-facing states stabilized by MgADP-VO4. These structures not only provide a structural framework for defining the alternating access transport cycle, but also reveal the paucity of cysteine residues in the glutathione binding site that could potentially form inhibitory mixed disulfides with GSSG. Despite extensive efforts, we were unable to prepare the ternary complex of AtAtm3 containing both GSSG and MgATP. A survey of structurally characterized type IV ABC transporters that includes AtAtm3 establishes that while nucleotides are found associated with all conformational states, they are effectively required to stabilize occluded, closed, and outward-facing conformations. In contrast, transport substrates have only been observed associated with inward-facing conformations. The absence of structures with dimerized nucleotide binding domains containing both nucleotide and transport substrate suggests that this form of the ternary complex exists only transiently during the transport cycle. KW - ABC transporters KW - glutathione transport KW - alternating access JF - eLife SN - 2050-084X PB - eLife Sciences Publications, Ltd ER -