PT - JOURNAL ARTICLE AU - Qiu, Biao AU - Boudker, Olga TI - Dancing with the ions: symport and antiport mechanisms of human glutamate transporters AID - 10.1101/2022.08.12.503774 DP - 2022 Jan 01 TA - bioRxiv PG - 2022.08.12.503774 4099 - http://biorxiv.org/content/early/2022/08/13/2022.08.12.503774.short 4100 - http://biorxiv.org/content/early/2022/08/13/2022.08.12.503774.full AB - Excitatory amino acid transporters (EAATs) pump glutamate into glial cells and neurons. EAATs achieve million-fold transmitter gradients by symporting it with three sodium ions and a proton and counter-transporting a potassium ion via an elevator mechanism. Despite the availability of structures, the symport and antiport mechanisms remain unclear. We report high-resolution Cryo-EM structures of human EAAT3 bound to the neurotransmitter glutamate with symported ions, potassium ions, sodium ions alone, or in the absence of ligands. We show that an evolutionarily conserved occluded translocation intermediate has a dramatically higher affinity for the neurotransmitter and the counter-transported potassium ion than outward- or inward-facing transporters and plays a crucial role in ion coupling. We propose a comprehensive ion coupling mechanism involving a choreographed interplay between bound solutes, conformations of conserved amino acid motifs, and movements of the gating hairpin and the substrate-binding domain.Competing Interest StatementThe authors have declared no competing interest.