Abstract
The human Alanine Serine Cysteine Transporter 2 (ASCT2) is a neutral amino acid exchanger that belongs to the solute carrier family 1 (SLC1A). SLC1A structures have revealed an elevator-type mechanism, in which the substrate is translocated across the cell membrane by a large displacement of the transport domain, whereas a small movement of hairpin 2 (HP2) gates the extracellular access to the substrate-binding site. However, it has remained unclear how substrate binding and release is gated on the cytoplasmic side. Here, we present an inward-open structure of the human ASCT2, revealing a hitherto elusive SLC1A conformation. Strikingly, the same structural element (HP2) serves as a gate in the inward-facing as in the outward-facing state. The structures reveal that SLC1A transporters work as one-gate elevators. Unassigned densities near the gate and surrounding the scaffold domain, may represent potential allosteric binding sites, which could guide the design of lipidic-inhibitors for anticancer therapy.
Highlights
The human Alanine Serine Cysteine Transporter 2 (ASCT2) is a neutral amino acid exchanger that belongs to the solute carrier family 1 (SLC1A)
We reckoned that binding of bulkier substrates might prevent full occlusion and capture the protein in an inward-open state—a strategy previously used to obtain the outward-open structures of GltPh and EAAT1, where binding of DL-threo-β-benzyloxyaspartic acid (DL-TBOA) and TFB-TBOA prevented closure of HP215,19
It appears that the arginine residue is the main determinant for acidic amino acid specificity, as the guanidium group interacts with the side chain carboxylate of glutamate and aspartate in transporters as GltPh, GltTk and EAAT1
Summary
The human Alanine Serine Cysteine Transporter 2 (ASCT2) is a neutral amino acid exchanger that belongs to the solute carrier family 1 (SLC1A). SLC1A structures have revealed an elevator-type mechanism, in which the substrate is translocated across the cell membrane by a large displacement of the transport domain, whereas a small movement of hairpin 2 (HP2) gates the extracellular access to the substrate-binding site. It has remained unclear how substrate binding and release is gated on the cytoplasmic side. Structures of the prokaryotic SLC1A transporters GltPh and GltTk and the human EAAT1 and ASCT2, in different conformational states, have provided insights into the transport cycle[14,15,16,17,18,19] They all share a similar homotrimeric protein architecture, where each protomer consists of a scaffold and a transport domain. All structures capture an inward-open state, where HP2 has moved indicating a one-gate elevator mechanism
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