Structural insights into the human neutral amino acid transporter ASCT2
Living cells maintain homeostasis by constantly exchanging metabolites, ions, and energy with their environment. Central to this process are membrane transporters, which regulate solute movement across membranes. Among them, amino acid transporters play a vital role in protein synthesis, signaling, and metabolism, requiring precise intracellular regulation.
This PhD thesis investigates ASCT2 (SLC1A5), a Na⁺-dependent neutral amino acid harmonizer. ASCT2 facilitates obligatory exchange of small polar amino acids, including glutamine, and is implicated in nutrient homeostasis, cancer metabolism, and cell-cell fusion. Despite structural similarity to concentrative EAAT transporters, ASCT2 is confined to exchange-only transport.
Chapter 2 reveals that ASCT2 binds three Na⁺ ions per transport cycle but retains one Na⁺ ion even under low-Na⁺ conditions. This constitutive binding, together with restricted dynamics in transmembrane helix 7, prevents full cargo release and explains its strict exchange mechanism.
Chapter 3 shows that the membrane environment shapes ASCT2 conformational flexibility: detergent solubilization stabilizes a single state, whereas nanodiscs capture heterogeneous and intermediate conformations, pinpointing the structural checkpoints against Na⁺ leakage during transport cycle.
Chapter 4 explores ASCT2’s role as a cellular receptor for a fusogenic human protein Syncytin-1. The cryo-EM structure of ASCT2 in complex with Syncytin-1 revealed selective recognition of the outward-facing state, mediated by a broad, asymmetric interaction interface.
Finally, Chapter 5 demonstrates the use of the fluorescent amino acid Anap as a probe for ASCT2 dynamics. Altogether, this work integrates structural and biochemical approaches to uncover how ASCT2 balances dual roles in transport and membrane fusion, advancing understanding of transporter biology.