The neurotransmitter dopamine has central roles in mood, appetite, arousal and movement¹. Despite its importance in brain physiology and function, and as a target for illicit and therapeutic drugs, the human dopamine transporter (hDAT) and mechanisms by which it is inhibited by small molecules and Zn²⁺ are without a high-resolution structural context. Here we determine the structure of hDAT in a tripartite complex with the competitive inhibitor and cocaine analogue, (–)-2-β-carbomethoxy-3-β-(4-fluorophenyl)tropane² (β-CFT), the non-competitive inhibitor MRS7292³ and Zn²⁺ (ref. ⁴). We show how β-CFT occupies the central site, approximately halfway across the membrane, stabilizing the transporter in an outward-open conformation. MRS7292 binds to a structurally uncharacterized allosteric site, adjacent to the extracellular vestibule, sequestered underneath the extracellular loop 4 (EL4) and adjacent to transmembrane helix 1b (TM1b), acting as a wedge, precluding movement of TM1b and closure of the extracellular gate. A Zn²⁺ ion further stabilizes the outward-facing conformation by coupling EL4 to EL2, TM7 and TM8, thus providing specific insights into how Zn²⁺ restrains the movement of EL4 relative to EL2 and inhibits transport activity.

神经递质多巴胺在情绪、食欲、觉醒和运动中发挥着核心作用¹ 。尽管人类多巴胺转运蛋白 (hDAT) 在大脑生理学和功能中很重要，并且是非法药物和治疗药物的靶标，但其被小分子和 Zn ²⁺抑制的机制尚无高分辨率的结构背景。在这里，我们确定了 hDAT 与竞争性抑制剂和可卡因类似物 (–)-2-β-甲甲氧基-3-β-(4-氟苯基)托烷² (β-CFT)、非竞争性抑制剂的三方复合物的结构。抑制剂 MRS7292 ³和 Zn ^{2 +} （参考文献⁴ ）。我们展示了 β-CFT 如何占据中心位点（大约穿过膜的一半），将转运蛋白稳定在向外开放的构象中。 MRS7292 与结构上未表征的变构位点结合，该位点邻近细胞外前庭，隔离在细胞外环 4 (EL4) 下方并邻近跨膜螺旋 1b (TM1b)，充当楔子，阻止 TM1b 的移动和细胞外门的关闭。 Zn ²⁺离子通过将 EL4 与 EL2、TM7 和 TM8 偶联，进一步稳定外向构象，从而提供有关 Zn ²⁺如何抑制 EL4 相对于 EL2 的运动并抑制转运活性的具体见解。