Department of Drug Design and Pharmacology, University of Copenhagen, Copenhagen, Denmark (J.A., N.S.-H., L.G.Z., K.S., A.S.K.); and Center for Insoluble Structures (inSPIN) and Interdisciplinary Nanoscience Center (iNANO), Department of Chemistry, Aarhus University, Aarhus, Denmark (H.K., B.S.).
Mol Pharmacol. 2014 May;85(5):703-14. doi: 10.1124/mol.113.091249. Epub 2014 Feb 10.
Inhibitors of the serotonin transporter (SERT) are widely used antidepressant agents, but the structural mechanism for inhibitory activity and selectivity over the closely related norepinephrine transporter (NET) is not well understood. Here we use a combination of chemical, biological, and computational methods to decipher the molecular basis for high-affinity recognition in SERT and selectivity over NET for the prototypical antidepressant drug fluoxetine (Prozac; Eli Lilly, Indianapolis, IN). We show that fluoxetine binds within the central substrate site of human SERT, in agreement with recent X-ray crystal structures of LeuBAT, an engineered monoamine-like version of the bacterial amino acid transporter LeuT. However, the binding orientation of fluoxetine is reversed in our experimentally supported model compared with the LeuBAT structures, emphasizing the need for careful experimental verification when extrapolating findings from crystal structures of bacterial transporters to human relatives. We find that the selectivity of fluoxetine and nisoxetine, a NET selective structural congener of fluoxetine, is controlled by residues in different regions of the transporters, indicating a complex mechanism for selective recognition of structurally similar compounds in SERT and NET. Our findings add important new information on the molecular basis for SERT/NET selectivity of antidepressants, and provide the first assessment of the potential of LeuBAT as a model system for antidepressant binding in human transporters, which is essential for future structure-based drug development of antidepressant drugs with fine-tuned transporter selectivity.
5-羟色胺转运体（SERT）抑制剂被广泛用作抗抑郁药，但对于与 SERT 密切相关的去甲肾上腺素转运体（NET）的抑制活性和选择性的结构机制还不太清楚。在这里，我们使用化学、生物学和计算方法的组合来破译 SERT 中高亲和力识别和对典型抗抑郁药氟西汀（百忧解；印第安纳波利斯礼来公司）的选择性的分子基础。我们表明，氟西汀结合在人 SERT 的中央底物结合位点内，这与 LeuBAT 的最近 X 射线晶体结构一致，LeuBAT 是细菌氨基酸转运蛋白 LeuT 的工程单胺样版本。然而，与 LeuBAT 结构相比，我们实验支持的模型中的氟西汀结合取向是相反的，这强调了当从细菌转运蛋白的晶体结构推断出人亲属的发现时，需要仔细进行实验验证。我们发现，氟西汀和尼索西汀（氟西汀的 NET 选择性结构同系物）的选择性由转运体中不同区域的残基控制，这表明在 SERT 和 NET 中对结构相似的化合物进行选择性识别的机制复杂。我们的发现增加了关于 SERT/NET 选择性抗抑郁药的分子基础的重要新信息，并首次评估了 LeuBAT 作为人类转运体中抗抑郁药结合的模型系统的潜力，这对于未来基于结构的具有精细转运体选择性的抗抑郁药药物开发至关重要。
