Genetically encoded photocrosslinkers locate the high-affinity binding site of antidepressant drugs in the human serotonin transporter.
Genetically encoded photocrosslinkers locate the high-affinity binding site of antidepressant drugs in the human serotonin transporter.
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DOI：
10.1038/ncomms11261
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发表时间：
2016-04-19
影响因子：
16.6
通讯作者：
Strømgaard K
中科院分区：
文献类型：
--
作者：
Rannversson H;Andersen J;Sørensen L;Bang-Andersen B;Park M;Huber T;Sakmar TP;Strømgaard K
尽管人血清素转运体（hSERT）在抑郁症治疗中的作用已经得到了很好的证实，但抗抑郁药物结合的分子细节仍然没有完全了解。在这里，我们利用琥珀密码子抑制膜结合转运蛋白编码光交联非天然氨基酸（UAA）到75个不同的位置在hSERT。UAA以高特异性掺入，并且功能活性转运蛋白与野生型转运蛋白相比具有相似的转运性质和药理学特征。我们采用紫外线诱导的交联与p-叠氮基-L-苯丙氨酸（azF）在hSERT中的选定位置映射丙咪嗪，一种典型的三环类抗抑郁药，和沃替西汀，一种新型的多模式抗抑郁药的结合位点。我们发现，这两种抗抑郁药交联azF纳入在不同的位置内的中央基板结合位点的hSERT，而没有交联观察到在前庭结合位点。综上所述，我们的数据提供了直接的证据，以确定高亲和力的抗抑郁剂的hSERT结合位点。 抗抑郁药物如何与人类血清素转运体结合的分子细节目前尚不清楚。在这里，作者将光交联剂引入蛋白质中，并绘制了几种抗抑郁药的结合位点。
Despite the well-established role of the human serotonin transporter (hSERT) in the treatment of depression, the molecular details of antidepressant drug binding are still not fully understood. Here we utilize amber codon suppression in a membrane-bound transporter protein to encode photocrosslinking unnatural amino acids (UAAs) into 75 different positions in hSERT. UAAs are incorporated with high specificity, and functionally active transporters have similar transport properties and pharmacological profiles compared with wild-type transporters. We employ ultraviolet-induced crosslinking with p-azido-L-phenylalanine (azF) at selected positions in hSERT to map the binding site of imipramine, a prototypical tricyclic antidepressant, and vortioxetine, a novel multimodal antidepressant. We find that the two antidepressants crosslink with azF incorporated at different positions within the central substrate-binding site of hSERT, while no crosslinking is observed at the vestibular-binding site. Taken together, our data provide direct evidence for defining the high-affinity antidepressant binding site in hSERT. Molecular details of how antidepressant drugs bind to the human serotonin transporter are not currently clear. Here, the authors introduce photo-cross-linkers into the protein and map the binding site of several antidepressants.