The high-affinity binding site for tricyclic antidepressants resides in the outer vestibule of the serotonin transporter.
The high-affinity binding site for tricyclic antidepressants resides in the outer vestibule of the serotonin transporter.
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DOI：
10.1124/mol.110.067538
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发表时间：
2010-12
影响因子：
3.6
通讯作者：
Sucic S
中科院分区：
文献类型：
--
作者：
Sarker S;Weissensteiner R;Steiner I;Sitte HH;Ecker GF;Freissmuth M;Sucic S
Aquifex aeolicus (LeuTAa) 的细菌亮氨酸转运蛋白的结构已被用作哺乳动物 Na+/Cl− 依赖性转运蛋白，特别是血清素转运蛋白 (SERT) 的模型。与三环类抗抑郁药配体的 LeuTAa 的晶体结构预示着抑制剂和底物的同时结合。这与 SERT 底物和抑制剂的相互竞争性抑制是不相容的。我们通过同源建模和对接研究探索了三环类抗抑郁药的结合模式。随后使用两种方法来区分三簇潜在的对接姿势：1）诊断性SERTY95F突变，该突变大大降低了对[3H]丙咪嗪的亲和力，但不影响底物结合； 2) 在存在和不存在卡马西平（即具有短侧链的三环丙咪嗪类似物，与[3H]丙咪嗪竞争结合SERT）的情况下的竞争结合实验。释放剂（对氯苯丙胺、亚甲基二氧基甲基苯丙胺/摇头丸）和卡马西平的结合是相互排斥的，但在卡马西平存在的情况下生成的迪克森图产生了血清素、MPP+、帕罗西汀和伊博加因的相交线。这些观察结果与模型一致，其中1）三环对接至外前庭，二甲基氨基丙基侧链指向底物结合位点； 2) 安非他明的结合会导致内前庭和外前庭发生结构变化，从而阻止三环的对接； 3）伊博加因（与面向内的构象结合）和卡马西平的同时结合表明内前庭中存在第二个结合位点，与单胺转运蛋白的假对称折叠一致。这可能是抗抑郁药的第二个低亲和力结合位点。
The structure of the bacterial leucine transporter from Aquifex aeolicus (LeuTAa) has been used as a model for mammalian Na+/Cl−-dependent transporters, in particular the serotonin transporter (SERT). The crystal structure of LeuTAa liganded to tricyclic antidepressants predicts simultaneous binding of inhibitor and substrate. This is incompatible with the mutually competitive inhibition of substrates and inhibitors of SERT. We explored the binding modes of tricyclic antidepressants by homology modeling and docking studies. Two approaches were used subsequently to differentiate between three clusters of potential docking poses: 1) a diagnostic SERTY95F mutation, which greatly reduced the affinity for [3H]imipramine but did not affect substrate binding; 2) competition binding experiments in the presence and absence of carbamazepine (i.e., a tricyclic imipramine analog with a short side chain that competes with [3H]imipramine binding to SERT). Binding of releasers (para-chloroamphetamine, methylene-dioxy-methamphetamine/ecstasy) and of carbamazepine were mutually exclusive, but Dixon plots generated in the presence of carbamazepine yielded intersecting lines for serotonin, MPP+, paroxetine, and ibogaine. These observations are consistent with a model, in which 1) the tricyclic ring is docked into the outer vestibule and the dimethyl-aminopropyl side chain points to the substrate binding site; 2) binding of amphetamines creates a structural change in the inner and outer vestibule that precludes docking of the tricyclic ring; 3) simultaneous binding of ibogaine (which binds to the inward-facing conformation) and of carbamazepine is indicative of a second binding site in the inner vestibule, consistent with the pseudosymmetric fold of monoamine transporters. This may be the second low-affinity binding site for antidepressants.