Do Hoa Quynh, Pirayesh Elham, Ferreira Garren, Pandhare Akash, Gallardo Zackary Ryan, Jansen Michaela
Cell Physiology and Molecular Biophysics, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, Texas.
Cell Physiology and Molecular Biophysics, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, Texas.
Biophys J. 2024 Jul 16;123(14):2185-2198. doi: 10.1016/j.bpj.2024.04.027. Epub 2024 Apr 27.
Bupropion is an atypical antidepressant and smoking cessation drug that causes adverse effects such as insomnia, irritability, and anxiety. Bupropion inhibits dopamine and norepinephrine reuptake transporters and eukaryotic cation-conducting pentameric ligand-gated ion channels, such as nicotinic acetylcholine and serotonin type 3A receptors, at clinically relevant concentrations. Here, we demonstrate that bupropion also inhibits a prokaryotic homolog of pentameric ligand-gated ion channels, the Gloeobacter violaceus ligand-gated ion channel (GLIC). Using the GLIC as a model, we used molecular docking to predict binding sites for bupropion. Bupropion was found to bind to several sites within the transmembrane domain, with the predominant site being localized to the interface between transmembrane segments M1 and M3 of two adjacent subunits. Residues W213, T214, and W217 in the first transmembrane segment, M1, and F267 and I271 in the third transmembrane segment, M3, most frequently reside within a 4 Å distance from bupropion. We then used single amino acid substitutions at these positions and two-electrode voltage-clamp recordings to determine their impact on bupropion inhibitory effects. The substitution T214F alters bupropion potency by shifting the half-maximal inhibitory concentration to a 13-fold higher value compared to wild-type GLIC. Residue T214 is found within a previously identified binding pocket for neurosteroids and lipids in the GLIC. This intersubunit binding pocket is structurally conserved and almost identical to a binding pocket described for neurosteroids in γ-aminobutyric acid type A receptors. Our data thus suggest that the T214 that lines a previously identified lipophilic binding pocket in GLIC and γ-aminobutyric acid type A receptors is also a modulatory site for bupropion interaction with the GLIC.
安非他酮是一种非典型抗抑郁药和戒烟药物，会引起失眠、易怒和焦虑等不良反应。在临床相关浓度下，安非他酮会抑制多巴胺和去甲肾上腺素再摄取转运体以及真核生物阳离子传导五聚体配体门控离子通道，如烟碱型乙酰胆碱受体和5-羟色胺3A型受体。在此，我们证明安非他酮还能抑制五聚体配体门控离子通道的原核同源物——紫球藻属配体门控离子通道（GLIC）。以GLIC为模型，我们利用分子对接预测安非他酮的结合位点。结果发现安非他酮与跨膜结构域内的多个位点结合，主要位点位于两个相邻亚基的跨膜片段M1和M3之间的界面处。第一个跨膜片段M1中的W213、T214和W217残基以及第三个跨膜片段M3中的F267和I271残基，最常位于距安非他酮4埃的距离内。然后，我们在这些位置进行单氨基酸替换，并通过双电极电压钳记录来确定它们对安非他酮抑制作用的影响。与野生型GLIC相比，T214F替换通过将半数最大抑制浓度提高到13倍，改变了安非他酮的效力。在GLIC中，T214残基位于先前确定的神经甾体和脂质结合口袋内。这个亚基间结合口袋在结构上是保守的，与γ-氨基丁酸A型受体中描述的神经甾体结合口袋几乎相同。因此，我们的数据表明，在GLIC和γ-氨基丁酸A型受体中构成先前确定的亲脂性结合口袋的T214也是安非他酮与GLIC相互作用的调节位点。
