SEROTONIN AND GABA TRANSPORTERS--STRUCTURE/FUNCTION

这项建议的主要目的是了解哺乳动物5-羟色胺转运体摄取5-羟色胺的分子机制。5-羟色胺转运体利用Na+电化学势将6种神经递质逆其浓度梯度转运到细胞内，从而维持较低的胞外神经递质浓度。这一过程背后的分子机制尚不清楚。拟议的一系列实验将有助于建立转运蛋白功能的综合图景，并确定参与运输周期特定步骤的结构域。根据这项研究的工作假设，转运蛋白包含一个类似通道的管腔，两侧有细胞外和细胞内的门。这些门的打开和关闭受有机底物和管腔内离子的结合控制。神经递质转运和离子电化学势之间的能量耦合是门的顺序开启和关闭的结果，也是神经递质结合依赖于共转运离子结合的结果。拟议的研究将利用电生理学和其他功能测量来表征突变的转运蛋白，并识别和研究与运输过程的特定方面有关的功能结构域。我们将研究蛋白质中的结构域：(1)参与转运体门控，(2)形成转运体管腔，(3)形成并积极参与离子和5-羟色胺结合部位。研究表明，人类5-羟色胺转运体活性的改变与几种精神障碍有关，可能与酒精和可卡因滥用有关。转运蛋白也是氟西汀、哌甲酸甲酯和苯丙胺等治疗药物的主要靶点。要了解5-羟色胺转运体在正常脑功能、精神障碍和药物滥用中的作用，需要更多地了解转运体功能的生物物理和分子机制。

The principal objective of this proposal is to understand the molecular mechanism of serotonin uptake by the mammalian serotonin transporter. The serotonin transporter utilizes the Na+ electrochemical potential to transpor6t neurotransmitters into the cell against their concentration gradient, so that a low extracellular neurotransmitter concentration can be maintained. The molecular mechanism that underlies this process is not known. The proposed set of experiments will help to establish an integrated picture of transporter function and identify structural domains that participate in specific steps of the transport cycle. According to this study's working hypothesis, the transporter protein contains a channel-like lumen flanked by extracellular and intracellular gates. The opening and closing of these gates is governed by the binding of organic substrate and ions in the lumen. The energetic coupling between neurotransmitter transport and the electrochemical potential of ions is a result of the sequential opening and closing of the gates as well as the dependance of neurotransmitter binding on the binding of co-transporting ions. The proposed research will utilize electrophysiology and other functional measurements in order to characterize mutated transporters, and to identify and study functional domains that contribute to specific aspects of the transport process. We will investigate domains in the protein that (1) participate in transporter gating, (2) form the transporter lumen, and (3) form and are actively involved with ion and serotonin binding sites. Studies have shown that an alterations in the activity of the human serotonin transporter is associated with several mental disorders and possibly with alcohol and cocaine abuse. The transporter is also a major target for therapeutic drugs such as fluoxetine, methylphenidate, and amphetamine. Understanding the roles of the serotonin transporter in normal brain function, mental disorders, and drug abuse requires more insight into the biophysical and molecular mechanisms of transporter function.