Department of Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign, Urbana, Illinois.
Department of Biochemistry, University of Illinois at Urbana-Champaign, Urbana, Illinois.
Biophys J. 2022 Mar 1;121(5):715-730. doi: 10.1016/j.bpj.2022.01.024. Epub 2022 Feb 1.
The serotonin transporter (SERT) initiates the reuptake of extracellular serotonin in the synapse to terminate neurotransmission. The cryogenic electron microscopy structures of SERT bound to ibogaine and the physiological substrate serotonin resolved in different states have provided a glimpse of the functional conformations at atomistic resolution. However, the conformational dynamics and structural transitions to intermediate states are not fully understood. Furthermore, the molecular basis of how serotonin is recognized and transported remains unclear. In this study, we performed unbiased microsecond-long simulations of the human SERT to investigate the structural dynamics to various intermediate states and elucidated the complete substrate import pathway. Using Markov state models, we characterized a sequential order of conformational-driven ion-coupled substrate binding and transport events and calculated the free energy barriers of conformation transitions associated with the import mechanism. We find that the transition from the occluded to inward-facing state is the rate-limiting step for substrate import and that the substrate decreases the free energy barriers to achieve the inward-facing state. Our study provides insights on the molecular basis of dynamics-driven ion-substrate recognition and transport of SERT that can serve as a model for other closely related neurotransmitter transporters.
血清素转运体（SERT）启动突触中外源性血清素的再摄取，以终止神经递质传递。与伊博加因和生理底物血清素结合的 SERT 的低温电子显微镜结构已在不同状态下得到解决，为原子分辨率的功能构象提供了一些了解。然而，对中间状态的构象动力学和结构转变还不完全了解。此外，血清素如何被识别和转运的分子基础仍不清楚。在这项研究中，我们对人 SERT 进行了无偏的微秒级模拟，以研究各种中间状态的结构动力学，并阐明了完整的底物导入途径。使用马尔可夫状态模型，我们描述了构象驱动的离子偶联底物结合和运输事件的顺序，并计算了与导入机制相关的构象转变的自由能障碍。我们发现，从封闭状态到内向状态的转变是底物导入的限速步骤，而底物降低了实现内向状态的自由能障碍。我们的研究提供了关于 SERT 的动力学驱动的离子-底物识别和转运的分子基础的见解，可作为其他密切相关的神经递质转运蛋白的模型。
