Wu Zhixiang, Han Zhongjie, Zhou Wenxue, Sun Xiaohan, Chen Lei, Yang Shuang, Hu Jianping, Li Chunhua
Faculty of Environmental and Life Sciences, Beijing University of Technology, Beijing, China.
Key Laboratory of Medicinal and Edible Plants Resources, Development of Sichuan Education Department, School of Pharmacy, Chengdu University, Chengdu, China.
Curr Res Struct Biol. 2022 May 25;4:192-205. doi: 10.1016/j.crstbi.2022.05.005. eCollection 2022.
The human equilibrative nucleoside transporter 1 (hENT1) is an effective controller of adenosine signaling by regulating its extracellular and intracellular concentration, and has become a solid drug target of clinical used adenosine reuptake inhibitors (AdoRIs). Currently, the mechanisms of adenosine transport and inhibition for hENT1 remain unclear, which greatly limits the in-depth understanding of its inner workings as well as the development of novel inhibitors. In this work, the dynamic details of hENT1 underlie adenosine transport and the inhibition mechanism of the non-nucleoside AdoRIs dilazep both were investigated by comparative long-time unbiased molecular dynamics simulations. The calculation results show that the conformational transitions of hENT1 from the outward open to metastable occluded state are mainly driven by TM1, TM2, TM7 and TM9. One of the trimethoxyphenyl rings in dilazep serves as the adenosyl moiety of the endogenous adenosine substrate to competitively occupy the orthosteric site of hENT1. Due to extensive and various interactions with N30, M33, M84, P308 and F334, the other trimethoxyphenyl ring is stuck in the opportunistic site near the extracellular side preventing the complete occlusion of thin gate simultaneously. Obviously, dilazep shows significant inhibitory activity by disrupting the local induce-fit action in substrate binding cavity and blocking the transport cycle of whole protein. This study not only reveals the nucleoside transport mechanism by hENT1 at atomic level, but also provides structural guidance for the subsequent design of novel non-nucleoside AdoRIs with enhanced pharmacologic properties.
人平衡核苷转运体1（hENT1）通过调节细胞外和细胞内腺苷浓度，有效控制腺苷信号传导，已成为临床使用的腺苷再摄取抑制剂（AdoRIs）的可靠药物靶点。目前，hENT1的腺苷转运和抑制机制尚不清楚，这极大地限制了对其内部工作原理的深入理解以及新型抑制剂的开发。在这项工作中，通过比较长时间的无偏分子动力学模拟，研究了hENT1在腺苷转运过程中的动态细节以及非核苷类AdoRIs药物双嘧达莫的抑制机制。计算结果表明，hENT1从外向开放状态到亚稳态封闭状态的构象转变主要由TM1、TM2、TM7和TM9驱动。双嘧达莫中的一个三甲氧基苯环充当内源性腺苷底物的腺苷部分，竞争性占据hENT1的正构位点。由于与N30、M33、M84、P308和F334存在广泛多样的相互作用，另一个三甲氧基苯环卡在细胞外侧附近的机会性位点，同时阻止了细门的完全封闭。显然，双嘧达莫通过破坏底物结合腔中的局部诱导契合作用并阻断整个蛋白质的转运循环，表现出显著的抑制活性。这项研究不仅在原子水平上揭示了hENT1的核苷转运机制，还为后续设计具有增强药理特性的新型非核苷类AdoRIs提供了结构指导。
