Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan; Theoretical Molecular Science Laboratory, RIKEN, Wako, Saitama 351-0198, Japan.
Structure. 2018 Sep 4;26(9):1284-1296.e4. doi: 10.1016/j.str.2018.05.012. Epub 2018 Jun 28.
Secondary active transporters translocate their substrates using the electrochemical potentials of other chemicals and undergo large-scale conformational changes. Despite extensive structural studies, the atomic details of the transport mechanism still remain elusive. We performed a series of all-atom molecular dynamics simulations of the triose-phosphate/phosphate translocator (TPT), which exports organic phosphates in the chloroplast stroma in strict counter exchange with inorganic phosphate (P). Biased sampling methods, including the string method and umbrella sampling, successfully reproduced the conformational changes between the inward- and outward-facing states, along with the substrate binding. The free energy landscape of this entire TPT transition pathway demonstrated the alternating access and substrate translocation mechanisms, which revealed that P is relayed by positively charged residues along the transition pathway. Furthermore, the conserved Glu207 functions as a "molecular switch", linking the local substrate binding and the global conformational transition. Our results provide atomic-detailed insights into the substrate transport mechanism of the antiporter.
次级主动转运蛋白利用其他化学物质的电化学势来转运其底物，并经历大规模的构象变化。尽管进行了广泛的结构研究，但转运机制的原子细节仍然难以捉摸。我们对三碳糖磷酸/磷酸转运蛋白（TPT）进行了一系列全原子分子动力学模拟，该蛋白在叶绿体基质中以严格的反向交换与无机磷酸盐（P）一起输出有机磷酸盐。有偏采样方法，包括字符串方法和伞形采样，成功地再现了内向和外向状态之间的构象变化，以及底物结合。整个 TPT 跃迁途径的自由能景观展示了交替访问和底物转运机制，揭示了 P 通过沿过渡途径的带正电荷的残基被传递。此外，保守的 Glu207 充当“分子开关”，将局部底物结合与全局构象转变联系起来。我们的结果提供了对反向转运蛋白底物转运机制的原子细节见解。
