Zhang Xintong, Xu Yanhong, Chen Qingyang, Li Chan, Zhang Yuan-Wei
School of Life Sciences, Guangzhou University, Guangzhou 510006, China.
Int J Mol Sci. 2025 Mar 26;26(7):3054. doi: 10.3390/ijms26073054.
The neurotransmitter sodium symporters (NSSs) play critical roles in the neurotransmission of monoamine and amino acid neurotransmitters and are the molecular targets of therapeutic agents in the treatment of several psychiatric disorders. Despite significant progress in characterizing structures and transport mechanisms, the management of conformational transitions by structural elements coupled with ion and substrate binding remains to be fully understood. In the present study, we biochemically identified a conserved GXP motif in the fifth transmembrane domain (TM5) of the serotonin transporter (SERT) that plays a vital role in its transport function by facilitating conformational transitions. Mutations of the conserved Gly278 or Pro288 in the GXP motif dramatically decreased specific transport activity by reducing the substrate binding-induced conformational transitions from an outward-open to an inward-open conformation. In addition, cysteine accessibility measurements demonstrated that the unwinding of the intracellular part of TM5 occurs during conformational transitions from an outward-open state, through an occluded state, to an inward-open state and that substrate binding triggers TM5 unwinding. Furthermore, mutations of the GXP motif were shown to result in destructive effects on TM5 unwinding, suggesting that the GXP motif controls conformational transitions through TM5 unwinding. Taken together, the present study provides new insights into the structural elements controlling conformational transitions in NSS transporters.
神经递质钠共转运体（NSSs）在单胺和氨基酸神经递质的神经传递中发挥着关键作用，并且是治疗多种精神疾病的治疗药物的分子靶点。尽管在表征结构和转运机制方面取得了重大进展，但与离子和底物结合相关的结构元件对构象转变的调控仍有待充分了解。在本研究中，我们通过生化方法在血清素转运体（SERT）的第五跨膜结构域（TM5）中鉴定出一个保守的GXP基序，该基序通过促进构象转变在其转运功能中发挥至关重要的作用。GXP基序中保守的甘氨酸278或脯氨酸288的突变通过减少底物结合诱导的从外向开放构象到内向开放构象的构象转变，显著降低了特异性转运活性。此外，半胱氨酸可及性测量表明，TM5细胞内部分的解旋发生在从外向开放状态通过闭塞状态到内向开放状态的构象转变过程中，并且底物结合触发TM5解旋。此外，GXP基序的突变被证明对TM5解旋有破坏作用，这表明GXP基序通过TM5解旋控制构象转变。综上所述，本研究为控制NSS转运体构象转变的结构元件提供了新的见解。
