Wang Yunyun, Shi Qian, Yang Qiulan, Yang Yang, Bian Xin
State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Frontiers Science Center for Cell Responses, Nankai University, Tianjin, China.
Institute of Molecular Medicine, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.
Methods Mol Biol. 2025;2888:221-236. doi: 10.1007/978-1-0716-4318-1_15.
Extended-synaptotagmins (E-Syts) are proteins located on the endoplasmic reticulum (ER) that tether the ER to the plasma membrane (PM) and regulate their lipid homeostasis via its lipid transfer module, the synaptotagmin-like mitochondrial lipid-binding protein (SMP) domain. Here, we describe in vitro DNA nanostructure-assisted lipid transfer assays investigating how the SMP domain transports lipids between membranes and associates with the membranes to extract and release lipids. The lipid transfer signal was detected through fluorescence resonance energy transfer (FRET). This method overcomes the limitations of commonly used lipid transfer assays in accurately controlling inter-liposome distance and liposome size, enabling us to further understand the details involved in the process of SMP domain-mediated lipid transfer. Similar platforms can be extended to studying the lipid transfer distance and membrane curvature sensitivity of other lipid transfer proteins.
延伸突触结合蛋白（E-Syts）是位于内质网（ER）上的蛋白质，它将内质网与质膜（PM）相连，并通过其脂质转移模块——突触结合蛋白样线粒体脂质结合蛋白（SMP）结构域来调节脂质稳态。在此，我们描述了体外DNA纳米结构辅助脂质转移测定法，该方法用于研究SMP结构域如何在膜之间运输脂质以及与膜结合以提取和释放脂质。脂质转移信号通过荧光共振能量转移（FRET）进行检测。该方法克服了常用脂质转移测定法在精确控制脂质体间距离和脂质体大小方面的局限性，使我们能够进一步了解SMP结构域介导的脂质转移过程中的细节。类似的平台可扩展用于研究其他脂质转移蛋白的脂质转移距离和膜曲率敏感性。
