Structural Mechanism of Soluble N-ethylmaleimide-Sensitive Factor Attachment Protein Receptor Complex Assembly in Lipid Bilayers Revealed by Solid-State NMR
Structural Mechanism of Soluble N-ethylmaleimide-Sensitive Factor Attachment Protein Receptor Complex Assembly in Lipid Bilayers Revealed by Solid-State NMR
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DOI：
10.1021/jacs.3c00764
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发表时间：
2023-05-09
影响因子：
15
通讯作者：
Ma，Cong
中科院分区：
文献类型：
--
作者：
Zhang，Yan;Hu，Yaru;Ma，Cong
突触囊泡融合是由N-乙基马来酰亚胺敏感因子附着蛋白受体(SNARE)蛋白介导的，这些蛋白包括突触素-2(Syb-2)、合成素-1(Syx-1)和SNAP-25。然而，从SNARE基序到跨膜区末端形成完全接触的α螺旋束是否是SNARE介导膜融合所必需的，目前仍存在争议。在这项研究中，我们通过结合基于偶极和标量的固体核磁共振实验表征了syb-2在不同组装状态下的构象。我们的光谱分析揭示了SYB-2TMD的高度动态性质，具有相当大的α螺旋含量。化学位移扰动和突变分析表明，Syb-2的Gly-100残基介导的Syb-2和Syx-1 TMD之间的偶联以及Syb-2 TMD的C末端片段的高迁移率是内膜合并所必需的。我们的结果为SYB-2 TMD在驱动膜融合中的作用提供了新的见解，提高了目前对SNARE复合体组装结构机制的理解。这项研究强调了膜环境对阐明膜蛋白的作用机制的重要意义。
Synaptic vesicle fusion is mediated by solubleN-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins, including synaptobrevin-2 (Syb-2), syntaxin-1 (Syx-1), and SNAP-25. However, it remains controversial whether the formation of thoroughly contacted α-helical bundle from the SNARE motifs to the end of the transmembrane domains (TMDs) is necessary for SNARE-mediated membrane fusion. In this study, we characterized the conformation of Syb-2 in different assembly states using a combination of dipolar- and scalar-based solid-state NMR experiments in lipid bilayers. Our spectral analysis revealed a highly dynamic nature of the Syb-2 TMD with considerable α-helical contents. Chemical shift perturbation and mutational analysis indicated that the coupling between Syb-2 and Syx-1 TMDs mediated by residue Gly-100 of Syb-2 together with high mobility of the C-terminal segment of Syb-2 TMD are required for inner membrane merger. Our results provide new insights into the role of the Syb-2 TMD in driving membrane fusion, which improves the current understanding of the structural mechanism of SNARE complex assembly. This study highlights the significance of membrane environments in elucidating the mechanism of membrane proteins.