Cheppali Sudheer K, Li Chang, Xing Wenjing, Sun Ruirui, Yang Mengyi, Xue Yi, Lu Si-Yao, Yao Jun, Sun Shan, Chen Chunlai, Sui Sen-Fang
State Key Laboratory of Membrane Biology, Beijing Frontier Research Center of Biological Structure, Beijing Advanced Innovation Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing, China.
School of Life Sciences, Tsinghua-Peking Joint Center for Life Sciences, Beijing Advanced Innovation Center for Structural Biology, Beijing Frontier Research Center of Biological Structure, Tsinghua University, Beijing, China.
Nat Commun. 2025 Jan 2;16(1):250. doi: 10.1038/s41467-024-55531-0.
SNARE (soluble N-ethylmaleimide sensitive factor attachment protein receptor) proteins are the minimal machinery required for vesicle fusion in eukaryotes. Formation of a highly stable four-helix bundle consisting of SNARE motif of these proteins, drives vesicle/membrane fusion involved in several physiological processes such as neurotransmission. Recycling/disassembly of the protein machinery involved in membrane fusion is essential and is facilitated by an AAA+ ATPase, N-ethylmaleimide sensitive factor (NSF) in the presence of an adapter protein, α-SNAP. Here we use single-molecule fluorescence spectroscopy approaches to elucidate the chain of events that occur during the disassembly of SNARE complex by NSF. Our observations indicate two major pathways leading to the sequential disassembly of the SNARE complex: one where a syntaxin separated intermediate state is observed before syntaxin disassembles first, and a second where Vamp disassembles from the other proteins first. These studies uncover two parallel sequential pathways for the SNARE disassembly by NSF along with a syntaxin separated intermediate that couldn't be observed otherwise.
SNARE（可溶性N-乙基马来酰亚胺敏感因子附着蛋白受体）蛋白是真核生物中囊泡融合所需的最小机制。由这些蛋白的SNARE基序形成的高度稳定的四螺旋束驱动了涉及神经传递等多种生理过程的囊泡/膜融合。参与膜融合的蛋白机制的循环/拆卸至关重要，在衔接蛋白α-SNAP存在的情况下，由一种AAA+ATP酶N-乙基马来酰亚胺敏感因子（NSF）促进。在这里，我们使用单分子荧光光谱方法来阐明NSF拆解SNARE复合体过程中发生的一系列事件。我们的观察结果表明，导致SNARE复合体顺序拆解的有两条主要途径：一条途径是在Syntaxin首先拆解之前观察到Syntaxin分离的中间状态，另一条途径是Vamp首先从其他蛋白上拆解下来。这些研究揭示了NSF拆解SNARE的两条平行顺序途径以及一个Syntaxin分离的中间状态，否则无法观察到该中间状态。
