NSF-mediated disassembly of on- and off-pathway SNARE complexes and inhibition by complexin.
NSF-mediated disassembly of on- and off-pathway SNARE complexes and inhibition by complexin.
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DOI：
10.7554/elife.36497
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发表时间：
2018-07-09
期刊：
影响因子：
7.7
通讯作者：
Brunger AT
中科院分区：
文献类型：
--
作者：
Choi UB;Zhao M;White KI;Pfuetzner RA;Esquivies L;Zhou Q;Brunger AT
atp酶NSF分解SNARE复合物对于神经递质释放和其他膜运输过程至关重要。我们开发了一种单分子FRET试验来监测nsf介导的单个SNARE复合物的反复拆卸和重组。对于三元神经元SNARE复合物，分解过程在100毫秒内完成。我们观察到短（<0.32 s）和长（≥0.32 s）的解体状态。长寿命状态表示完全拆卸的SNARE复合体，而短寿命状态对应于失败的拆卸或立即重新组装。较高的离子强度或αSNAP浓度降低都降低了分解速率，同时增加了短寿命态的频率。NSF还能够分解反平行三元SNARE复合物，这意味着它可以用于质量控制。最后，络合素-1与α - snap结合SNARE复合体竞争；络合素-1的加入与α - snap浓度的降低具有相似的作用，可能对顺式和反式SNARE复合物的分解具有差异调节作用。
SNARE complex disassembly by the ATPase NSF is essential for neurotransmitter release and other membrane trafficking processes. We developed a single-molecule FRET assay to monitor repeated rounds of NSF-mediated disassembly and reassembly of individual SNARE complexes. For ternary neuronal SNARE complexes, disassembly proceeds in a single step within 100 msec. We observed short- (<0.32 s) and long-lived (≥0.32 s) disassembled states. The long-lived states represent fully disassembled SNARE complex, while the short-lived states correspond to failed disassembly or immediate reassembly. Either high ionic strength or decreased αSNAP concentration reduces the disassembly rate while increasing the frequency of short-lived states. NSF is also capable of disassembling anti-parallel ternary SNARE complexes, implicating it in quality control. Finally, complexin-1 competes with αSNAP binding to the SNARE complex; addition of complexin-1 has an effect similar to that of decreasing the αSNAP concentration, possibly differentially regulating cis and trans SNARE complexes disassembly.