Volatile anesthetics bind rat synaptic snare proteins
Volatile anesthetics bind rat synaptic snare proteins
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DOI：
10.1097/00000542-200510000-00015
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发表时间：
2005-10-01
期刊：
影响因子：
8.8
通讯作者：
Crowder, M
中科院分区：
文献类型：
--
作者：
Nagele, P;Mendel, JB;Crowder, M
背景资料：挥发性全身麻醉药（VA）具有许多突触作用，其中之一是抑制兴奋性神经递质的释放;然而，尚未发现突触前VA结合蛋白。秀丽隐杆线虫的遗传数据已经导致了这样的假设，即与突触前蛋白syntaxin 1A相互作用的蛋白质是VA靶标。出于这一假设，作者测量的能力，突触融合蛋白IA和蛋白质的相互作用，与突触融合蛋白结合氟烷和isofluranes.Methods：重组大鼠突触融合蛋白1A，SNAP-25 B，VAMP 2，和三元陷阱复合物，它们形成了测试。通过F-19-核磁共振弛豫测量检测VA与这些蛋白质的结合。通过圆二色性和形成复合物的能力检查蛋白质的结构改变。结果：挥发性麻醉剂不与VAMP 2结合。在临床范围内的浓度下，VA确实与SNA-P-25 B结合;然而，仅在含有SNAP-25 B同源多聚体的制剂中检测到结合。VA还在临床浓度下与突触融合蛋白和SNARE复合物结合。在突触融合蛋白的N-末端添加His（6）标签后，尽管突触融合蛋白具有正常的二级结构和形成SNARE复合物的能力，但其结合VA的能力被破坏;凝血酶切割标签恢复了VA结合。因此，VA结合位点具有结构要求，并且不仅仅是任何α-螺旋束。VA在超临床浓度产生的SNARE复合物的螺旋度增加，否则，VA结合产生的SNARE complex.Conclusion：SNARE蛋白的稳定性或二级结构没有明显的改变是挥发性麻醉剂的潜在突触靶点。
Background: Volatile general anesthetics (VAs) have a number of synaptic actions, one of which is to inhibit excitatory neurotransmitter release; however, no presynaptic VA binding proteins have been identified. Genetic data in Caenorhabditis elegans have led to the hypothesis that a protein that interacts with the presynaptic protein syntaxin 1A is a VA target. Motivated by this hypothesis, the authors measured the ability of syntaxin IA and proteins that interact with syntaxin to bind to halothane and isoflurane.Methods: Recombinant rat syntaxin 1A, SNAP-25B, VAMP2, and the ternary SNARE complex that they form were tested. Binding of VAs to these proteins was detected by F-19-nuclear magnetic resonance relaxation measurements. Structural alterations in the proteins were examined by circular dichroism and ability to form complexes.Results: Volatile anesthetics did not bind to VAMP2. At concentrations in the clinical range, VAs did bind to SNA-P-25B; however, binding was detected only in preparations containing SNAP-25B homomultimers. VAs also bound at clinical concentrations to both syntaxin and the SNARE complex. Addition of an N-terminal His(6) tag to syntaxin abolished its ability to bind VAs despite normal secondary structure and ability to form SNARE complexes; thrombin cleavage of the tag restored VA binding. Thus, the VA binding site(s) has structural requirements and is not simply any a-helical bundle. VAs at supraclinical concentrations produced an increase in helicity of the SNARE complex; otherwise, VA binding produced no gross alteration in the stability or secondary structure of the SNARE complex.Conclusion: SNARE proteins are potential synaptic targets of volatile anesthetics.