Membrane Docking of the Synaptotagmin 7 C2A Domain: Electron Paramagnetic Resonance Measurements Show Contributions from Two Membrane Binding Loops.
Membrane Docking of the Synaptotagmin 7 C2A Domain: Electron Paramagnetic Resonance Measurements Show Contributions from Two Membrane Binding Loops.
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DOI：
10.1021/acs.biochem.5b00421
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发表时间：
2015-09-22
期刊：
影响因子：
2.9
通讯作者：
Knight JD
中科院分区：
文献类型：
--
作者：
Osterberg JR;Chon NL;Boo A;Maynard FA;Lin H;Knight JD
突触结合蛋白（Syt）家族的蛋白质起着重要的作用，在囊泡对接和融合过程中的钙离子诱导的胞吐在各种各样的细胞类型。其作为Ca 2+传感器的作用主要来自其两个C2结构域C2 A和C2B，其在结合Ca 2+时插入阴离子脂质膜。Syt亚型1和7在其Ca 2+敏感性方面显著不同; Syt 7的C2 A结构域比Syt 1的C2 A结构域更紧密地结合Ca 2+和膜，至少部分是由于疏水作用的贡献更大。虽然Syt 1的结构和膜活性已被广泛研究，但Syt 7和Syt 1之间差异的结构起源尚不清楚。本研究使用定点自旋标记和电子顺磁共振光谱法来确定Syt 7 C2 A结构域的深度参数，以与Syt 1 C2 A的类似先前测量进行比较。在一种新的方法中，Syt 1和Syt 7 C2 A的膜对接的几何形状建模映射深度参数到多个分子动力学模拟的Ca 2+结合蛋白质的结构。该模型揭示了Ca 2+结合环（CBL）1和3的膜渗透，并且膜结合对CBL 3中的突变更敏感。平均而言，Syt 7 C2 A比Syt 1 C2 A更深地插入膜中，尽管不同结构模型之间的深度不同。这一观察结果为Syt 7 C2 A的疏水驱动膜对接提供了部分结构解释。
The synaptotagmin (Syt) family of proteins plays an important role in vesicle docking and fusion during Ca2+-induced exocytosis in a wide variety of cell types. Its role as a Ca2+ sensor derives primarily from its two C2 domains, C2A and C2B, which insert into anionic lipid membranes upon binding Ca2+. Syt isoforms 1 and 7 differ significantly in their Ca2+ sensitivity; the C2A domain from Syt7 binds Ca2+ and membranes much more tightly than the C2A domain from Syt1, due at least in part to greater contributions from the hydrophobic effect. While the structure and membrane activity of Syt1 have been extensively studied, the structural origins of differences between Syt7 and Syt1 are unknown. The present study used site-directed spin labeling and electron paramagnetic resonance spectroscopy to determine depth parameters for the Syt7 C2A domain, for comparison to analogous previous measurements with Syt1 C2A. In a novel approach, the membrane docking geometry of both Syt1 and Syt7 C2A was modeled by mapping depth parameters onto multiple molecular dynamics simulated structures of the Ca2+-bound protein. The models reveal membrane penetration of Ca2+ binding loops (CBLs) 1 and 3, and membrane binding is more sensitive to mutations in CBL3. On average, Syt7 C2A inserts more deeply in the membrane than Syt1 C2A, although depths vary among the different structural models. This observation provides a partial structural explanation for the hydrophobically driven membrane docking of Syt7 C2A.