Pobbati Ajaybabu V, Razeto Adelia, Böddener Matthias, Becker Stefan, Fasshauer Dirk
Department of Neurobiology, Max-Planck-Institute for Biophysical Chemistry, Am Fassberg 11, 37077 Göttingen, Germany.
J Biol Chem. 2004 Nov 5;279(45):47192-200. doi: 10.1074/jbc.M408767200. Epub 2004 Aug 16.
Upon Ca2+ influx synaptic vesicles fuse with the plasma membrane and release their neurotransmitter cargo into the synaptic cleft. Key players during this process are the Q-SNAREs syntaxin 1a and SNAP-25 and the R-SNARE synaptobrevin 2. It is thought that these membrane proteins gradually assemble into a tight trans-SNARE complex between vesicular and plasma membrane, ultimately leading to membrane fusion. Tomosyn is a soluble protein of 130 kDa that contains a COOH-terminal R-SNARE motif but lacks a transmembrane anchor. Its R-SNARE motif forms a stable core SNARE complex with syntaxin 1a and SNAP-25. Here we present the crystal structure of this core tomosyn SNARE complex at 2.0-A resolution. It consists of a four-helical bundle very similar to that of the SNARE complex containing synaptobrevin. Most differences are found on the surface, where they prevented tight binding of complexin. Both complexes form with similar rates as assessed by CD spectroscopy. In addition, synaptobrevin cannot displace the tomosyn helix from the tight complex and vice versa, indicating that both SNARE complexes represent end products. Moreover, data bank searches revealed that the R-SNARE motif of tomosyn is highly conserved throughout all eukaryotic kingdoms. This suggests that the formation of a tight SNARE complex is important for the function of tomosyn.
钙离子流入时，突触小泡与质膜融合，并将其神经递质货物释放到突触间隙中。这一过程中的关键蛋白是Q-SNARE蛋白 syntaxin 1a和SNAP-25以及R-SNARE蛋白突触小泡蛋白2。据认为，这些膜蛋白逐渐组装成囊泡膜和质膜之间紧密的反式SNARE复合体，最终导致膜融合。Tomosyn是一种130 kDa的可溶性蛋白，含有一个COOH末端R-SNARE基序，但缺乏跨膜锚定结构。其R-SNARE基序与syntaxin 1a和SNAP-25形成稳定的核心SNARE复合体。在此，我们展示了该核心Tomosyn SNARE复合体在2.0埃分辨率下的晶体结构。它由一个四螺旋束组成，与含有突触小泡蛋白的SNARE复合体非常相似。大多数差异出现在表面，在那里它们阻止了复合体蛋白的紧密结合。通过圆二色光谱法评估，两种复合体以相似的速率形成。此外，突触小泡蛋白不能从紧密复合体中取代Tomosyn螺旋，反之亦然，这表明两种SNARE复合体都代表终产物。此外，数据库搜索显示，Tomosyn的R-SNARE基序在所有真核生物界中高度保守。这表明紧密SNARE复合体的形成对Tomosyn的功能很重要。
