STRUCTURE OF TUBULIN AT 6.5 ANGSTROM AND LOCATION OF THE TAXOL-BINDING SITE
STRUCTURE OF TUBULIN AT 6.5 ANGSTROM AND LOCATION OF THE TAXOL-BINDING SITE
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DOI：
10.1038/375424a0
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发表时间：
1995-06-01
期刊：
影响因子：
64.8
通讯作者：
DOWNING, KH
中科院分区：
文献类型：
--
作者：
NOGALES, E;WOLF, SG;DOWNING, KH
微管蛋白是微管的主要成分，是α和β两条链的异源二聚体，两者的相对分子质量均为50，000（M（r）50 K），同源性为40-50%。微管蛋白的同种型多样性（2）和构象的灵活性，到目前为止，使得不可能获得用于X射线工作的晶体（3）。微管蛋白的结构知识仅限于来自定向微管的X射线衍射（4）和来自负染色中微管和锌诱导的结晶片的电子显微镜（5，6）的约20埃。这些片层由类似于微管中的原丝组成，但以反平行排列（7），它们的二维特征对于高分辨率电子显微镜是理想的（8，9）。在这里，我们展示了微管蛋白的三维重建，分辨率为6.5埃，通过锌诱导的蛋白质二维晶体的电子晶体学获得。α和β亚基在拓扑学上相似，与它们的序列同源性一致（10）。可以根据二级结构定义几个特征。一个明显的α-螺旋部分，相邻的interdimer和inter-protofilament接触，暂时归因于一个片段附近的羧基末端的蛋白质。我们可以根据紫杉醇结合的投影研究来分配α-和β-亚基 *，其显示每个微管蛋白异源二聚体有一个紫杉醇位点，与微管中紫杉醇的已知化学计量一致（11）。这些研究表明，紫杉醇影响原丝之间的相互作用;据我们所知，这是第一次在微管蛋白分子中观察到配体结合位点。
TUBULIN, the major component of microtubules, is a heterodimer of two chains, alpha and beta(1), both of relative molecular mass 50,000 (M(r) 50K) and sith 40-50% identity. The isotypic variety(2) and conformational flexibility of tubulin have so far made it impossible to obtain crystals for X-ray work(3). Structural knowledge of tubulin has been limited to about 20 Angstrom from X-ray diffraction of oriented microtubules(4), and from electron microscopy of microtubules and zinc-induced crystalline sheets in negative stain(5,6). The sheets consist of protofilaments similar to those in microtubules but associated in an antiparallel arrangement(7) and their two-dimensional character is ideal for high-resolution electron microscopy(8,9). Here we present a three-dimensional reconstruction of tubulin to 6.5 Angstrom resolution, obtained by electron crystallography of zinc-induced two-dimensional crystals of the protein, The alpha- and beta-subunits appear topologically similar, in agreement with their sequence homology(10). Several features can be defined in terms of secondary structure. An apparent alpha-helical portion, adjacent to both interdimer and inter-protofilament contacts, is tentatively attributed to a segment near the carboxy terminus of the protein. We can assign the alpha- and beta-subunits on the basis of projection studies of the binding of taxol*, which show one taxol site per tubulin heterodimer, in agreement with the known stoichiometry of taxol in microtubules(11). These studies indicate that taxol affects the interaction between protofilaments; to our knowledge, this is the first time that a ligand-binding site has been visualized in the tubulin molecule.