Institute of Molecular Science, Shanxi University, Taiyuan 030006, China.
Spectrochim Acta A Mol Biomol Spectrosc. 2012 Feb 15;87:163-70. doi: 10.1016/j.saa.2011.11.032. Epub 2011 Nov 23.
Centrin, a member of calcium-binding proteins, is an essential component for microtubule-organizing center (MTOC). Lanthanide (Ln) ions can increase amounts, enhance stability and orderliness of microtubules which is an important component of cytoskeleton. To investigate the structural basis of the effect of Ln ions on orderliness of microtubules, we focused on the interactions between the isolated N-terminal domain of Euplotes centrin (N-EoCen) and Ln by some combined biophysical and biochemical methods. Our results suggest that Ln ions may bind to the canonical calcium binding sites on N-EoCen. Taking advantage of ligand competition, we first determined the metal-binding affinities of Nd(3+), Eu(3+), Gd(3+) and Tm(3+) with N-EoCen. Major changes of N-EoCen in secondary and tertiary structure are noted while Ln ions bind with N-EoCen through CD spectra and 2-p-toluidinylnaphthalene-6-sulfonate (TNS) fluorescence. N-EoCen exists in the form of monomer and dimer in the presence of Ln ions. These results can provide some insights into the structural basis of how Ln ions achieve biological effect in cell through the centrin protein.
中心体蛋白是一种钙结合蛋白，是微管组织中心（MTOC）的必需组成部分。镧系（Ln）离子可以增加微管的数量，增强其稳定性和有序性，微管是细胞骨架的重要组成部分。为了研究 Ln 离子对微管有序性的影响的结构基础，我们使用一些组合生物物理和生化方法，集中研究了分离的真涡虫中心体蛋白的 N 端结构域（N-EoCen）与 Ln 之间的相互作用。我们的结果表明，Ln 离子可能与 N-EoCen 上的典型钙结合位点结合。利用配体竞争，我们首先确定了 Nd(3+)、Eu(3+)、Gd(3+)和 Tm(3+)与 N-EoCen 的金属结合亲和力。通过圆二色性（CD）光谱和 2-对甲苯磺酰基萘-6-磺酸钠（TNS）荧光实验，我们发现 N-EoCen 的二级和三级结构发生了重大变化，当 Ln 离子与 N-EoCen 结合时。在 Ln 离子存在的情况下，N-EoCen 以单体和二聚体的形式存在。这些结果可以为 Ln 离子如何通过中心体蛋白在细胞中实现生物学效应提供一些结构基础方面的见解。
