Laboratory of Structural Cell Biology, Fundación Instituto Leloir and Instituto de Investigaciones Bioquímicas de Buenos Aires, Avenida Patricias Argentinas 435, C1405BWE Buenos Aires, Argentina.
J Biol Chem. 2010 Feb 12;285(7):4544-53. doi: 10.1074/jbc.M109.034512. Epub 2009 Dec 15.
Calreticulin is an abundant endoplasmic reticulum resident protein that fulfills at least two basic functions. Firstly, due to its ability to bind monoglucosylated high mannose oligosaccharides, calreticulin is a central component of the folding quality control system of glycoproteins. On the other hand, thanks to its capacity to bind high amounts of calcium, calreticulin is one of the main calcium buffers in the endoplasmic reticulum. This last activity resides on a highly negatively charged domain located at the C terminus. Interestingly, this domain has been proposed to regulate the intracellular localization of calreticulin. Structural information for this domain is currently scarce. Here we address this issue by employing a combination of biophysical techniques and molecular dynamics simulation. We found that calreticulin C-terminal domain at low calcium concentration displays a disordered structure, whereas calcium addition induces a more rigid and compact conformation. Remarkably, this change develops when calcium concentration varies within a range similar to that taking place in the endoplasmic reticulum upon physiological fluctuations. In addition, a much higher calcium concentration is necessary to attain similar responses in a peptide displaying a randomized sequence of calreticulin C-terminal domain, illustrating the sequence specificity of this effect. Molecular dynamics simulation reveals that this ordering effect is a consequence of the ability of calcium to bring into close proximity residues that lie apart in the primary structure. These results place calreticulin in a new setting in which the protein behaves not only as a calcium-binding protein but as a finely tuned calcium sensor.
钙网蛋白是内质网中丰富的驻留蛋白，至少具有两种基本功能。首先，由于其结合单葡萄糖化高甘露糖寡糖的能力，钙网蛋白是糖蛋白折叠质量控制系统的核心组成部分。另一方面，由于其结合大量钙的能力，钙网蛋白是内质网中主要的钙缓冲剂之一。最后一项活动位于 C 末端的一个带负电荷的高域上。有趣的是，该结构域被提议调节钙网蛋白的细胞内定位。目前，关于该结构域的结构信息非常有限。在这里，我们通过结合生物物理技术和分子动力学模拟来解决这个问题。我们发现，钙网蛋白 C 末端结构域在低钙浓度下显示出无序结构，而钙的加入诱导出更刚性和紧凑的构象。值得注意的是，当钙浓度在生理波动期间内质网中发生变化的范围内变化时，会发生这种变化。此外，需要更高的钙浓度才能在显示钙网蛋白 C 末端结构域随机序列的肽中获得类似的反应，这表明了这种效应的序列特异性。分子动力学模拟表明，这种有序化效应是钙能够将远离一级结构的残基紧密结合在一起的结果。这些结果将钙网蛋白置于一个新的环境中，在这个环境中，该蛋白不仅作为钙结合蛋白，而且作为一个精细调节的钙传感器发挥作用。
