Preparation and crystallization of the disulfide-linked HLA-G dimer.
Preparation and crystallization of the disulfide-linked HLA-G dimer.
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DOI：
10.1016/j.bbapap.2005.10.006
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发表时间：
2006-05
期刊：
影响因子：
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通讯作者：
M. Shiroishi;D. Kohda;K. Maenaka
中科院分区：
文献类型：
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作者：
M. Shiroishi;D. Kohda;K. Maenaka
人类白细胞抗原-G是一种非经典的MHC-I类分子，它与抑制性受体如白细胞免疫球蛋白样受体结合，诱导广泛的耐受性免疫效应。在溶液中和细胞表面均可表达类二硫化物二聚体。然而，人类白细胞抗原-G二聚体的三维结构尚不清楚。在这里，我们报告了通过添加二硫苏糖醇(DTT)来结晶二硫键连接的二聚体形式的人类白细胞抗原-G，使得能够收集3.2？个数据集。我们还发现，DTT促进了复性的HLA-G的二硫键交换，其游离半胱氨酸受到保护，从而促进了其二聚化。这项技术也可以应用于二硫化物介导的含有游离半胱氨酸的复性蛋白的二聚体/多聚体的形成。
HLA-G is a non-classical MHC class I, which binds to inhibitory receptors, such as Leukocyte Ig-like receptors, to induce a wide range of tolerogenic immunological effects. HLA-G can be expressed as a disulfide-liked dimer both in solution and at the cell surface. However, the three-dimensional structure of the HLA-G dimer is unknown. Here, we report the crystallization of the disulfide-linked dimer form of HLA-G by adding dithiothreitol (DTT), enabling a 3.2-Å data set to be collected. We also show that DTT promotes disulfide bond exchange of refolded HLA-G, whose free cysteine was protected, thus facilitating its dimerization. This technique could also be applied for disulfide-mediated dimer/multimer formation of refolded proteins harbouring free cysteines.