Structures of the Phosphorylated and VO3-Bound 2H-Phosphatase Domain of Sts-2
Structures of the Phosphorylated and VO3-Bound 2H-Phosphatase Domain of Sts-2
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DOI：
10.1021/bi9008648
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发表时间：
2009-09-01
期刊：
影响因子：
2.9
通讯作者：
Nassar, Nicolas
中科院分区：
文献类型：
--
作者：
Chen, Yunting;Jakoncic, Jean;Nassar, Nicolas
T细胞受体(TCR)信号转导抑制因子1和2(Sts-1和Sts-2)蛋白的C末端结构域与2H-磷酸酶家族具有同源性。相应的Sts-1结构域Sts-1(PGM)的磷酸酶活性是其负性调节TCR和表皮生长因子受体(EGFR)等膜结合受体信号的关键。在磷酸酶反应过程中短暂磷酸化的亲核组氨酸对活性是必不可少的。在这里，我们介绍了Sts-2(PGM)的磷酸化活性形式并结合到VO_3上的晶体结构，它代表了脱磷反应路径上的中间体和过渡态类似物的结构。在前一种结构中，所提出的亲核His366是唯一的光短化残基，通过与活性中心内保守的碱性残基的几次相互作用而稳定。在后一种结构中，钒原子位于由VO_3分子的三个氧原子、His366的NE_2原子和顶端水分子W-a的三个氧原子组成的三角双金字塔的中间。V-NE2键长(2.25埃)表明VO_3不是共价键合在His366上的，反应机理是部分缔合的。这两个结构还表明Glu476在激活一个独特位置的水分子方面发挥了作用。在这两种结构中，活性中心的构象与apo-Sts-2(PGM)中的构象非常相似，表明催化残基的空间排列在脱磷反应过程中没有改变。
The C-terminal domain of the suppressor of T cell receptor (TCR) signaling 1 and 2 (Sts-1 and -2) proteins has homology to the 2H-phosphatase family of enzymes. The phosphatase activity of the correspondent Sts-1 domain, Sts-1(PGM), is key for its ability to negatively regulate the signaling of membrane-bound receptors including TCR and the epidermal growth factor receptor (EGFR). A nucleophilic histidine, which is transiently phosphorylated during the phosphatase reaction, is essential for the activity. Here, we present the crystal structure of Sts-2(PGM) in the phosphorylated active form and bound to VO3, which represent structures of an intermediate and of a transition state analogue along the path of the dephosphorylation reaction. In the former structure, the proposed nucleophilic His366 Is the only phoshorylated, residue and is stabilized by several interactions with conserved basic residues within the active site. In the latter structure, the vanadium atom sits in the middle of a trigonal bipyramid formed by the three oxygen atoms of the VO3 molecule, atom NE2 of His366, and an apical water molecule W-a. The V-NE2 bond length (2.25 angstrom) suggests that VO3 is not covalently attached to His366 and that the reaction mechanism is partially associative. The two structures also suggest a role for Glu476 in activating a uniquely positioned water molecule. In both structures, the conformation of the active site is remarkably similar to the one seen in apo-Sts-2(PGM) suggesting that the spatial arrangement of the catalytic residues does not change during the dephosphorylation reaction.