Crystallization and preliminary X-ray crystallographic study of the editing domain of Thermus thermophilus isoleucyl-tRNA synthetase complexed with pre- and post-transfer editing-substrate analogues
Crystallization and preliminary X-ray crystallographic study of the editing domain of Thermus thermophilus isoleucyl-tRNA synthetase complexed with pre- and post-transfer editing-substrate analogues
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DOI：
10.1107/s0907444904019511
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发表时间：
2004-10-01
影响因子：
2.2
通讯作者：
Yokoyama, S
中科院分区：
文献类型：
--
作者：
Fukunaga, R;Yokoyama, S
异亮氨酰-tRNA合成酶（IleRS）的CP 1结构域（编辑结构域）在转移前编辑中水解错误激活的Val-AMP，在转移后编辑中水解错误充电的Val-tRNA（Ile）。分离表达嗜热栖热菌IleRS的CP 1结构域，纯化并与Val-AMS（Val-AMP类似物）和瓦尔-2AA（Val-tRNA（Ile）类似物）共结晶。两种不同的表达构建体用于每种共结晶。Val-AMS配合物晶体属四面体空间群P4（1）2（1）2，晶胞参数a = B = 102.00，c = 84.88埃。不对称单元含有两个分子的CP 1结构域，每蛋白质重量的相应晶体体积为2.7埃（3）Da（-1），溶剂含量为53.5%。瓦尔-2AA配合物晶体属四面体空间群P4（1）22，晶胞参数a = B = 72.59，c= 83.68埃。不对称单元包含一个分子的CP 1结构域，每蛋白质重量的相应晶体体积为2.8埃（3）Da（-1），溶剂含量为55.8%。在100 K下从每个单晶收集衍射至1.7埃分辨率的数据集。
The CP1 domain (the editing domain) of isoleucyl-tRNA synthetase (IleRS) hydrolyzes misactivated Val-AMP in pre-transfer editing and mischarged Val-tRNA(Ile) in post-transfer editing. The CP1 domain of Thermus thermophilus IleRS was expressed in isolation, purified and cocrystallized with Val-AMS (a Val-AMP analogue) and with Val-2AA (a Val-tRNA(Ile) analogue). Two different expression constructs were used for each cocrystallization. The complex crystals with Val-AMS belong to the tetragonal space group P4(1)2(1)2, with unit-cell parameters a = b = 102.00, c = 84.88 Angstrom. The asymmetric unit contains two molecules of the CP1 domain, with a corresponding crystal volume per protein weight of 2.7 Angstrom(3) Da(-1) and a solvent content of 53.5%. The complex crystals with Val-2AA belong to the tetragonal space group P4(1)22, with unit-cell parameters a = b = 72.59, c= 83.68 Angstrom. The asymmetric unit contains one molecule of the CP1 domain, with a corresponding crystal volume per protein weight of 2.8 Angstrom(3) Da(-1) and a solvent content of 55.8%. Data sets diffracting to 1.7 Angstrom resolution were collected from each single crystal at 100 K.