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

Acta Crystallographica. Section D, Biological Crystallography
Ryuya Fukunaga, Shigeyuki Yokoyama

Abstract

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 A. The asymmetric unit contains two molecules of the CP1 domain, with a corresponding crystal volume per protein weight of 2.7 A3 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 A. The asymmetric unit contains one molecule of the CP1 domain, with a corresponding crystal volume per protein weight of 2.8 A3 Da(-1) and a solvent content of 55.8%. Data sets diffracting to 1.7 A resolution were collected from each single crystal at 100 K.

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