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Updated: May 27, 2026
X-Ray Crystallography to Study the Oligomeric State Transition of the Thermotoga maritima M42 Aminopeptidase TmPep1050
Published on: May 13, 2020
Crystallization and preliminary X-ray diffraction analysis of an archaeal tRNA-modification enzyme, TiaS, complexed
Takuo Osawa1, Hideko Inanaga, Satoshi Kimura
1Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), 1-1-1 Higashi, Tsukuba-shi, Ibaraki 305-8566, Japan.
Abstract:
The cytidine at the first anticodon position of archaeal tRNA(Ile2), which decodes the isoleucine AUA codon, is modified to 2-agmatinylcytidine (agm(2)C) to guarantee the fidelity of protein biosynthesis. This post-transcriptional modification is catalyzed by tRNA(Ile)-agm(2)C synthetase (TiaS) using ATP and agmatine as substrates. Archaeoglobus fulgidus TiaS was overexpressed in Escherichia coli cells and purified. tRNA(Ile2) was prepared by in vitro transcription with T7 RNA polymerase. TiaS was cocrystallized with both tRNA(Ile2) and ATP by the vapour-diffusion method. The crystals of the TiaS-tRNA(Ile2)-ATP complex diffracted to 2.9 Å resolution using synchrotron radiation at the Photon Factory. The crystals belonged to the primitive hexagonal space group P3(2)21, with unit-cell parameters a = b = 131.1, c = 86.6 Å. The asymmetric unit is expected to contain one TiaS-tRNA(Ile2)-ATP complex, with a Matthews coefficient of 2.8 Å(3) Da(-1) and a solvent content of 61%.
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