Crystallization of leucyl-tRNA synthetase complexed with tRNA{sup Leu} from the archaeon Pyrococcus horikoshii
- Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo (Japan)
The leucyl-tRNA synthetase (LeuRS) from P. horikoshii has been overexpressed in Escherichia coli and purified, and cocrystallizations with each of the tRNA{sup Leu} isoacceptors have been attempted. Cocrystals were obtained by the hanging-drop vapour-diffusion method, but only when the tRNA{sup Leu} isoacceptor with the anticodon CAA was used. All five tRNA{sup Leu} isoacceptors from the archaeon Pyrococcus horikoshii have been transcribed in vitro and purified. The leucyl-tRNA synthetase (LeuRS) from P. horikoshii was overexpressed in Escherichia coli and purified, and cocrystallizations with each of the tRNA{sup Leu} isoacceptors were attempted. Cocrystals were obtained by the hanging-drop vapour-diffusion method, but only when the tRNA{sup Leu} isoacceptor with the anticodon CAA was used. Electrophoretic analyses revealed that the crystals contain both LeuRS and tRNA{sup Leu}, suggesting that they are LeuRS–tRNA{sup Leu} complex crystals. A data set diffracting to 3.3 Å resolution was collected from a single crystal at 100 K. The crystal belongs to the orthorhombic space group P2{sub 1}2{sub 1}2, with unit-cell parameters a = 118.18, b = 120.55, c = 231.13 Å. The asymmetric unit is expected to contain two complexes of LeuRS–tRNA{sup Leu}, with a corresponding crystal volume per protein weight of 2.9 Å{sup 3} Da{sup −1} and a solvent content of 57.3%.
- OSTI ID:
- 22356082
- Journal Information:
- Acta Crystallographica. Section F, Journal Name: Acta Crystallographica. Section F Journal Issue: Pt 1 Vol. 61; ISSN ACSFCL; ISSN 1744-3091
- Country of Publication:
- United Kingdom
- Language:
- English
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