title: Crystallization of leucyl-tRNA synthetase complexed with tRNALeu from the archaeon Pyrococcus horikoshii.
abstract: All five tRNALeu 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 tRNALeu isoacceptors were attempted. Cocrystals were obtained by the hanging-drop vapour-diffusion method, but only when the tRNALeu isoacceptor with the anticodon CAA was used. Electrophoretic analyses revealed that the crystals contain both LeuRS and tRNALeu, suggesting that they are LeuRS-tRNALeu complex crystals. A data set diffracting to 3.3 A resolution was collected from a single crystal at 100 K. The crystal belongs to the orthorhombic space group P2(1)2(1)2, with unit-cell parameters a = 118.18, b = 120.55, c = 231.13 A. The asymmetric unit is expected to contain two complexes of LeuRS-tRNALeu, with a corresponding crystal volume per protein weight of 2.9 A3 Da(-1) and a solvent content of 57.3%.
authors.affiliation: Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Japan.
authors.forename: Ryuya
authors.initials: R
authors.lastname: Fukunaga
authors.forename: Ryuichiro
authors.initials: R
authors.lastname: Ishitani
authors.forename: Osamu
authors.initials: O
authors.lastname: Nureki
authors.forename: Shigeyuki
authors.initials: S
authors.lastname: Yokoyama
mesh_terms: D019843:Archaeal Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D018360:Crystallography, X-Ray; D007935:Leucine-tRNA Ligase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D008970:Molecular Weight; D011485:Protein Binding; D043902:Pyrococcus horikoshii / Q000201:enzymology*; D019642:RNA, Archaeal / Q000378:metabolism; D012356:RNA, Transfer, Leu / Q000378:metabolism*; D014961:X-Ray Diffraction
chemical_list: D019843:Archaeal Proteins; D019642:RNA, Archaeal; D012356:RNA, Transfer, Leu; D007935:Leucine-tRNA Ligase