Crystallization and preliminary X-ray crystallographic analysis of bacterial tRNA(Sec) in complex with seryl-tRNA synthetase.
Selenocysteine (Sec) is translationally incorporated into proteins in response to the UGA codon. The tRNA specific to Sec (tRNA(Sec)) is first ligated with serine by seryl-tRNA synthetase (SerRS). To elucidate the tertiary structure of bacterial tRNA(Sec) and its specific interaction with SerRS, the bacterial tRNA(Sec) from Aquifex aeolicus was crystallized as the heterologous complex with the archaeal SerRS from Methanopyrus kandleri. Although X-ray diffraction by crystals of tRNA(Sec) in complex with wild-type SerRS was rather poor (to 5.7 Å resolution), the resolution was improved by introducing point mutations targeting the crystal-packing interface. Heavy-atom labelling also contributed to resolution improvement. A 3.2 Å resolution diffraction data set for phase determination was obtained from a K(2)Pt(CN)(4)-soaked crystal.
Department of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Yuzuru
Y
Itoh
Shun Ichi
SI
Sekine
Shigeyuki
S
Yokoyama
D001419:Bacteria / Q000201:enzymology*; D003460:Crystallization; D018360:Crystallography, X-Ray; D019605:Euryarchaeota / Q000201:enzymology*; D017354:Point Mutation; D011485:Protein Binding; D012343:RNA, Transfer / Q000737:chemistry* / Q000378:metabolism; D012714:Serine-tRNA Ligase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D012343:RNA, Transfer; D012714:Serine-tRNA Ligase