Usui Takuya, Ono Sayaka, Nakamura Akiyoshi, Kato Koji, Ose Toyoyuki, Yao Min
Graduate School of Life Science, Hokkaido University, Kita 10, Nishi 8, Kita-ku, Sapporo, Hokkaido 060-0810, Japan.
Faculty of Advanced Life Science, Hokkaido University, Kita 10, Nishi 8, Kita-ku, Sapporo, Hokkaido 060-0810, Japan.
Acta Crystallogr F Struct Biol Commun. 2025 Feb 1;81(Pt 2):35-40. doi: 10.1107/S2053230X25000044. Epub 2025 Jan 9.
The bacterial enzyme tRNA 2-selenouridine synthase (SelU) catalyzes the conversion of 5-substituted 2-thiouridine (R5S2U) to 5-substituted 2-selenouridine (R5Se2U) at the wobble positions of several tRNAs. Seleno-modification potentially regulates translation efficiency in response to selenium availability. Notably, SelU uses the 2-geranylthiouridine (R5geS2U) intermediate for sulfur removal, and this geranylthiol (geS) is a unique leaving group among tRNA-maturation enzymes. However, the underlying sequence of the SelU reaction remains unclear. Here, a crystallographic study of the Escherichia coli SelU-tRNA complex is reported. Robust and well formed SelU-tRNA crystals were obtained after several optimizations, including co-expression with tRNA and additive screening. Diffraction data were collected at a resolution of 3.10 Å using a wavelength of 1.0000 Å. The crystals belonged to space group C2, and the phase was determined by molecular replacement using the AlphaFold2-predicted SelU structure as a search model. Electron-density mapping revealed the presence of two SelU-tRNA complexes in the asymmetric unit.
细菌酶tRNA 2-硒代尿苷合酶（SelU）可催化几种tRNA摆动位置上的5-取代2-硫代尿苷（R5S2U）转化为5-取代2-硒代尿苷（R5Se2U）。硒修饰可能会根据硒的可利用性来调节翻译效率。值得注意的是，SelU利用2-香叶基硫代尿苷（R5geS2U）中间体进行脱硫，而这种香叶基硫醇（geS）是tRNA成熟酶中独特的离去基团。然而，SelU反应的潜在序列仍不清楚。在此，报道了大肠杆菌SelU-tRNA复合物的晶体学研究。经过包括与tRNA共表达和添加剂筛选在内的多次优化后，获得了坚固且形态良好的SelU-tRNA晶体。使用波长为1.0000 Å收集了分辨率为3.10 Å的衍射数据。晶体属于空间群C2，通过以AlphaFold2预测的SelU结构作为搜索模型进行分子置换来确定相位。电子密度图显示在不对称单元中存在两个SelU-tRNA复合物。
