Wang Kai Tuo, Ma Linglong, Nan Jie, Su Xiao Dong, Li Lanfen
The National Laboratory of Protein Engineering and Plant Genetic Engineering, School of Life Sciences, Peking University, Beijing 100871, People's Republic of China.
Acta Crystallogr Sect F Struct Biol Cryst Commun. 2010 Nov 1;66(Pt 11):1484-6. doi: 10.1107/S1744309110035074. Epub 2010 Oct 28.
The RlmL (YcbY) protein in Escherichia coli is an rRNA methyltransferase that is specific for m(2)G2445 modification of 23S RNA. The rlmL gene was cloned into the expression vector pET28a and expressed in the host E. coli strain BL21 (DE3). Recombinant protein with a six-histidine tag was purified by Ni(2+)-affinity chromatography followed by gel filtration. Crystals were grown using the hanging-drop vapour-diffusion method and a detergent was used as an additive to improve diffraction quality. The final crystals diffracted to 2.2 Å resolution. The crystals belonged to space group P2(1), with unit-cell parameters a = 73.6, b = 140.8, c = 102.9 Å, β = 102.3°. The crystal has a most probable solvent content of 62.8% with two molecules in the asymmetric unit.
大肠杆菌中的RlmL（YcbY）蛋白是一种rRNA甲基转移酶，对23S RNA的m（2）G2445修饰具有特异性。将rlmL基因克隆到表达载体pET28a中，并在宿主大肠杆菌菌株BL21（DE3）中表达。带有六个组氨酸标签的重组蛋白通过Ni（2+）亲和层析，然后进行凝胶过滤进行纯化。使用悬滴气相扩散法生长晶体，并使用去污剂作为添加剂来提高衍射质量。最终晶体的衍射分辨率达到2.2 Å。晶体属于空间群P2（1），晶胞参数a = 73.6，b = 140.8，c = 102.9 Å，β = 102.3°。该晶体的最可能溶剂含量为62.8%，不对称单元中有两个分子。
