Sergiev Petr V, Lesnyak Dmitry V, Bogdanov Alexey A, Dontsova Olga A
Department of Chemistry and AN Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia.
J Mol Biol. 2006 Nov 17;364(1):26-31. doi: 10.1016/j.jmb.2006.09.008. Epub 2006 Sep 8.
Escherichia coli ribosomal RNA contains five guanosine residues methylated at N2. The ygjO gene was previously predicted to methylate 16 S rRNA residue G966 due to its high sequence homology with the protein RsmC, responsible for G1207 methylation. We have identified the target of YgjO as being m2G1835 of the 23 S rRNA and not m2G966 of the 16 S rRNA as expected. Knock-out of the ygjO gene leads to loss of modification at G1835, as revealed by reverse transcription. Moreover, the modification could be restored by in vivo complementation of the ygjO knock-out strain with a plasmid expressing ygjO. Recombinant YgjO protein is able to methylate in vitro protein-free 23 S rRNA, but not assembled 50 S subunits purified from the ygjO knock-out strain. The nucleotide m2G1835 is located in a functionally extremely important region of the ribosome, being located within intersubunit bridges of group B2. Growth competition assays reveal that the lack of the G1835 methylation causes growth retardation, especially at temperatures higher than optimal and in poor media. Based on these results we suggest that YgjO be renamed to RlmG in accordance with the accepted nomenclature for rRNA methyltransferases.
大肠杆菌核糖体RNA含有五个在N2位置甲基化的鸟苷残基。由于ygjO基因与负责G1207甲基化的RsmC蛋白具有高度序列同源性，之前预测它会使16S rRNA残基G966甲基化。我们已确定YgjO的靶标是23S rRNA的m2G1835，而非预期的16S rRNA的m2G966。如逆转录所示，ygjO基因敲除导致G1835处的修饰缺失。此外，通过用表达ygjO的质粒对ygjO基因敲除菌株进行体内互补，可恢复该修饰。重组YgjO蛋白能够在体外使无蛋白的23S rRNA甲基化，但不能使从ygjO基因敲除菌株中纯化的组装好的50S亚基甲基化。核苷酸m2G1835位于核糖体功能极其重要的区域，处于B2组的亚基间桥内。生长竞争试验表明，G1835甲基化的缺失会导致生长迟缓，尤其是在高于最适温度的条件下以及在贫瘠培养基中。基于这些结果，我们建议根据rRNA甲基转移酶的公认命名法将YgjO重新命名为RlmG。
