Lesnyak Dmitry V, Sergiev Petr V, Bogdanov Alexey A, Dontsova Olga A
Department of Bioinformatics and Bioengineering, Moscow State University, Moscow, Russia.
J Mol Biol. 2006 Nov 17;364(1):20-5. doi: 10.1016/j.jmb.2006.09.009. Epub 2006 Sep 8.
N2-methylguanosine 2445 of the 23 S rRNA is located in a cluster of modified nucleotides concentrated at the peptidyl transferase center of the ribosome. Here we describe the identification of a gene, ycbY, as encoding an enzyme responsible for methylation of G2445. Knock-out of the ycbY gene leads to loss of modification at G2445 as revealed by reverse transcription. The modification is restored in the ycbY knock-out strain if co-transformed with a plasmid expressing the ycbY gene product. Recombinant YcbY protein is able to methylate 23 S rRNA purified from the ycbY knock-out strain in vitro, assembled 50 S subunits are not a substrate for the methylase. Knock-out of the ycbY gene leads to growth retardation. Growth competition with the parental wild-type strain leads to a gradual decrease in the knock-out strain cells proportion in the media. It is likely that the G2445 modification is necessary for prevention of non-functional secondary or tertiary structure formation at the peptidyl transferase center. Based on these results we suggest that YcbY be renamed to RlmL in accordance with the accepted nomenclature for rRNA methyltransferases.
23S rRNA的N2-甲基鸟苷2445位于核糖体肽基转移酶中心集中的修饰核苷酸簇中。在此，我们描述了一个基因ycbY的鉴定，该基因编码一种负责G2445甲基化的酶。如逆转录所示，ycbY基因的敲除导致G2445处修饰的缺失。如果与表达ycbY基因产物的质粒共转化，ycbY敲除菌株中的修饰得以恢复。重组YcbY蛋白能够在体外使从ycbY敲除菌株中纯化的23S rRNA甲基化，组装好的50S亚基不是甲基化酶的底物。ycbY基因的敲除导致生长迟缓。与亲本野生型菌株的生长竞争导致培养基中敲除菌株细胞比例逐渐降低。G2445修饰可能对于防止肽基转移酶中心形成无功能的二级或三级结构是必要的。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YcbY重新命名为RlmL。
