Lartigue Carole, Lebaudy Anne, Blanchard Alain, El Yacoubi Basma, Rose Simon, Grosjean Henri, Douthwaite Stephen
INRA, UMR 1332 de Biologie du Fruit et Pathologie, F-33140 Villenave d'Ornon, France Université de Bordeaux, UMR 1332 de Biologie du Fruit et Pathologie, F-33140 Villenave d'Ornon, France.
Department of Microbiology and Cell Science, University of Florida, FL 32611, USA.
Nucleic Acids Res. 2014 Jul;42(12):8073-82. doi: 10.1093/nar/gku518. Epub 2014 Jun 17.
Efficient protein synthesis in all organisms requires the post-transcriptional methylation of specific ribosomal ribonucleic acid (rRNA) and transfer RNA (tRNA) nucleotides. The methylation reactions are almost invariably catalyzed by enzymes that use S-adenosylmethionine (AdoMet) as the methyl group donor. One noteworthy exception is seen in some bacteria, where the conserved tRNA methylation at m5U54 is added by the enzyme TrmFO using flavin adenine dinucleotide together with N5,N10-methylenetetrahydrofolate as the one-carbon donor. The minimalist bacterium Mycoplasma capricolum possesses two homologs of trmFO, but surprisingly lacks the m5U54 tRNA modification. We created single and dual deletions of the trmFO homologs using a novel synthetic biology approach. Subsequent analysis of the M. capricolum RNAs by mass spectrometry shows that the TrmFO homolog encoded by Mcap0476 specifically modifies m5U1939 in 23S rRNA, a conserved methylation catalyzed by AdoMet-dependent enzymes in all other characterized bacteria. The Mcap0476 methyltransferase (renamed RlmFO) represents the first folate-dependent flavoprotein seen to modify ribosomal RNA.
所有生物体中高效的蛋白质合成需要特定核糖体核糖核酸（rRNA）和转运RNA（tRNA）核苷酸的转录后甲基化。甲基化反应几乎总是由以S-腺苷甲硫氨酸（AdoMet）作为甲基供体的酶催化。一个值得注意的例外是在一些细菌中，其中保守的tRNA在m5U54处的甲基化是由TrmFO酶使用黄素腺嘌呤二核苷酸以及N5,N10-亚甲基四氢叶酸作为一碳供体添加的。简约细菌山羊支原体拥有两个trmFO同源物，但令人惊讶的是缺乏m5U54 tRNA修饰。我们使用一种新型合成生物学方法创建了trmFO同源物的单缺失和双缺失。随后通过质谱对山羊支原体RNA进行分析表明，由Mcap0476编码的TrmFO同源物特异性修饰23S rRNA中的m5U1939，这是在所有其他已表征细菌中由AdoMet依赖性酶催化的保守甲基化。Mcap0476甲基转移酶（重新命名为RlmFO）代表了第一个被发现可修饰核糖体RNA的叶酸依赖性黄素蛋白。
