<br>tRNA 修饰中新型 5-氨甲基-2-硫尿苷甲基转移酶的鉴定,Nucleic Acids Research

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tRNA 修饰中新型 5-氨甲基-2-硫尿苷甲基转移酶的鉴定
Nucleic Acids Research ( IF 16.6 ) Pub Date : 2023-02-10 , DOI: 10.1093/nar/gkad048
Gyuhyeok Cho ₁ , Jangmin Lee ₁ , Jungwook Kim ₁

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tRNA 34 位的尿苷能够与 mRNA 的 3' 端密码子碱基配对，通常会被修饰以影响翻译过程中解码特性的许多方面。 5-甲基尿苷 (xm5U) 的衍生物在尿嘧啶碱基的 C5 处包含甲基氨基甲基 (mnm-) 或羧甲基氨基甲基 (cmnm-) 基团，在许多原核 tRNA 的 34 位上广泛保守。在大肠杆菌等革兰氏阴性细菌中，双功能 MnmC 参与 mnm5(s2)U 生物合成的最后两个反应，其中该酶首先将 cmnm5(s2)U 转化为 5-氨甲基-(2-硫代) )尿苷(nm5(s2)U)，随后安装甲基以完成mnm5(s2)U的形成。尽管在革兰氏阳性细菌和植物的 tRNA 中也发现了 mnm5s2U，但它们的基因组不包含 mnmC 直向同源物，并且负责这种修饰的基因尚不清楚。我们通过比较基因组学、基因互补实验和体外测定发现，MnmM（以前称为 YtqB）是枯草芽孢杆菌中将 nm5s2U 转化为 mnm5s2U 的甲基转移酶。此外，我们还确定了与 tRNAGln 反密码子茎环复合的 MnmM 的 X 射线晶体结构。这些结构为 U33-nm5s2U34-U35 作为 MnmM 特异性的关键决定因素的重要性提供了分子基础。

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Identification of a novel 5-aminomethyl-2-thiouridine methyltransferase in tRNA modification

The uridine at the 34th position of tRNA, which is able to base pair with the 3′-end codon on mRNA, is usually modified to influence many aspects of decoding properties during translation. Derivatives of 5-methyluridine (xm5U), which include methylaminomethyl (mnm-) or carboxymethylaminomethyl (cmnm-) groups at C5 of uracil base, are widely conserved at the 34th position of many prokaryotic tRNAs. In Gram-negative bacteria such as Escherichia coli, a bifunctional MnmC is involved in the last two reactions of the biosynthesis of mnm5(s2)U, in which the enzyme first converts cmnm5(s2)U to 5-aminomethyl-(2-thio)uridine (nm5(s2)U) and subsequently installs the methyl group to complete the formation of mnm5(s2)U. Although mnm5s2U has been identified in tRNAs of Gram-positive bacteria and plants as well, their genomes do not contain an mnmC ortholog and the gene(s) responsible for this modification is unknown. We discovered that MnmM, previously known as YtqB, is the methyltransferase that converts nm5s2U to mnm5s2U in Bacillus subtilis through comparative genomics, gene complementation experiments, and in vitro assays. Furthermore, we determined X-ray crystal structures of MnmM complexed with anticodon stem loop of tRNAGln. The structures provide the molecular basis underlying the importance of U33-nm5s2U34-U35 as the key determinant for the specificity of MnmM.

更新日期：2023-02-10

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