Sequence-specific and Shape-selective RNA Recognition by the Human RNA 5-Methylcytosine Methyltransferase NSun6
Sequence-specific and Shape-selective RNA Recognition by the Human RNA 5-Methylcytosine Methyltransferase NSun6
复制标题
人类 RNA 5-甲基胞嘧啶甲基转移酶 NSun6 的序列特异性和形状选择性 RNA 识别
DOI：
10.1074/jbc.m116.742569
复制
发表时间：
2016-11-11
影响因子：
4.8
通讯作者：
Wang, En-Duo
中科院分区：
文献类型：
--
作者：
Long, Tao;Li, Jing;Wang, En-Duo
人NSun 6是一种RNA甲基转移酶，催化甲基从S-腺苷-L-甲硫氨酸（SAM）转移到tRNA（Thr）和tRNA（Cys）的C72。在目前的研究中，我们使用质谱法证明了人类NSun 6确实如预期的那样将5-甲基胞嘧啶（m（5）C）引入tRNA。为了进一步揭示人NSun 6的tRNA识别机制，我们测定了人NSun 6对不同tRNA底物及其突变体的甲基化活性及其动力学参数。我们发现，人类NSun 6需要一个折叠良好的全长tRNA作为其底物。在受体区中，CCA末端、靶位点C72、C74碱基U 73以及受体茎的第二和第三碱基对（2：71和3：70）都是人NSun 6的重要RNA识别元件。此外，tRNA底物D-茎中的两个特异性碱基对（11：24和12：23）参与与人NSun 6的相互作用。总之，我们的研究结果表明，人类NSun 6依赖于一个微妙的RNA识别网络，该网络涉及tRNA底物的一级序列和三级结构。
Human NSun6 is an RNA methyltransferase that catalyzes the transfer of the methyl group from S-adenosyl-l-methionine (SAM) to C72 of tRNA(Thr) and tRNA(Cys). In the current study, we used mass spectrometry to demonstrate that human NSun6 indeed introduces 5-methylcytosine (m(5)C) into tRNA, as expected. To further reveal the tRNA recognition mechanism of human NSun6, we measured the methylation activity of human NSun6 and its kinetic parameters for different tRNA substrates and their mutants. We showed that human NSun6 requires a well folded, full-length tRNA as its substrate. In the acceptor region, the CCA terminus, the target site C72, the discriminator base U73, and the second and third base pairs (2:71 and 3:70) of the acceptor stem are all important RNA recognition elements for human NSun6. In addition, two specific base pairs (11:24 and 12:23) in the D-stem of the tRNA substrate are involved in interacting with human NSun6. Together, our findings suggest that human NSun6 relies on a delicate network for RNA recognition, which involves both the primary sequence and tertiary structure of tRNA substrates.