Nucleic Acids ReS:中科院生化细胞所王恩多研究组揭示人tRNA修饰酶
2017年5月22日，国际核酸类重要学术期刊《Nucleic Acids Research》杂志在线发表了中国科学院生物化学与细胞生物学研究所王恩多研究组题为“Structural basis for substrate binding and catalytic mechanism of a human RNA:m5C methyltransferase NSun6”的最新研究成果。刘如娟副研究员和博士研究生龙韬等人为论文第一作者，王恩多研究员为论文通讯作者。
5-甲基胞嘧啶(m5C)是广泛存在于DNA和RNA的一种核苷酸修饰。作为一种重要的表观遗传标记物，m5C修饰在DNA中已经被研究得很透彻;而RNA中的m5C修饰的研究则相对滞后。NSun甲基转移酶家族是真核生物中主要的RNA:m5C修饰酶，由七个成员组成，其多个成员的基因突变被发现与智力障碍、男性不育及癌症等人类疾病密切相关。然而，由于缺乏三级结构信息，NSun家族成员的RNA底物识别机制以及RNA:m5C的催化机制长期悬而未决。与此同时，RNA: m5C的生物学功能也有待阐明。人细胞质tRNA:m5C甲基转移酶NSun6(hNSun6)是2015年被鉴定出来的催化tRNA氨基酸接受茎上第72位m5C修饰的甲基转移酶。在之前的工作中，研究人员已经鉴定了hNSun6识别tRNA的元件，揭示了hNSun6是专一于tRNA的甲基修饰酶(Long et al, 2016, JBC)。
研究通过解析hNSun6与底物tRNA的共晶结构以及定点突变、ITC、结合迁移及酶学动力学等生物化学手段揭示了hNSun6的催化机理。研究发现hNSun6通过PUA结构域识别tRNA的CCA末端和D茎区域，通过MTase结构域精确识别C72和U73。这些识别作用使得底物tRNA发生了构象变化，从而使得目标碱基得以暴露并被催化。另外，结合生化实验还阐明了两个保守的半胱氨酸残基在催化中不同的角色。
A hNSun6与tRNA复合物结构 B复合物中tRNA(右)与经典tRNA(左)的构象比对 C和D hNSun6催化活性中心的详细结构
原文链接：
Structural basis for substrate binding and catalytic mechanism of a human RNA:m5C methyltransferase NSun6
原文摘要：
5-methylcytosine (m5C) modifications of RNA are ubiquitous in nature and play important roles in many biological processes such as protein translational regulation, RNA processing and stress response. Aberrant expressions of RNA:m5C methyltransferases are closely associated with various human diseases including cancers. However, no structural information for RNA-bound RNA:m5C methyltransferase was available until now, hindering elucidation of the catalytic mechanism behind RNA:m5C methylation. Here, we have solved the structures of NSun6, a human tRNA:m5C methyltransferase, in the apo form and in complex with a full-length tRNA substrate. These structures show a non-canonical conformation of the bound tRNA, rendering the base moiety of the target cytosine accessible to the enzyme for methylation. Further biochemical assays reveal the critical, but distinct, roles of two conserved cysteine residues for the RNA:m5C methylation. Collectively, for the first time, we have solved the complex structure of a RNA:m5C methyltransferase and addressed the catalytic mechanism of the RNA:m5C methyltransferase family, which may allow for structure-based drug design toward RNA:m5C methyltransferase–related diseases.
作者：王恩多
