Archaeal NSUN6 catalyzes m5C72 modification on a wide-range of specific tRNAs
Archaeal NSUN6 catalyzes m5C72 modification on a wide-range of specific tRNAs
复制标题
古菌 NSUN6 催化多种特定 tRNA 上的 m(5)C72 修饰
DOI：
10.1093/nar/gky1236
复制
发表时间：
2019-02-28
影响因子：
14.9
通讯作者：
Liu, Ru-Juan
中科院分区：
文献类型：
--
作者：
Li, Jing;Li, Hao;Liu, Ru-Juan
摘要 人NOL1/NOP2/Sun RNA甲基转移酶家族成员6（hNSun6）在四种特定tRNA的C72处产生5-甲基胞嘧啶（m5C），其同源物仅存在于高等真核生物和超嗜热古细菌中。古菌 NSun6 同源物具有保守的催化残基，但其 RNA 识别基序与真核 NSun6 有明显差异。迄今为止，古细菌 NSun6 同源物的生化特性和功能尚不清楚。在古细菌 Pyrococcus horikoshii OT3 中，编码 NSun6 同源物的基因是 PH1991。我们证明PH1991蛋白可以在体外催化某些特定PhtRNA上m5C72的形成，因此被命名为PhNSun6。值得注意的是，PhNSun6 比 hNSun6 具有更广泛的 tRNA 底物范围，这归因于其 tRNA 底物特异性。利用生化和晶体学实验进一步阐明了该机制。从结构上来说，PhNSun6 中核苷酸 73 的结合口袋特异性地容纳含有 U73 或 G73 的 PhtRNA。此外，PhNSun6缺乏真核NSun6特异性的富含Lys的环，导致PhNSun6无法识别D-茎区。从功能上来说，m5C72修饰可以略微提高PhtRNA的热稳定性，但不影响PhtRNA的氨基酸接受活性。
Abstract Human NOL1/NOP2/Sun RNA methyltransferase family member 6 (hNSun6) generates 5-methylcytosine (m5C) at C72 of four specific tRNAs, and its homologs are present only in higher eukaryotes and hyperthermophilic archaea. Archaeal NSun6 homologs possess conserved catalytic residues, but have distinct differences in their RNA recognition motifs from eukaryotic NSun6s. Until now, the biochemical properties and functions of archaeal NSun6 homologs were unknown. In archaeon Pyrococcus horikoshii OT3, the gene encoding the NSun6 homolog is PH1991. We demonstrated that the PH1991 protein could catalyze m5C72 formation on some specific PhtRNAs in vitro and was thus named as PhNSun6. Remarkably, PhNSun6 has a much wider range of tRNA substrates than hNSun6, which was attributed to its tRNA substrate specificity. The mechanism was further elucidated using biochemical and crystallographic experiments. Structurally, the binding pocket for nucleotide 73 in PhNSun6 is specific to accommodate U73 or G73-containing PhtRNAs. Furthermore, PhNSun6 lacks the eukaryotic NSun6-specific Lys-rich loop, resulting in the non-recognition of D-stem region by PhNSun6. Functionally, the m5C72 modification could slightly promote the thermal stability of PhtRNAs, but did not affect the amino acid accepting activity of PhtRNAs.