Crystal Structure of the Human tRNA m(1)A58 Methyltransferase-tRNA(3)(Lys) Complex: Refolding of Substrate tRNA Allows Access to the Methylation Target.
Crystal Structure of the Human tRNA m(1)A58 Methyltransferase-tRNA(3)(Lys) Complex: Refolding of Substrate tRNA Allows Access to the Methylation Target.
复制标题
DOI：
10.1016/j.jmb.2015.10.005
复制
发表时间：
2015-12-04
影响因子：
5.6
通讯作者：
Stroud RM
中科院分区：
文献类型：
--
作者：
Finer-Moore J;Czudnochowski N;O'Connell JD 3rd;Wang AL;Stroud RM
人tRNA 3 Lys是HIV逆转录的引物，3′端与HIV RNA上的引物结合位点互补。互补性终止于第18个碱基A58，其在tRNA 3 Lys中被修饰以去除Watson-Crick配对。出于测试修饰在终止引物结合序列从而限制连续转录中的作用的动机，我们询问了RNA的修饰如何完成。tRNA m1 A58甲基转移酶（m1 A58 MTase）从辅因子S-腺苷-L-甲硫氨酸甲基化A58的N1，其被埋在tRNA的T β C环中。这种保守的tRNA修饰对于酿酒酵母中起始tRNA的稳定性是必不可少的。本文报道了人tRNA m1 A58 MTase与人tRNA 3 Lys和产物S-腺苷-L-高半胱氨酸复合的三种结构，显示了异二聚体的二聚体，其中每个异二聚体包含催化链Trm 61和同源但非催化链Trm 6，其被重新用作反式作用的tRNA结合亚基; tRNA通过二聚体界面结合，使得来自相对异二聚体的Trm 6将A58带入Trm 61的活性位点。T环和D环分开，显示了通常埋在tRNA中的A58是如何被修饰的。这一结果对我们理解折叠tRNA内部位点的修饰机制具有广泛的影响。这些结构可作为设计抑制剂的模板，这些抑制剂可用于测试tRNA m1 A58 MTase对逆转录病毒引发和转录的影响。
Human tRNA3 Lys is the primer for reverse transcription of HIV; the 3′ end is complementary to the primer-binding site on HIV RNA. The complementarity ends at the 18th base, A58, which in tRNA3 Lys is modified to remove Watson–Crick pairing. Motivated to test the role of the modification in terminating the primer-binding sequence and thus limiting run-on transcription, we asked how the modification of RNA could be accomplished. tRNA m1A58 methyltransferase (m1A58 MTase) methylates N1 of A58, which is buried in the TψC-loop of tRNA, from cofactor S-adenosyl-L-methionine. This conserved tRNA modification is essential for stability of initiator tRNA in Saccharomyces cerevisiae. Reported here, three structures of human tRNA m1A58 MTase in complex with human tRNA3 Lys and the product S-adenosyl-L-homocysteine show a dimer of heterodimers in which each heterodimer comprises a catalytic chain, Trm61, and a homologous but noncatalytic chain, Trm6, repurposed as a tRNA-binding subunit that acts in trans; tRNAs bind across the dimer interface such that Trm6 from the opposing heterodimer brings A58 in to the active site of Trm61. T-loop and D-loop are splayed apart showing how A58, normally buried in tRNA, becomes accessible for modification. This result has broad impact on our understanding of the mechanisms of modifying internal sites in folded tRNA. The structures serve as templates for design of inhibitors that could be used to test tRNA m1A58 MTase's impact on retroviral priming and transcription.