Crystal Structure of archaeal tRNA(m1G37)methyltransferase aTrm5
Crystal Structure of archaeal tRNA(m1G37)methyltransferase aTrm5
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DOI：
10.1002/prot.22019
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发表时间：
2008-09-01
影响因子：
2.9
通讯作者：
Yokoyama, Shigeyuki
中科院分区：
文献类型：
--
作者：
Goto-Ito, Sakurako;Ito, Takuhiro;Yokoyama, Shigeyuki
在tRNA的第37位（与反密码子相邻的第3（1）位），鸟苷的NI原子发生甲基化，生成修饰的核苷m（1）G37。在古细菌和真核生物中，m（1）G37的合成由tRNA（m（1）G37）甲基转移酶（古细菌或真核生物Trm 5，a/eTrm 5）催化。在这里，我们报告的晶体结构的古菌Trm 5（aTrm 5）从詹氏甲烷球菌（以前称为詹氏甲烷球菌）在复杂的甲基供体类似物在2.2埃分辨率。晶体结构显示，整个蛋白质由三个结构域D1、D2和D3组成。在a/eTrm 5一级结构中，D2和D3高度保守，而D1不保守。D3结构是Rossmann折叠，这是典型的I类甲基转移酶的标志。α/eTrm 5-定义结构域D2表现出与一些I类甲基转移酶的结构相似性。与此相反，DAM搜索与DI结构没有产生结构同源物。在晶体结构中，D3接触D1和D2两者。参与D1：D3相互作用的残基是不保守的，而参与D2：D3相互作用的残基是很保守的。D1和D2彼此不接触，并且它们之间的接头是无序的。以可溶形式制备对应于D1和D2-D3区的aTrm 5片段。DI片段的NMR分析显示DI本身折叠良好，并且它不与D2-D3片段或tRNA相互作用。D2-D3片段的NMR分析显示，它是良好折叠的，不依赖于DI，并且它与tRNA相互作用。此外，D2-D3片段与全长tRNA甲基化酶一样具有活性。D2-D3表面的正电荷可能参与tRNA结合。因此，这些发现表明D1和D3之间的相互作用不是持久的，并且D2-D3区域在tRNA甲基化中起主要作用。
Methylation of the NI atom of guanosine at position 37 in tRNA, the position 3(1)-adjacent to the anticodon, generates the modified nucleoside m(1)G37. In archaea and eukaryotes, m(1)G37 synthesis is catalyzed by tRNA(m(1)G37)methyltransferase (archaeal or eukaryotic Trm5, a/eTrm5). Here we report the crystal structure of archaeal Trm5 (aTrm5) from Methanocaldococcus jannaschii (formerly known as Methanococcus jannaschii) in complex with the methyl donor analogue at 2.2 angstrom resolution. The crystal structure revealed that the entire protein is composed of three structural domains, D1, D2, and D3. In the a/eTrm5 primary structures, D2 and D3 are highly conserved, while D1 is not conserved. The D3 structure is the Rossmann fold, which is the hallmark of the canonical class-I methyltransferases. The a/eTrm5-defining domain, D2, exhibits structural similarity to some class-I methyltransferases. In contrast, a DAM search with the DI structure yielded no structural homologues. In the crystal structure, D3 contacts both DI and D2. The residues involved in the DI:D3 interactions are not conserved, while those participating in the D2:D3 interactions are well conserved. DI and D2 do not contact each other, and the linker between them is disordered. aTrm5 fragments corresponding to the DI and D2-D3 regions were prepared in a soluble form. The NMR analysis of the DI fragment revealed that DI is well folded by itself, and it did not interact with either the D2-D3 fragment or the tRNA. The NMR analysis of the D2-D3 fragment revealed that it is well folded, independently of DI, and that it interacts with tRNA. Furthermore, the D2-D3 fragment was as active as the full-length enzyme for tRNA methylation. The positive charges on the surface of D2-D3 may be involved in tRNA binding. Therefore, these findings suggest that the interaction between DI and D3 is not persistent, and that the D2-D3 region plays the major role in tRNA methylation.