Structure of a TrmA-RNA complex:: A consensus RNA fold contributes to substrate selectivity and catalysis in m5U methyltransferases
Structure of a TrmA-RNA complex:: A consensus RNA fold contributes to substrate selectivity and catalysis in m5U methyltransferases
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DOI：
10.1073/pnas.0802247105
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发表时间：
2008-05-13
影响因子：
11.1
通讯作者：
Finer-Moore, Janet
中科院分区：
文献类型：
--
作者：
Alian, Akram;Lee, Tom T.;Finer-Moore, Janet
TrmA催化大多数tRNA中U 54的S-腺苷甲硫氨酸（S-腺苷甲硫氨酸）依赖性甲基化。我们解决了大肠杆菌5-甲基尿苷（（MU）-U-5）54 tRNA甲基转移酶（MTase）TrmA的结构在共价复合物与19-nt的T臂类似物10 2.4埃分辨率。将TrmA催化碱基Glu-358突变为Gln抑制了催化作用，并允许分离共价TrmA-RNA复合物用于结晶。蛋白质-RNA界面包括T环的6 nt和茎的两个近端碱基对。U 54从环中翻转出来进入活性位点。A58占据翻转的U 54的空间，并且是共线基础堆叠G53-A58-G57-C56-U 55的一部分。RNA折叠与未结合的tRNA或T臂类似物中的T环构象不同，但与结合在M5 U MTase鲁马的活性位点处的RNA环的折叠几乎相同。在这两种酶中，这种共有折叠将靶U和以下两个碱基呈递到蛋白质上的保守结合沟。在这个折叠之外，鲁马和TrmA底物具有完全不同的结构和蛋白质界面。环残基以外的目标U 54，使超过一半的氢键的蛋白质通过糖磷酸部分，占，部分原因是广泛的共识序列的TrmA底物。
TrmA catalyzes S-adenosylmethionine (AdoMet)-dependent methylation of U54 in most tRNAs. We solved the structure of the Escherichia coli 5-methyluridine ((MU)-U-5) 54 tRNA methyltransferase (MTase) TrmA in a covalent complex with a 19-nt T arm analog 10 2.4-angstrom resolution. Mutation of the TrmA catalytic base Glu-358 to Gin arrested catalysis and allowed isolation of the covalent TrmA-RNA complex for crystallization. The protein-RNA interface includes 6 nt of the T loop and two proximal base pairs of the stem. U54 is flipped out of the loop into the active site. A58 occupies the space of the everted U54 and is part of a collinear base stack G53-A58-G57-C56-U55. The RNA fold is different from T loop conformations in unbound tRNA or T arm analogs, but nearly identical to the fold of the RNA loop bound at the active site of the M5U MTase RumA. In both enzymes, this consensus fold presents the target U and the following two bases to a conserved binding groove on the protein. Outside of this fold, the RumA and TrmA substrates have completely different structures and protein interfaces. Loop residues other than the target U54 make more than half of their hydrogen bonds to the protein via sugar-phosphate moieties, accounting, in part, for the broad consensus sequence for TrmA substrates.