Insights into the Catalytic Mechanism of 16S rRNA Methyltransferase RsmE (m3U1498) from Crystal and Solution Structures
Insights into the Catalytic Mechanism of 16S rRNA Methyltransferase RsmE (m3U1498) from Crystal and Solution Structures
复制标题
从晶体和溶液结构洞察16S rRNA甲基转移酶RsmE (m3U1498)的催化机制
DOI：
10.1016/j.jmb.2012.08.016
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发表时间：
2012-11-02
影响因子：
5.6
通讯作者：
Dong, Yu-Hui
中科院分区：
文献类型：
--
作者：
Zhang, Heng;Wan, Hua;Dong, Yu-Hui
RsmE 是新 RNA 甲基转移酶 (MTase) 家族的创始成员，负责大肠杆菌 16S 核糖体 RNA 中 U1498 的甲基化。它在细菌和植物中非常保守，可能在核糖体亚基间通讯中发挥重要作用。单体的晶体结构表明它由两个不同但结构相关的结构域组成：PUA（假尿苷合酶和古氨酸特异性转糖基酶）样RNA识别和结合结构域以及具有深三叶结的保守MTase结构域。小角 X 射线散射数据分析表明，RsmE 形成灵活的二聚体构象，这对于底物结合可能至关重要。通过等温滴定量热法测定的 S-腺苷-L-甲硫氨酸 (AdoMet) 结合特征表明，同二聚体的亚基中仅结合了一个 AdoMet 分子。基于 RsmE AdoMet 尿苷酸复合物模型的突变体体外甲基化测定显示了参与底物结合和催化的关键残基。将RsmE与密切相关的MTase进行综合比较，并结合生化实验表明，二聚体RsmE中一个亚基的MTase结构域负责与一个AdoMet分子的结合和催化过程，而另一个亚基中的PUA样结构域主要负责对一个底物分子（核糖体RNA片段和核糖体蛋白复合物）的识别。甲基化过程是两个亚基协作所必需的，二聚化对于催化功能至关重要。总的来说，我们的研究提供了有关 RsmE 结构功能关系的新信息，从而提出了一种新的催化机制。 (C) 2012 Elsevier Ltd. 保留所有权利。
RsmE is the founding member of a new RNA methyltransferase (MTase) family responsible for methylation of U1498 in 16S ribosomal RNA in Escherichia coli. It is well conserved across bacteria and plants and may play an important role in ribosomal intersubunit communication. The crystal structure in monomer showed that it consists of two distinct but structurally related domains: the PUA (pseudouridine synthases and archaeosine-specific transglycosylases)-like RNA recognition and binding domain and the conserved MTase domain with a deep trefoil knot. Analysis of small-angle X-ray scattering data revealed that RsmE forms a flexible dimeric conformation that may be essential for substrate binding. The S-adenosyl-L-methionine (AdoMet)-binding characteristic determined by isothermal titration calorimetry suggested that there is only one AdoMet molecule bound in the subunit of the homodimer. In vitro methylation assay of the mutants based on the RsmE AdoMet uridylic acid complex model showed key residues involved in substrate binding and catalysis. Comprehensive comparisons of RsmE with closely related MTases, combined with the biochemical experiments, indicated that the MTase domain of one subunit in dimeric RsmE is responsible for binding of one AdoMet molecule and catalytic process while the PUA-like domain in the other subunit is mainly responsible for recognition of one substrate molecule (the ribosomal RNA fragment and ribosomal protein complex). The methylation process is required by collaboration of both subunits, and dimerization is functionally critical for catalysis. In general, our study provides new information on the structure function relationship of RsmE and thereby suggests a novel catalytic mechanism. (C) 2012 Elsevier Ltd. All rights reserved.