Two enzymes bound to one transfer RNA assume alternative conformations for consecutive reactions
Two enzymes bound to one transfer RNA assume alternative conformations for consecutive reactions
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DOI：
10.1038/nature09411
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发表时间：
2010-09-01
期刊：
影响因子：
64.8
通讯作者：
Yokoyama, Shigeyuki
中科院分区：
文献类型：
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作者：
Ito, Takuhiro;Yokoyama, Shigeyuki
在大多数细菌和所有古细菌中，谷氨酰胺-tRNA合成酶（GluRS）将tRNA（Glu）和tRNA（Gln）同时谷氨酰胺化，然后Glu-tRNA（Gln）被tRNA依赖的氨基转移酶选择性地转化为Gln-tRNA(Gln)（1,2）。这两种酶识别其底物tRNA的机制，以及它们如何在Gln-tRNA（Gln）合成中相互合作，仍有待确定。本文报道了由tRNA（Gln）、GluRS和异三聚体氨基转移酶GatCAB组成的细菌Thermotoga maritima的“谷氨酰胺转酶体”的形成及其3.35埃分辨率下的晶体结构。GluRS的反密码子结合体识别tRNA（Gln）和tRNA（Glu）的共同特征，而GatCAB的尾体以tRNA（Gln）特异性的方式识别l形tRNA（Gln）的外角。GluRS处于生产形式，因为它的催化体与tRNA（Gln）的氨基酸受体臂结合。相比之下，GatCAB则是非生产形式：GatCAB的催化体与GluRS的催化体接触，位于tRNA（Gln）受体干附近，在合适的位置等待GluRS完成Glu-tRNA（Gln）的形成。我们发现了GluRS的催化体和反密码子结合体之间以及GatCAB的催化体和尾体之间的铰链，这使得GluRS和GatCAB都可以采用生产性和非生产性形式。这两种酶的催化体竞争tRNA（Gln）的受体臂，因此不能同时呈现它们的生产形式。从目前的谷氨酰化状态（具有生产性GluRS和非生产性GatCAB）过渡到假定的酰胺化状态（具有非生产性GluRS和生产性GatCAB），由于空间原因，需要两种酶处于非生产性形式的中间状态。所提出的机制解释了转胺体如何高效地完成Gln- trna （Gln）形成的两个连续步骤，同时释放不稳定的中间Glu-tRNA（Gln）的风险很低。
In most bacteria and all archaea, glutamyl-tRNA synthetase (GluRS) glutamylates both tRNA(Glu) and tRNA(Gln), and then Glu-tRNA(Gln) is selectively converted to Gln-tRNA(Gln) by a tRNAdependent amidotransferase(1,2). The mechanisms by which the two enzymes recognize their substrate tRNA(s), and how they cooperate with each other in Gln-tRNA(Gln) synthesis, remain to be determined. Here we report the formation of the 'glutamine transamidosome' from the bacterium Thermotoga maritima, consisting of tRNA(Gln), GluRS and the heterotrimeric amidotransferase GatCAB, and its crystal structure at 3.35 angstrom resolution. The anticodon-binding body of GluRS recognizes the common features of tRNA(Gln) and tRNA(Glu), whereas the tail body of GatCAB recognizes the outer corner of the L-shaped tRNA(Gln) in a tRNA(Gln)-specific manner. GluRS is in the productive form, as its catalytic body binds to the amino-acid-acceptor arm of tRNA(Gln). In contrast, GatCAB is in the non-productive form: the catalytic body of GatCAB contacts that of GluRS and is located near the acceptor stem of tRNA(Gln), in an appropriate site to wait for the completion of Glu-tRNA(Gln) formation by GluRS. We identified the hinges between the catalytic and anticodon-binding bodies of GluRS and between the catalytic and tail bodies of GatCAB, which allow both GluRS and GatCAB to adopt the productive and non-productive forms. The catalytic bodies of the two enzymes compete for the acceptor arm of tRNA(Gln) and therefore cannot assume their productive forms simultaneously. The transition from the present glutamylation state, with the productive GluRS and the non-productive GatCAB, to the putative amidation state, with the nonproductive GluRS and the productive GatCAB, requires an intermediate state with the two enzymes in their non-productive forms, for steric reasons. The proposed mechanism explains how the transamidosome efficiently performs the two consecutive steps of Gln-tRNA(Gln) formation, with a low risk of releasing the unstable intermediate Glu-tRNA(Gln).