Structure and Binding Mode of a Ribosome Recycling Factor (RRF) from Mesophilic Bacterium* 210
Structure and Binding Mode of a Ribosome Recycling Factor (RRF) from Mesophilic Bacterium* 210
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嗜温细菌核糖体回收因子 (RRF) 的结构和结合模式* 210
DOI：
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发表时间：
2003
影响因子：
4.8
通讯作者：
Yuji Kobayashi
中科院分区：
文献类型：
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作者：
H. Nakano;Takuya Yoshida;S. Uchiyama;Masako Kawachi;Hitomi Matsuo;Takayuki Kato;A. Ohshima;Y. Yamaichi;T. Honda;H. Kato;Y. Yamagata;T. Ohkubo;Yuji Kobayashi
对嗜热菌核糖体循环因子（RRFs）的X射线和核磁共振分析表明，它们具有类似tRNA的L型构象，由两个结构域组成。从嗜温菌副溶血弧菌（Vibrio parahaemolyticus）中克隆了一个RRF，并对其进行了X射线分析。在大肠杆菌多聚核糖体和延伸因子G（EF-G）的体外测定系统中，该RRF被证明是有活性的。相反，上述来自嗜热细菌的RRF在这样的系统中是无活性的。对各种RRF结构中两个结构域之间的相对取向进行分析，包括来自嗜温细菌的RRF，揭示了结构域II围绕结构域I的螺旋束的长轴旋转。为了阐明RRF的核糖体结合位点，表达并表征了对应于RRF结构域I的肽片段（RRF-DI）。RRF-DI与70 S核糖体和50 S亚基的结合亲和力与野生型RRF相似。但它不与30 S亚基结合。这些发现促使我们重新研究RRF对tRNA的模仿的概念，并提出了一个新的模型，其中结构域I对应于tRNA的受体臂，结构域II对应于反密码子臂。然而，新模型与已发表的生物学发现更一致。
X-ray and NMR analyses on ribosome recycling factors (RRFs) from thermophilic bacteria showed that they display a tRNA-like L-shaped conformation consisting of two domains. Since then, it has been accepted that domain I, consisting of a three-helix bundle, corresponds to the anticodon arm of tRNA and domain II and a β/α/β sandwich structure, corresponds to the acceptor arm. In this study, we obtained a RRF from a mesophilic bacterium,Vibrio parahaemolyticus, by gene cloning and carried out an x-ray analysis on it at 2.2 Å resolution. This RRF was shown to be active in an in vitro assay system usingEscherichia coli polysomes and elongation factor G (EF-G). In contrast, the above-mentioned RRFs from thermophilic bacteria were inactive in such a system. Analysis of the relative orientations between the two domains in the structures of various RRFs, including this RRF from mesophilic bacterium, revealed that domain II rotates about the long axis of the helix bundle of domain I. To elucidate the ribosome binding site of RRF, the peptide fragment (RRF-DI) corresponding to domain I of RRF was expressed and characterized. RRF-DI is bound to 70 S ribosome and the 50 S subunit with an affinity similar to that of wild-type RRF. But it does not bind to the 30 S subunit. These findings caused us to reinvestigate the concept of the mimicry of RRF to tRNA and to propose a new model where domain I corresponds to the acceptor arm of tRNA and domain II corresponds to the anticodon arm. This is just the reverse of a model that is now widely accepted. However, the new model is in better agreement with published biological findings.