Visualization of two transfer RNAs trapped in transit during elongation factor G-mediated translocation
Visualization of two transfer RNAs trapped in transit during elongation factor G-mediated translocation
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DOI：
10.1073/pnas.1320387110
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发表时间：
2013-12-24
影响因子：
11.1
通讯作者：
Spahn, Christian M. T.
中科院分区：
文献类型：
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作者：
Ramrath, David J. F.;Lancaster, Laura;Spahn, Christian M. T.
在蛋白质合成过程中，信使RNA（mRNA）和转运RNA（tRNA）通过核糖体的偶联易位发生在每个肽键形成之后。该反应由核糖体复合物内的大规模构象变化促进，并由延伸因子G（EF-G）催化。以前的EF-G与核糖体相互作用的结构分析要么使用不含tRNA或仅含单个tRNA的模型复合物，要么使用EF-G直接结合于后易位状态的核糖体的复合物。在这里，我们提出了一个多粒子冷冻EM重建的易位中间含有两个tRNA被困在运输中，结合在嵌合intrasubunit ap/P和pe/E杂交状态。下游ap/P-tRNA与EF-G的结构域IV和30 S亚基体内的P位点元件接触，而上游pe/E-tRNA与旋转的30 S头部的P位点元件保持紧密的相互作用。值得注意的是，在这种状态下可以看到tRNA对的紧密压缩。这里提出的易位中间体代表了以前在理解易位机制中缺失的环节，揭示了核糖体使用两种不同的分子棘轮，涉及亚基内和亚基间的旋转运动，以驱动tRNA和mRNA的同步运动。
During protein synthesis, coupled translocation of messenger RNAs (mRNA) and transfer RNAs (tRNA) through the ribosome takes place following formation of each peptide bond. The reaction is facilitated by large-scale conformational changes within the ribosomal complex and catalyzed by elongtion factor G (EF-G). Previous structural analysis of the interaction of EF-G with the ribosome used either model complexes containing no tRNA or only a single tRNA, or complexes where EF-G was directly bound to ribosomes in the posttranslocational state. Here, we present a multiparticle cryo-EM reconstruction of a translocation intermediate containing two tRNAs trapped in transit, bound in chimeric intrasubunit ap/P and pe/E hybrid states. The downstream ap/P-tRNA is contacted by domain IV of EF-G and P-site elements within the 30S subunit body, whereas the upstream pe/E-tRNA maintains tight interactions with P-site elements of the swiveled 30S head. Remarkably, a tight compaction of the tRNA pair can be seen in this state. The translocational intermediate presented here represents a previously missing link in understanding the mechanism of translocation, revealing that the ribosome uses two distinct molecular ratchets, involving both intra-and intersubunit rotational movements, to drive the synchronous movement of tRNAs and mRNA.