GTP hydrolysis by EF-G synchronizes tRNA movement on small and large ribosomal subunits
GTP hydrolysis by EF-G synchronizes tRNA movement on small and large ribosomal subunits
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DOI：
10.1002/embj.201387465
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发表时间：
2014-05-02
期刊：
影响因子：
11.4
通讯作者：
Rodnina, Marina V.
中科院分区：
文献类型：
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作者：
Holtkamp, Wolf;Cunha, Carlos E.;Rodnina, Marina V.
延伸因子G（EF-G）在肽延伸的每个循环中促进两个tRNA和mRNA通过核糖体的运动。在易位过程中，tRNA暂时占据小（30 S）和大（50 S）核糖体亚基的中间位置。EF-G和GTP水解如何控制这些运动仍不清楚。我们使用荧光标记，专门监测运动的任一30 S或50 S亚基结合EF-G突变体和易位特异性抗生素，以调查时间和能量的易位。我们表明，EF-G-GTP促进同步运动的肽基-tRNA的两个亚基进入早期易位后的状态，这类似于嵌合状态的结构研究确定。没有GTP水解的EF-G结合仅促进50 S亚基上的部分tRNA移动。然而，快速的30 S易位和伴随的50 S易位的完成需要GTP水解和EF-G的功能结构域4。我们的研究结果揭示了两种不同的模式，利用EF-G结合和GTP水解的能量，并建议耦合GTP水解易位介导的30 S亚基的重排。
Elongation factor G (EF-G) promotes the movement of two tRNAs and the mRNA through the ribosome in each cycle of peptide elongation. During translocation, the tRNAs transiently occupy intermediate positions on both small (30S) and large (50S) ribosomal subunits. How EF-G and GTP hydrolysis control these movements is still unclear. We used fluorescence labels that specifically monitor movements on either 30S or 50S subunits in combination with EF-G mutants and translocation-specific antibiotics to investigate timing and energetics of translocation. We show that EF-G-GTP facilitates synchronous movements of peptidyl-tRNA on the two subunits into an early post-translocation state, which resembles a chimeric state identified by structural studies. EF-G binding without GTP hydrolysis promotes only partial tRNA movement on the 50S subunit. However, rapid 30S translocation and the concomitant completion of 50S translocation require GTP hydrolysis and a functional domain 4 of EF-G. Our results reveal two distinct modes for utilizing the energy of EF-G binding and GTP hydrolysis and suggest that coupling of GTP hydrolysis to translocation is mediated through rearrangements of the 30S subunit.