Structural basis for the enhancement of eIF4A helicase activity by eIF4G
Structural basis for the enhancement of eIF4A helicase activity by eIF4G
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DOI：
10.1101/gad.1335305
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发表时间：
2005-09-15
影响因子：
10.5
通讯作者：
Wagner, G
中科院分区：
文献类型：
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作者：
Oberer, M;Marintchev, A;Wagner, G
真核翻译起始因子4A （eIF4A）和4G （eIF4G）对翻译活性核糖体的组装至关重要。它们与eIF4E一起形成eIF4F复合物，将40S亚基招募到mRNA的5'端。双结构域RNA解旋酶eIF4A本身是一种非常弱的解旋酶，但与支架蛋白eIF4G相互作用后活性增强。本研究表明，尽管两个eIF4A结构域都在结合eIF4G的中间结构域（eIF4G-m，氨基酸745-1003）中发挥作用，但主要的相互作用表面位于c端结构域。我们使用核磁共振波谱来确定结合位点，并发现接触面邻近RNA-， ATP-和eif4a - ntd相互作用区域。界面残基的突变取消了结合，确认了界面，并表明eIF4G-m的n端与eIF4A的c端结构域相互作用。数据表明，eIF4G-m形成一个软钳来稳定eIF4A封闭的域间取向。该模型可以解释eIF4A的所有结合伙伴（eIF4G、RNA、ATP）之间的协同性以及eIF4F复合物中eIF4A活性的刺激。
The eukaryotic translation initiation factors 4A (eIF4A) and 4G (eIF4G) are crucial for the assembly of the translationally active ribosome. Together with eIF4E, they form the eIF4F complex, which recruits the 40S subunit to the 5' cap of mRNA. The two-domain RNA helicase eIF4A is a very weak helicase by itself, but the activity is enhanced upon interaction with the scaffolding protein eIF4G. Here we show that, albeit both eIF4A domains play a role in binding the middle domain of eIF4G (eIF4G-m, amino acids 745-1003), the main interaction surface is located on the C-terminal domain. We use NMR spectroscopy to define the binding site and find that the contact surface is adjacent to the RNA-, ATP-, and eIF4A-NTD-interacting regions.Mutations of interface residues abrogated binding, confirmed the interface, and showed that the N-terminal end of eIF4G-m interacts with the C-terminal domain of eIF4A. The data suggest that eIF4G-m forms a soft clamp to stabilize the closed interdomain orientation of eIF4A. This model can explain the cooperativity between all binding partners of eIF4A (eIF4G, RNA, ATP) and stimulation of eIF4A activity in the eIF4F complex.