A conserved motif within the flexible C-terminus of the translational regulator 4E-BP is required for tight binding to the mRNA cap-binding protein eIF4E
A conserved motif within the flexible C-terminus of the translational regulator 4E-BP is required for tight binding to the mRNA cap-binding protein eIF4E
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DOI：
10.1042/bj20101481
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发表时间：
2012-01-01
影响因子：
4.1
通讯作者：
Tomoo, Koji
中科院分区：
文献类型：
--
作者：
Paku, Keum Soon;Umenaga, Yu;Tomoo, Koji
虽然翻译调节因子4 E-BP [eIF（真核起始因子）4 E结合蛋白]的中心α-螺旋Y（X）（4）L Phi基序（X，可变氨基酸; Phi，疏水性氨基酸）是mRNA帽结合蛋白eIF 4 E的核心结合区，但其N-和C-末端柔性区与eIF 4 E相互作用的功能仍有待阐明。为了确定C-末端区域在这种相互作用中的作用，通过SPR（表面等离子体共振）分析和ITC（等温滴定量热法）研究了4 E-BPn（n = 1-3）亚型的全长和连续C-末端缺失突变体的结合特征。因此，在C-末端区域内的保守PGVTS/T基序被证明作为第二结合区域，并在与eIF 4 E的紧密结合中发挥重要作用。4 E-BP亚型与eIF 4 E的结合常数在该保守区域存在的情况下比在该区域不存在的情况下增加了约1000倍。4 E-BP 1中该保守区域的连续缺失表明，瓦尔（81）的缺失导致4 E-BP结合能力的显著降低。分子动力学模拟表明，PGVTS/T保守区的功能类似于一种糊状物，通过疏水相互作用将eIF 4 E N-末端和4 E-BP C-末端柔性区的根部粘附在一起，其中缬氨酸位于两个柔性区的交叉位置。结论：4 E-BP柔性C端保守的PGVTS/T基序在增强eIF 4 E与核心Y（X）（4）L Phi基序的结合中起辅助但不可或缺的作用。
Although the central alpha-helical Y(X)(4)L Phi motif (X, variable amino acid; Phi, hydrophobic amino acid) of the translational regulator 4E-BP [eIF (eukaryotic initiation factor) 4E-binding protein] is the core binding region for the mRNA cap-binding protein eIF4E, the functions of its N- and C-terminal flexible regions for interaction with eIF4E remain to be elucidated. To identify the role for the C-terminal region in such an interaction, the binding features of full-length and sequential C-terminal deletion mutants of 4E-BPn (n = 1-3) subtypes were investigated by SPR (surface plasmon resonance) analysis and ITC (isothermal titration calorimetry). Consequently, the conserved PGVTS/T motif within the C-terminal region was shown to act as the second binding region and to play an important role in the tight binding to eIF4E. The 4E-BP subtypes increased the association constant with eIF4E by approximately 1000-fold in the presence of this conserved region compared with that in the absence of this region. The sequential deletion of this conserved region in 4E-BP1 showed that deletion of Val(81) leads to a considerable decrease in the binding ability of 4E-BP. Molecular dynamics simulation suggested that the conserved PGVTS/T region functions as a kind of paste, adhering the root of both the eIF4E N-terminal and 4E-BP C-terminal flexible regions through a hydrophobic interaction, where valine is located at the crossing position of both flexible regions. It is concluded that the conserved PGVTS/T motif within the flexible C-terminus of 4E-BP plays an auxiliary, but indispensable, role in strengthening the binding of eIF4E to the core Y(X)(4)L Phi motif.