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Updated: Jun 30, 2025
Monitoring eIF4F Assembly by Measuring eIF4E-eIF4G Interaction in Live Cells
Published on: May 1, 2020
The human eIF4E:4E-BP2 complex structure for studying hyperphosphorylation
Juan Zeng1, CuiMin Lu1, Xuan Huang1
1School of Biomedical Engineering, Guangdong Medical University, Dongguan 523808, China. azengjuan@gdmu.edu.cn.
Abstract:
The cap-dependent mRNA translation is dysregulated in many kinds of cancers. The interaction between eIF4E and eIF4G through a canonical eIF4E-binding motif (CEBM) determines the efficacy of the cap-dependent mRNA translation. eIF4E-binding proteins (4E-BPs) share the CEBM and compete with eIF4G for the same binding surface of eIF4E and then inhibit the mRNA translation. 4E-BPs function as tumor repressors in nature. Hyperphosphorylation of 4E-BPs regulates the structure folding and causes the dissociation of 4E-BPs from eIF4E. However, until now, there has been no structure of the full-length 4E-BPs in complex with eIF4E. The regulation mechanism of phosphorylation is still unclear. In this work, we first investigate the interactions of human eIF4E with the CEBM and an auxiliary eIF4E-binding motif (AEBM) in eIF4G and 4E-BPs. The results unravel that the structure and interactions of the CEBM are highly conserved between eIF4G and 4E-BPs. However, the extended CEBM (ECEBM) in 4E-BPs forms a longer helix than that in eIF4G. The residue R62 in the ECEBM of 4E-BP2 forms salt bridges with E32 and E70 of eIF4E. The residue R63 of 4E-BP2 forms two special hydrogen bonds with N77 of eIF4E. Both of these interactions are missing in eIF4G. The AEBM of 4E-BPs folds into a β-sheet conformation, which protects V81 inside a hydrophobic core in 4E-BP2. In eIF4G, the AEBM exists in a random coil state. The hydrophilic residues S637 and D638 of eIF4G open the hydrophobic core for solvents. The results show that the ECEBM and AEBM may be responsible for the competing advantage of 4E-BP2. Finally, based on our previous work (J. Zeng, F. Jiang and Y. D. Wu, J. Chem. Theory Comput., 2017, 13, 320), the human eIF4E:4E-BP2 complex (eIF4E:BP2P18-I88) including all reported phosphorylation sites is predicted. The eIF4E:BP2P18-I88 complex is different from the existing experimental eIF4E:eIF4G complex and provides an important structure for further studying the regulation mechanism of phosphorylation in 4E-BPs.
Insights
4E-binding proteins (4E-BPs) regulate mRNA translation by competing with eIF4G for eIF4E binding. This study reveals unique interactions of 4E-BP2 with eIF4E, explaining its tumor-suppressive role and providing a structural basis for phosphorylation regulation.
Area of Science:
- Molecular Biology
- Structural Biology
- Cancer Research
Background:
- Cap-dependent mRNA translation is crucial for protein synthesis and often dysregulated in cancers.
- eIF4E-binding proteins (4E-BPs) act as tumor suppressors by inhibiting translation, but their precise interaction mechanisms with eIF4E remain unclear.
- Understanding these interactions is vital for developing targeted cancer therapies.
Purpose of the Study:
- To investigate the structural basis of interactions between human eIF4E and the binding motifs of eIF4G and 4E-BPs.
- To elucidate the role of unique structural features in 4E-BP2 in its competitive advantage over eIF4G.
- To predict the structure of the full-length human eIF4E:4E-BP2 complex, including phosphorylation sites.
Main Methods:
- Comparative analysis of canonical (CEBM) and auxiliary (AEBM) eIF4E-binding motifs in eIF4G and 4E-BPs.
- Molecular modeling and structural prediction of protein complexes.
- Utilizing previous computational work to predict the eIF4E:4E-BP2 complex structure.
Main Results:
- The CEBM structures are conserved, but 4E-BP2's extended CEBM (ECEBM) forms a longer helix with unique salt bridges and hydrogen bonds to eIF4E.
- 4E-BP2's AEBM adopts a protective β-sheet conformation, unlike eIF4G's random coil, shielding hydrophobic residues.
- A predicted structure of the human eIF4E:4E-BP2 complex reveals differences from the eIF4E:eIF4G complex, offering insights into phosphorylation regulation.
Conclusions:
- The distinct structural features of 4E-BP2's ECEBM and AEBM contribute to its superior binding affinity to eIF4E.
- The predicted eIF4E:4E-BP2 complex structure provides a foundation for understanding phosphorylation-mediated regulation of translation inhibition.
- These findings have implications for cancer therapy by targeting the eIF4E-4E-BP interaction pathway.
