Structure of the eukaryotic translation initiation factor eIF4E in complex with 4EGI-1 reveals an allosteric mechanism for dissociating eIF4G

Evangelos Papadopoulos; Simon Jenni; Eihab Kabha; Khuloud J. Takrouri; Tingfang Yi; Nicola Salvi; Rafael E. Luna; Evripidis Gavathiotis; Poornachandran Mahalingam; Haribabu Arthanari; Ricard Rodriguez-Mias; Revital Yefidoff-Freedman; Bertal H. Aktas; Michael Chorev; Jose A. Halperin; Gerhard Wagner

doi:10.1073/pnas.1410250111

New Research In

Contributed by Gerhard Wagner, June 13, 2014 (sent for review March 29, 2014)

Significance

eIF4E is critical for protein synthesis and becomes hyperactive in cancer cells. Small-molecule inhibitors of the eIF4E/eIF4G initiation factor complex have recently been found to exhibit antitumor activity in vitro and in vivo. However, their mode of action at the atomic level has remained elusive. Here, we report high-resolution crystal structures of complexes of 4EGI-1 analogue inhibitors with eIF4E. We find that inhibition of eIF4G binding must be allosteric, because the 4EGI-1 and eIF4G bind at distant epitopes on eIF4E. Compound binding induces extension of an α-helix that stretches between the two binding sites. Indeed, mutations increasing helix propensity in this region reduce eIF4G affinity in the absence of the inhibitor, which is consistent with the proposed allosteric model.

Abstract

The interaction of the eukaryotic translation initiation factor eIF4E with the initiation factor eIF4G recruits the 40S ribosomal particle to the 5′ end of mRNAs, facilitates scanning to the AUG start codon, and is crucial for eukaryotic translation of nearly all genes. Efficient recruitment of the 40S particle is particularly important for translation of mRNAs encoding oncoproteins and growth-promoting factors, which often harbor complex 5′ UTRs and require efficient initiation. Thus, inhibiting the eIF4E/eIF4G interaction has emerged as a previously unpursued route for developing anticancer agents. Indeed, we discovered small-molecule inhibitors of this eIF4E/eIF4G interaction (4EGIs) that inhibit translation initiation both in vitro and in vivo and were used successfully in numerous cancer–biology and neurobiology studies. However, their detailed molecular mechanism of action has remained elusive. Here, we show that the eIF4E/eIF4G inhibitor 4EGI-1 acts allosterically by binding to a site on eIF4E distant from the eIF4G binding epitope. Data from NMR mapping and high-resolution crystal structures are congruent with this mechanism, where 4EGI-1 attaches to a hydrophobic pocket of eIF4E between β-sheet₂ (L₆₀-T₆₈) and α-helix₁ (E₆₉-N₇₇), causing localized conformational changes mainly in the H₇₈-L₈₅ region. It acts by unfolding a short 3₁₀-helix (S₈₂-L₈₅) while extending α-helix₁ by one turn (H₇₈-S₈₂). This unusual helix rearrangement has not been seen in any previous eIF4E structure and reveals elements of an allosteric inhibition mechanism leading to the dislocation of eIF4G from eIF4E.

Footnotes

↵¹To whom correspondence should be addressed. Email: gerhard_wagner{at}hms.harvard.edu.

Author contributions: E.P., B.H.A., M.C., J.A.H., and G.W. designed research; E.P., S.J., N.S., and R.R.-M. performed research; E.K., K.J.T., T.Y., R.E.L., E.G., P.M., H.A., and R.Y.-F. contributed new reagents/analytic tools; E.P., S.J., and E.G. analyzed data; and E.P., R.E.L., and G.W. wrote the paper.
The authors declare no conflict of interest.
Data deposition: The crystallography, atomic coordinates, and structure factors have been deposited in the Protein Data Bank, www.pdb.org (PDB ID codes 4TPW, 4TQB, and 4TQC).
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1410250111/-/DCSupplemental.

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