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.

Although the central α-helical Y(X)4LΦ motif (X, variable amino acid; Φ, 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.

Structural scaffold for eIF4E binding selectivity of 4E-BP isoforms: crystal structure of eIF4E binding region of 4E-BP2 and its comparison with that of 4E-BP1.

To clarify the higher eukaryotic initiation factor 4E (eIF4E) binding selectivity of 4E-binding protein 2 (4E-BP2) than of 4E-BP1, as determined by Trp fluorescence analysis, the crystal structure of the eIF4E binding region of 4E-BP2 in complex with m(7) GTP-bound human eIF4E has been determined by X-ray diffraction analysis and compared with that of 4E-BP1. The crystal structure revealed that the Pro47-Ser65 moiety of 4E-BP2 adopts a L-shaped conformation involving extended and α-helical structures and extends over the N-terminal loop and two different helix regions of eIF4E through hydrogen bonds, and electrostatic and hydrophobic interactions; these features were similarly observed for 4E-BP1. Although the pattern of the overall interaction of 4E-BP2 with eIF4E was similar to that of 4E-BP1, a notable difference was observed for the 60-63 sequence in relation to the conformation and binding selectivity of the 4E-BP isoform, i.