Effect of N-terminal region of eIF4E and Ser65-phosphorylation of 4E-BP1 on interaction between eIF4E and 4E-BP1 fragment peptide.
To clarify the contribution of N-terminal region of eukaryotic initiation factor 4E (eIF4E) to the interaction with 4E-BP and to investigate the effect of 4E-BP phosphorylation on the interaction with eIF4E, the interaction profiles of the Ser65-unphosphorylated and phosphorylated peptides (Thr37-Thr70 fragment of 4E-BP1) with full-length and N-terminal 33 residues-deleted eIF4Es were investigated by fluorescence and SPR methods. The effect of N-terminal region of eIF4E on the interaction with 4E-BP1 peptides was shown to be dependent on the interaction state, that is, the steady-state fluorescence and kinetic-state SRP analyses showed the positive and negative contributions of the N-terminal region to the interaction with the peptide, respectively, despite its unphosphorylated or phosphorylated state. The comparison of the association constants of the peptide with those of full-length 4E-BP1 indicated the importance of N-terminal (1-36) and/or C-terminal (71-118) sequence of 4E-BP1 for the interaction, although the MD simulations suggested that the alpha-helical region (Arg56-Cys62) of 4E-BP1 peptide is sufficient for keeping the interaction. The MD simulations also indicated that a charge-dependent rigid hydration shell formed around the phosphate group makes the molecular conformation rigid, and single Ser65 phosphorylation is insufficient for releasing 4E-PB1 peptide from eIF4E.
Tomoo
Koji
K
Department of Physical Chemistry, Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki, Osaka 569-1094.
Abiko
Fumi
F
Miyagawa
Hiroo
H
Kitamura
Kunihiro
K
Ishida
Toshimasa
T
D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D001665:Binding Sites; D018797:Cell Cycle Proteins; D039561:Eukaryotic Initiation Factor-4E / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation
D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; C087000:EIF4EBP1 protein, human; D039561:Eukaryotic Initiation Factor-4E; D010750:Phosphoproteins