Regulation of human eIF4E by 4E-BP1: binding analysis using surface plasmon resonance.
The interaction between recombinant human eukaryotic initiation factor (eIF)4E and recombinant glutathione S-transferase (GST)-fused human 4E-binding protein (BP)1 was analyzed by using the surface plasmon resonance method. The association rate of 4E-BP1 with eIF4E increased by about two orders of magnitude in the presence of m7GTP (a model compound of mRNA cap structure), but the dissociation rate was scarcely affected, indicating the cap-dependent binding of 4E-BP1 to eIF4E. On the other hand, phosphorylation of 4E-BP1 weakened its interaction with eIF4E whether m7GTP was present or not. However, phosphorylation of 4E-BP1 already associated with eIF4E or its m7GTP complex did not cause any change of the association, probably because of incomplete phosphorylation. This suggests that the regulation of eIF4E by 4E-BP phosphorylation is performed at its free state, not when it is in the associated state with eIF4E. Given these results, we discuss how 4E-BP regulates eIF4E at the first step of translation initiation.
Youtani
T
T
Department of Physical Chemistry, Osaka University of Pharmaceutical Sciences, Takatsuki, Japan.
Tomoo
K
K
Ishida
T
T
Miyoshi
H
H
Miura
K
K
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D039561:Eukaryotic Initiation Factor-4E; D005982:Glutathione Transferase / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D008954:Models, Biological; D010448:Peptide Initiation Factors / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020349:Surface Plasmon Resonance / Q000379:methods*; D013997:Time Factors
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; C087000:EIF4EBP1 protein, human; D039561:Eukaryotic Initiation Factor-4E; D010448:Peptide Initiation Factors; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; D006160:Guanosine Triphosphate; D005982:Glutathione Transferase