title: 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.
abstract: 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. The 4E-BP subtypes increased the association constant with eIF4E by approximately 1000-fold in the presence of this conserved region compared with that in the absence of this region. The sequential deletion of this conserved region in 4E-BP1 showed that deletion of Val81 leads to a considerable decrease in the binding ability of 4E-BP. Molecular dynamics simulation suggested that the conserved PGVTS/T region functions as a kind of paste, adhering the root of both the eIF4E N-terminal and 4E-BP C-terminal flexible regions through a hydrophobic interaction, where valine is located at the crossing position of both flexible regions. It is concluded that the conserved PGVTS/T motif within the flexible C-terminus of 4E-BP plays an auxiliary, but indispensable, role in strengthening the binding of eIF4E to the core Y(X)4LΦ motif.
authors.affiliation: Department of Physical Chemistry, Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki, Osaka 569-1094, Japan.
authors.forename: Keum Soon
authors.initials: KS
authors.lastname: Paku
authors.forename: Yu
authors.initials: Y
authors.lastname: Umenaga
authors.forename: Tsunego
authors.initials: T
authors.lastname: Usui
authors.forename: Ai
authors.initials: A
authors.lastname: Fukuyo
authors.forename: Atsuo
authors.initials: A
authors.lastname: Mizuno
authors.forename: Yasuko
authors.initials: Y
authors.lastname: In
authors.forename: Toshimasa
authors.initials: T
authors.lastname: Ishida
authors.forename: Koji
authors.initials: K
authors.lastname: Tomoo
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D017124:Conserved Sequence; D004926:Escherichia coli; D039561:Eukaryotic Initiation Factor-4E / Q000235:genetics / Q000378:metabolism*; D039642:Eukaryotic Initiation Factors / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000502:physiology; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*
chemical_list: C104151:EIF4EBP2 protein, human; D039561:Eukaryotic Initiation Factor-4E; D039642:Eukaryotic Initiation Factors; D012333:RNA, Messenger