Conformational ensembles of the disordered 4E‐BP2:eIF4E complex restrained by smFRET experiments
Abstract Eukaryotic cap‐dependent translation initiation is regulated by binding of the predominantly folded eukaryotic initiation factor 4E (eIF4E) to the intrinsically disordered eIF4E binding proteins (4E‐BPs). Here, we report full‐length atomistic conformational ensembles generated by IDPConformerGenerator and optimized by X‐EISDv2 workflow for both apo 4E‐BP2, the neuronal 4E‐BP, and 4E‐BP2 in complex with eIF4E, using data from single‐molecule fluorescence and nuclear magnetic resonance (NMR), together with select coordinates from a 4E‐BP1:eIF4E crystal structure. Structural sampling within dynamic complexes is often underappreciated, with NMR and crystal structure data for 4E‐BP:eIF4E suggesting different degrees of structural heterogeneity. Our ensemble models validated by solution spectroscopy data enable comparison of free 4E‐BP2 and its complex with eIF4E. This shows a delocalization of contacts around canonical regions, which supports previous findings of unidirectional conditional occupancy of the binding sites. Two new contact regions emerged: one between the disordered N‐termini of eIF4E and 4E‐BP2, which may play an allosteric role in tuning the binding affinity, and the other between the C‐terminus of 4E‐BP2 and an extended region of eIF4E, which is consistent with the extended, dynamic binding interface that we reported previously. These results support a model of translation regulation in which the dynamic 4E‐BP2:eIF4E complex facilitates accessibility of regulatory sites of 4E‐BP2 when bound.