Asymmetric co-evolution of the MEK-ERK binding interface

The highly conserved ERK regulates diverse cellular processes by phosphorylating a wide range of intracellular substrates. Its catalytic activity relies on phosphorylation by a single upstream kinase, MEK, which interacts with only a few binding partners. Here we test whether the asymmetry in protein-protein interaction (PPI) network architecture influences the co-evolution of the MEK-ERK complex. Phylogenetic sequence analysis across metazoan species revealed accelerated divergence in MEK's intrinsically disordered N-terminal docking motif (D-site), while ERK remained highly conserved. Structure prediction with AlphaFold2 and extensive molecular dynamics (MD) simulations showed that five conserved D-site residues form stable hydrophobic and electrostatic contacts with ERK's D-recruitment site (DRS). Functional assays in Drosophila melanogaster confirmed that these D-site interactions are essential for proper downstream signaling and support an allosteric role for this motif. Our results demonstrate that MEK uses a structurally simple yet evolutionarily adaptable motif to regulate MEK-ERK complex stability and binding dynamics. The D-site is strongly conserved within phylogenetic groups such as insects or terrestrial vertebrates, yet diverges across them, reflecting evolutionary pressures that balance functional conservation with signaling adaptability. The presented approach illustrates how the combined approach using sequencing data, molecular simulations and targeted perturbations can be used to address fundamental question about the evolution of PPI networks.