Mechanism of MEK1 activation by phosphorylation

Abstract

Phosphorylation is the most common post-translational protein modification, often acting as the activator of protein function. In the case of MEK1, a member of the MAP kinase family, phosphorylation of both S218 and S222 are necessary for activation. Yet the molecular activation mechanism and its cooperative nature are poorly understood, especially due to the lack of experimental phosphorylated MEK1 structures. We performed molecular dynamics simulations to investigate the structural and dynamical consequences of single and double phosphorylation in MEK1. We find that successive phosphorylation progressively unwinds the helix containing the phosphorylation sites, thereby rotating the phosphate groups to directly interact with the catalytic site. Consequently, the solvent accessible surface area of the catalytic residues increase with phosphorylation. Yet, only in the double-phosphorylated state do all four critical catalytic residues become solvent exposed. By calculating the conformational entropy, we find that only in the double-phosphorylated state do all four catalytic residues have mutual information with each other, suggesting that phosphorylation also induces correlated motion at the active site. To validate our approach, we show that our simulations predict the alignment of the R-spine motif in the double-phosphorylated state, in agreement with the other kinases for which this alignment has been experimentally observed. These results show that phosphorylation can, via a partial unfolding mechanism, increase the solvent exposure of and dynamical coupling within the active site, both of which are critical for enzymatic activity.