Investigating Effects of Phosphorylation on the Structure and Dynamics of WNK Kinase
by Using µs-long Molecular Dynamics Simulation

Nisha Amarnath Jonniya, Parimal Kar

Discipline of Biosciences and Biomedical Engineering, Indian Institute of Technology Indore, Khandwa Road, Indore 453552, Madhya Pradesh, India


WNK (With No Lysine) kinase is a serine/threonine kinase that plays a significant role in the regulation of cation chloride cotransporters located within a kidney and regulate blood pressure and body fluid homeostasis. It has been found that mutations in WNK kinase (WNK1 and WNK4) leads to Pseudohypoaldosteronism type II (PHA II), also known as Gordon’s syndrome. The characteristic feature of WNK kinase is the unique placement of catalytic lysine Lys-233 in β strand 2 of glycine-rich loop. Experimental evidence suggests that WNK1 kinase must be autophosphorylated at Ser382 found in the activation loop for its kinase activity. We have conducted all-atom molecular dynamics simulations of 1 µs to investigate the effect of phosphorylation on the structure and dynamics of the kinase in the apo and ligand-bound (WNK463) states. Our simulations reveal that phosphorylation and the binding of inhibitor stabilize the highly flexible activation loop as compared to its apo structure. The dynamic cross correlation analysis shows that phosphorylation in the activation loop influences the dynamic infrastructure and interfere with the long distance allosteric signaling that propagates throughout the whole molecule by diminishing the correlated motion and increasing the anti-corelated motion between the domains to occupy the inhibitors. In the case of phosphorylated complex, activation loop and αC helix lie farther away from the hinge region compared to the apo state to accomodate the inhibitor. Further analysis shows the formation of a distinct saltbridge between Glu-268 in αC helix and Arg348 of catalytic loop which has not been mentioned previously.This salt-bridge is found to be more stable in the case of phosphorylated WNK compared to unphosphorylated structure. We have also investigated the binding mechanism of WNK463 to WNK via molecular mechanics-Poisson-Boltzmann surface area (MM/PBSA) method. We shall discuss how different molecular forces compete with each other resulting in the complex formation. Overall, our study might provide an important information on the structural dynamics of WNK kinase, which will aid in the development of new inhibitor.