Simulations of FGFR2 Kinase Activation Loop Dynamics and their Effects on Catalytic Activity

Abstract

Unregulated activity of tyrosine kinases (TKs) is responsible for numerous developmental, oncologic and musculoskeletal diseases, yet the regulatory mechanisms of TKs are still in need of elucidation. Fibroblast growth factor receptor (FGFR) kinases are a TK family whose catalytic activity appears to be regulated by phosphorylation of two adjacent tyrosine residues in the kinase “activation loop.” However, there is no clear mechanism to explain how tyrosine phosphorylation in the activation loop enables substrate catalysis. Using enhanced-sampling molecular dynamics (MD) simulations of FGFR2 kinase, we demonstrate by simulation that phosphorylation of Tyr657 in FGFR2 does not disable an inhibitory mechanism, as seen in other homologous structures such as FGFR1 kinase, but actively promotes substrate catalysis by altering the conformational dynamics of the kinase. Phosphorylation of Tyr657 allows for greater coordination of the kinase's N-lobe and C-lobe, facilitating enhanced coordination of the enzyme and substrate for catalysis. Moreover, inward motion of Tyr657, promoted by phosphorylation, stabilizes the presence of Arg664 in the enzyme's active site. We propose that the presence of Arg664 near ATP and the substrate tyrosine residue, previously observed in a crystal structure of FGFR3 kinase, locks the substrate tyrosine in a precise position, sandwiched by two arginine residues, favoring catalysis. Additionally, Arg664 helps to coordinate the position of ATP, and may also lower the activation energy for the catalytic phosphorylation reaction. Our study of activation loop dynamics in the pTyr657-in state of FGFR2 kinase may provide avenues for targeted therapies to counteract pathologic over-activation of FGFR2 kinase leading to uncontrolled signal transduction and uninhibited cell growth.