Anatomy of a structural pathway for activation of the catalytic domain of Src kinase Hck.

Src kinase activity is implicated in the regulation of downstream signal transduction pathways involved in cell growth processes. Crystallographic studies indicate that activation of Hematopoietic cell kinase (Hck), a member of the Src kinase family, is accompanied structurally by a large conformational change in two specific parts of its catalytic domain: the alpha-C helix and the activation loop. In the present study, molecular dynamics (MD) simulations are used to characterize the transformation pathway from the inactive to the active state. Four different conditions are considered: the presence or absence of Tyr416 phosphorylation in the activation loop, and the presence or absence of substrate ATP-2Mg(+2) in the active site. Effective free energy landscapes for local residues are determined using a combination of restrained MD simulations with a Root Mean Square Distance (RMSD) biasing potential to enforce the change followed by free MD simulations to allow relaxation from artificially enforced intermediates. A conceptual subdivision of the kinase catalytic domain into four moving parts: the flexible activation loop segment, the buried activation loop segment, the alpha-C helix, and the N-terminal end linker, leads to a concise hypothesis in which each of the moving parts are only required to be coupled to their nearest neighbor to ensure bidirectional allostery in the regulation of protein tyrosine kinases. Both Tyr416 phosphorylation and ATP-2Mg(+2) affect the local backbone torsional free energy landscapes accompanying the structural transition. When these two factors are present together, a metastable coordinated state of ATP-2Mg(+2) and the phosphorylated Tyr416 is observed that offers a possible explanation for the inhibition of protein kinase activity due to increase in Mg(+2) ion concentration.