Tyrosine kinases are enzymes playing a critical role in cellular signaling. Mutations causing increased in kinase activity are often associated with cancer and various pathologies. One example in Src tyrosine kinases is offered by the substitution of the highly conserved tryptophan 260 by an alanine (W260A), which has been shown to cause an increase in activity. Here, molecular dynamics simulations based on atomic models are carried out to characterize the conformational changes in the linker region and the catalytic (kinase) domain of Src kinase to elucidate the impact of the W260A mutation. Umbrella sampling calculations show that the conformation of the linker observed in the assembled down-regulated state of the kinase is most favored when the kinase domain is in the inactive state, whereas the conformation of the linker observed in the re-assembled up-regulated state of the kinase is favored when the kinase domain is in the unphosphorylated active-like state. The calculations further indicate that there are only small differences between the WT and W260A mutant. In both cases, the intermediates states are very similar and the down-regulated inactive conformation is the most stable state. However, the calculations also show that the free energy cost to reach the unphosphorylated active-like conformation is slightly smaller for the W260A mutant compared with WT. A simple kinetic model is developed and submitted to a Bayesian Monte Carlo analysis to illustrate how such small differences can contribute to accelerate the trans-autophosphorylation reaction and yield a large increase in the activity of the mutant as observed experimentally.
Keywords: Src kinase; W260A mutation; autophosphorylation; umbrella sampling.
© 2015 The Protein Society.