Conformational flexibility of human casein kinase catalytic subunit explored by metadynamics

Abstract

Casein kinase CK2 is an essential enzyme in higher organisms, catalyzing the transfer of the γ phosphate from ATP to serine and threonine residues on protein substrates. In a number of animal tumors, CK2 activity has been shown to escape normal cellular control, making it a potential target for cancer therapy. Several crystal structures of human CK2 have been published with different conformations for the CK2α catalytic subunit. This variability reflects a high flexibility for two regions of CK2α: the interdomain hinge region, and the glycine-rich loop (p-loop). Here, we present a computational study simulating the equilibrium between three conformations involving these regions. Simulations were performed using well-tempered metadynamics combined with a path collective variables approach. This provides a reference pathway describing the conformational changes being studied, based on analysis of free energy surfaces. The free energies of the three conformations were found to be close and the paths proposed had low activation barriers. Our results indicate that these conformations can exist in water. This information should be useful when designing inhibitors specific to one conformation.

Figure 1

Overview of the flexible regions near the ATP binding site of the catalytic subunit of protein kinase in the totally active conformation (PDB: 2PVR). (a) Two conformations are superposed: in blue the collapsed p-loop, in green the stretched p-loop conformation. (b) The two conformations superposed are the closed and the open hinge region conformation respectively in blue and green. Figures were prepared with PYMOL (DeLano Scientific, San Carlos, CA, http://www.pymol.org). To see this figure in color, go online.

Figure 2

FES reconstructed as a function of s and z of the p-loop equilibrium. The energy separation between contours is 2 kcal/mol. Basins A–E are referring to the conformations in Fig. 3, a–e. To see this figure in color, go online.

Figure 3

(a–e) Steps of the p-loop collapse. Typical structure of basins A–E defined in Fig 2. Figures were prepared with PYMOL (DeLano Scientific, http://www.pymol.org). To see this figure in color, go online.

Figure 4

FES reconstructed as a function of s and z of the hinge region equilibrium. The energy separation between contours is 2 kcal/mol. Basins A–D are referring to the conformations below the FES. Figures were prepared with PYMOL (Delano Scientific, http://www.pymol.org). To see this figure in color, go online.

Scheme 1

Hypothetical equilibrium of CK2α proposed by Niefind et al. (9)