Solution Structure of a GSK 3$\beta$ Binding Motif, A $AXIN^{pep}$

Axin is a scaffold protein of the APC/axin/GSK complex, binding to all of the other signalling components. Axin interacts with Glycogen synthase kinase 3 (GSK 3) and functions as a negative regulator of Wnt signalling pathways. To determine the solution structure of the GSK3 binding regions of the axin, we initiated NMR study of axin fragment comprising residues 3using circular dichroism (CD) and two-dimensional NMR spectroscopy. The CD spectra of 3 in the presence of 30% TFE displayed a standard 3-helical conformation, exhibiting the bound structure of 3 to GSK3. On the basis of experimental restraints including , and coupling constants, the solution conformation of was determined with program CNS. The 20 lowest energy structures were selected out of 50 final simulated-annealing structures in both water and TFE environment, respectively. The for the 20 structures in TFE solution were 0.086 nm for backbone atoms and 0.195 nm for all heavy atoms, respectively. The Ramachandran plot indicates that the , angles of the 20 final structures is properly distributed in energetically acceptable regions. in aqueous solutions consists of a stable -helix spanning residues form to , which is an interacting motif with GSK3.