An unexpected switch in peptide binding mode: from simulation to substrate specificity

A ten microsecond molecular dynamics simulation of a kallikrein-related peptidase 7 peptide complex revealed an unexpected change in binding mode. After more than two microseconds unrestrained sampling we observe a spontaneous transition of the binding pose including a 180 degrees rotation around the P1 residue. Subsequently, the substrate peptide occupies the prime side region rather than the cognate non-prime side in a stable conformation. We characterize the unexpected binding mode in terms of contacts, solvent-accessible surface area, molecular interactions and energetic properties. We compare the new pose to inhibitor-bound structures of kallikreins with occupied prime side and find that a similar orientation is adopted. Finally, we apply in silico mutagenesis based on the alternative peptide binding position to explore the prime side specificity of kallikrein-related peptidase 7 and compare it to available experimental data. Our study provides the first microsecond time scale simulation data on a kallikrein protease and shows previously unexplored prime side interactions. Therefore, we expect our study to advance the rational design of inhibitors targeting kallikrein-related peptidase 7, an emerging drug target involved in several skin diseases as well as cancer.