Protein kinase monopolar spindle 1 plays an important role in spindle assembly 
checkpoint at the onset of mitosis. Over expression of MPS1 correlated with a 
wide range of human tumors makes it an attractive target for finding an 
effective and specific inhibitor. In this work, we performed molecular dynamics 
simulations of protein MPS1 itself as well as protein bound systems with the 
inhibitor and natural substrate based on crystal structures. The reported orally 
bioavailable 1 h-pyrrolo [3,2-c] pyridine inhibitors of MPS1 maintained stable 
binding in the catalytic site, while natural substrate ATP could not stay. 
Comparative study of stability and flexibility of three systems reveals position 
shifting of β-sheet region within the catalytic site, which indicates inhibition 
mechanism was through stabilizing the β-sheet region. Binding free energies 
calculated with MM-GB/PBSA method shows different binding affinity for inhibitor 
and ATP. Finally, interactions between protein and inhibitor during molecular 
dynamic simulations were measured and counted. Residue Gly605 and Leu654 were 
suggested as important hot spots for stable binding of inhibitor by molecular 
dynamic simulation. Our results reveal an important position shifting within 
catalytic site for non-inhibited proteins. Together with hot spots found by 
molecular dynamic simulation, the results provide important information of 
inhibition mechanism and will be referenced for designing novel inhibitors.