Prolyl oligopeptidase (POP), a member of the prolyl endopeptidase family, is known to play a role in several neurological disorders. Its primary function is to cleave a wide range of small oligopeptides, including neuroactive peptides. We have used force biased molecular dynamics simulation to study the binding mechanism of POP. We examined three possible binding pathways using Steered Molecular Dynamics (SMD) and Umbrella Sampling (US) on a crystal structure of porcine POP with bound Z-pro-prolinal (ZPP). Using SMD, an exit pathway between the first and seventh blade of the β-propeller domain of POP was found to be a nonviable route. US on binding pathways through the β-propeller tunnel and the TYR190-GLN208 flexible loop at the interface between both POP domains allowed us to isolate the flexible loop pathway as the most probable. Further analysis of that pathway suggests a long-range covariation of the interdomain H-bond network, which indicates the possibility of large-scale domain reorientation observed in bacterial homologues and hypothesized to also occur in human POP.