The association and dissociation of proteins and ligands are crucial in biophysics for potential drug development [Baron and McCammon, Annu. Rev. Phys. Chem. 64, 151–175 (2013)]. However, identifying and characterizing the reaction pathways for these rare events has been a long-standing challenge. Molecular dynamics (MD) simulations are limited in exploring biophysical processes on experimental timescales, so ligand transport processes through complex transient tunnels formed by proteins during dynamics are often simulated using enhanced sampling MD [Rydzewski and Nowak, Phys. Life Rev. 22–23, 58–74 (2017)]. Erroneously identified ligand binding pathways can affect thermodynamic and kinetic characteristics calculated from MD trajectories. A system that has the potential to be a therapeutic target for neurodegenerative diseases is prolyl oligopeptidase (PREP). This is due to its involvement in promoting protein aggregation and disrupting cellular function through affecting protein–protein interactions (PPI). The recent discovery of a new type of PREP inhibitor that targets PPI raises important questions about the diversity of ligand binding pathways in PREP and their impact on protein dynamics [Pätsi et al., J. Med. Chem. 67, 5421–5436 (2024); Kilpeläinen et al., J. Med. Chem. 66, 7475–7496 (2023); and Walczewska-Szewc et al., Phys. Chem. Chem. Phys. 24, 4366–4373 (2022)]. In this article, using results from enhanced sampling MD, we visually present how the binding process in PREP depends on subtle changes in inhibitors, which could be crucial in treating neurodegenerative disorders.