Subsequently, 4 moved closer to Phe461 and Thr471 (Figure ), resulting in unbinding through the internal path. The side chain of Phe461 sterically prevents 4 from exiting the binding site, thus an additional rotation of the Phe461 side chain is required for complete ligand unbinding (Figure ). The results show that the neutral and cationic antagonists have a different behavior, with the cationic antagonists 2 and 3 unbinding through the internal path only, and the neutral antagonists 4, 5 and 6 unbinding through both paths, because 2 and 3 form an ionic interaction with Glu427, which prevents the side chain of Glu427 from forming the hydrogen bond with the side chain of Tyr567* observed in crystal structures with neutral antagonists 4, 5 and 6. The lack of this hydrogen bond facilitates the exit of cationic antagonists through the internal path, while the neutral antagonists can undergo both possible unbinding paths.