The peripheral anionic site on acetylcholinesterase (AChE), located at the active site gorge entry, encompasses overlapping binding sites for allosteric activators and inhibitors. Yet the molecular mechanisms coupling this site to the active center at the base of the gorge to modulate catalysis remain unclear. Crystal structures of mAChE bound with decidium, propidium and gallamine unveiled new determinants contributing to ligand interactions at the peripheral site. Subsequent studies using the syn and anti regioisomers of the click-chemistry inhibitor, TZ2PA6, that link propidium and tacrine moieties via distinctively substituted triazoles, revealed the inherent flexibility and a unique conformation of the peripheral site, along with substantial binding contributions from the triazoles with the Tyr337 region within the gorge. The recently solved structures of the mAChE mutant, Tyr337Ala, complexed with the TZ2PA6 isomers now reveals distinctive and time-dependent conformations of the complexes that are consistent with the triazole contribution to the energetics of inhibitor binding manifested in the respective dissociation rates of the complexes.