Abstract

Tacrine (1,2,3,4-tetrahydro-9-aminoacridine) showed an apparent noncompetitive inhibition of Torpedo acetylcholinesterase (AChE) with a dissociation constant, Ki, of 8.5 nM. It altered the CD bands of AChE in the near-UV region, which monitor the local conformation of aromatic side groups, but not those in the far-UV region, which measure the secondary structure. An extrinsic CD band was induced at 348 nm, with a molar ellipticity of 35,000 deg cm2 dmol-1 (bases on tacrine), when each AChE subunit (Mr = 67,000) was saturated with one tacrine (mol/mol). With this band as a probe, the bound tacrine could be displaced by edrophonium or decamethonium, both of which are known to bind to the anionic site at the active center of AChE, but not by propidium, which binds to the peripheral site of the enzyme. Tacrine protected AChE from inactivation by diisopropylfluorophosphate (DFP). AChE completely lost its enzymatic activity when 1 mol of DFP was bound per mol of subunit upon incubation of 7 microM AChE (subunit) with 100 microM DFP for 40 min, but tacrine-treated AChE retained 60% of its activity and bound only 0.2 mol of DFP per mol of subunit under similar conditions. The corresponding CD, at 348 nm, of the AChE-tacrine-DFP complex increased or decreased gradually, depending on the order of addition of tacrine and DFP, and reached an equilibrium value (80% of its original) after 2 days. The difference absorption spectrum of the AChE-tacrine-DFP complex was the same as that of the AChE-tacrine complex. These results suggest that the protective effect of tacrine may be due to steric hindrance at the esteratic site of the enzyme.