Spontaneously reactivation of acetylcholinesterase inhibited by diisopropylfluorophosphate

When electric eel acetylcholinesterase is inhibited by diisopropylfluorophosphate at 0 degree C most of the enzyme is irreversibly inactivated. However, 0.13--0.18% of the initial activity returns spontaneously after the unbound inhibitor is removed or when the inhibited enzyme is diluted into a large excess of competing substrate. A subsequent inhibition-reactivation cycle results in an essentially complete return of activity with minimal aging. The extent of aging increases substantially when inhibition and reactivation are performed at temperatures above 0 degree C. The free energy of activation for spontaneous reactivation was determined to be 20.1 kcal . mol-1. This large free energy of activation indicates that the reactivation process is a typical dephosphorylation reaction. The computer program used in determining the rate constants and the final extent of reactivation may be widely applicable in similar kinetic studies.