ASSESSMENT OF THE COMPETITION BETWEEN TACRINE AND GALLAMINE FOR BINDING-SITES ON ACETYLCHOLINESTERASE

Gallamine binds to a peripheral anionic site on bovine erythrocyte acetylcholinesterase at low ionic strength and exerts allosteric effects on substrate hydrolysis (increase in maximum velocity), decarbamylation of carbamylated enzyme (increase in rate) and ageing of isopropylmethyl-phosphonylated enzyme (decrease in rate). Tacrine also inhibits ageing, but has no effect on decarbamylation and is a mixed competitive/noncompetitive inhibitor of substrate hydrolysis. Experiments were performed in order to determine if gallamine and tacrine compete for the same binding site. Kinetic parameters were determined in the presence of gallamine alone, tacrine alone, and mixtures of the two compounds in varying concentrations and ratios. Competition between edrophonium and tacrine or gallamine as inhibitors of substrate hydrolysis was also studied. The effects on substrate hydrolysis were complex, and departures from simple first-order Michaelis-Menten kinetics were observed. Further, analysis of the results by the median-effect technique indicated that the nature of the competition (mutually exclusive or non-exclusive) varied with the concentrations of substrate and inhibitors. Tacrine at relatively high concentrations antagonized the gallamine-induced acceleration of decarbamylation, but experiments at two concentrations of gallamine suggested that the antagonism was noncompetitive rather than competitive. The rate constant for ageing of a mixture of gallamine and tacrine was comparable with or lower than the lowest rate constant of either effector acting alone. Taken together, the results are inconsistent with competition between tacrine and gallamine for a single binding site on the enzyme, contrary to a proposal to this effect in the literature.