Kinetic analysis of calcium activation of brain acetylcholinesterase forms.
Calcium activation of acetylcholine hydrolysis by bovine brain acetylcholinesterase (Acetylcholine hydrolase, EC 3.1.1.7) forms has been analyzed in terms of changes in kinetic constants and thermodynamic activation parameters. De-acetylation was determined to be the major rate-influencing step in acetylcholine hydrolysis by both 60 000- and 240 000-dalton forms of the brain enzyme and 10 mM Ca2+ increased the rate constant for this step (k+3) by approximately 30% for both forms. For the smaller acetylcholinesterase form the effects of Ca2+ on de-acetylation was equivalent to its effect on the overall rate constant (k) and occurred without an effect on pK. In the case of the 240 000-dalton species, the overall rate constant was increased by Ca2+ by 33% at pH 8.0 and 81% at pH 7.25 and involved a pK shift of -0.2 pH units. For both enzyme forms the rate constants for acetylation (k+2) were increased by Ca2+. Thermodynamic analysis suggested that Ca2+ activation of the acetylation step was entropically driven. Differences between the two enzymes forms in terms of Ca2+ appear to result from association of low molecular weight species.
Kinetic analysis of calcium activation of brain acetylcholinesterase forms. Calcium activation of acetylcholine hydrolysis by bovine brain acetylcholinesterase (Acetylcholine hydrolase, EC 3.1.1.7) forms has been analyzed in terms of changes in kinetic constants and thermodynamic activation parameters. De-acetylation was determined to be the major rate-influencing step in acetylcholine hydrolysis by both 60 000- and 240 000-dalton forms of the brain enzyme and 10 mM Ca2+ increased the rate constant for this step (k+3) by approximately 30% for both forms. For the smaller acetylcholinesterase form the effects of Ca2+ on de-acetylation was equivalent to its effect on the overall rate constant (k) and occurred without an effect on pK. In the case of the 240 000-dalton species, the overall rate constant was increased by Ca2+ by 33% at pH 8.0 and 81% at pH 7.25 and involved a pK shift of -0.2 pH units. For both enzyme forms the rate constants for acetylation (k+2) were increased by Ca2+. Thermodynamic analysis suggested that Ca2+ activation of the acetylation step was entropically driven. Differences between the two enzymes forms in terms of Ca2+ appear to result from association of low molecular weight species.