AMINIUM CATION-RADICAL MECHANISM PROPOSED FOR MONOAMINE-OXIDASE-B CATALYSIS - ARE THERE ALTERNATIVES

1. The interaction of bovine liver mitochondrial monoamine oxidase B (MAO B) with a series of benzylamine analogues was investigated to provide mechanistic information relative to the proposed cation radical mechanism and to provide information on the structural requirements of the substrate binding site. 2. Steady-state kinetic analysis of MAO B with 11 ring-substituted benzylamine analogues showed substitution does not alter the reaction pathway. All amine analogues tested exhibit sizeable deuterium kinetic isotope effects. 3. Anaerobic stopped-flow kinetic studies showed (1) C-H bond cleavage is rate-limiting in enzyme-bound flavin reduction and (2) that no specially detectable flavin radicals are observed. 4. The binding affinity of para-substituted benzylamine analogues to MAO B increased as the hydrophobicity of the substituent increased. In contrast, meta-substitution of the ring showed reduced affinity with an increase in the van der Waals volume of the substituent. 5. The rate of enzyme reduction by para-substitution exhibited a strong negative dependence with the Taft (E(s)) steric value of the substituent. In contrast, the rate of enzyme reduction by meta-substituted benzylamines is independent of the nature of the substituent. 6. para-Substituted N,N-dimethylbenzylamine analogues are not substrates for MAO B but are competitive inhibitors of benzylamine oxidation with a weaker affinity with increasing van der Waals volume of the substituent. In contrast, meta-substituted N,N-dimethyl benzylamine analogues are weak substrates for MAO B with oxidation occurring exclusively at the benzyl carbon. 7. The consequences of these results on the possible mechanisms (aminium cation radical, H-. abstraction, and nucleophilic mechanism) for C-H bond cleavage proposed for MAO B are discussed.