Catalytic Turnover of Substrate Benzylamines by the Quinone-Dependent Plasma Amine Oxidase Leads to H2O2-Dependent Inactivation: Evidence for Generation of a Cofactor-Derived Benzoxazole†

Incubation of bovine plasma amine oxidase (BPAO) with benzylamine and various p-substituted analogues results in a time-dependent inactivation that is attributable to buildup of the H2O2-turnover product on the basis of protection afforded by coincubation with catalase. The mechanism of inactivation is distinct from that effected by H2O2 itself, which requires higher concentrations. Solution studies using models for the 2,4,5-trihydroxyphenylalanine quinone (TPQ) cofactor reveal a loss of catalytic activity arising from oxidation of the dihydrobenzoxazole tautomer of the product Schiff base, that competes with hydrolytic release of benzaldehyde product. The resulting stable benzoxazole exhibits a characteristic absorption depending on the nature of the benzylamine p-substituent. For benzylamine itself, the model benzoxazole absorbs at 313 nm, in an area of strong absorption by the enzyme, whereas for 4-nitrobenzylamine, the absorption of the model benzoxazole is sufficiently red-shifted (at 365 nm) to be ...