The hydroperoxidase activity of deuteroferriheme: kinetic study of a peroxidase model

Abstract

Deuteroferriheme has been shown to display peroxidase-like activity. Its reaction with hypochlorite results in its oxidation to one or more reaction intermediates known as the "intermediate state" which is analogous to that obtained with peroxidase enzymes. The intermediate state is characterized by its spectrum which is identical to that obtained from heme oxidation by other oxidants such as peroxo acids, chlorite, iodosobenzene and iodobenzene diacetate. The stoichiometric ratio of deuteroferriheme oxidation is 2:1. This is interpreted as an aggregation of a two electron oxidized dfh monomer with an unoxidized dfh monomer. It was found that the rate constant (k(,m) = k'/(alpha)) for the reaction of monomeric heme with hypochlorite was independent of pH in the range of 6.99 - 8.77. Above pH 8.77 the value of k'/(alpha) increases. This is interpreted as a contribution of dimer reactivity. Rate constants for the oxidation of dimer, k(,d), were found to be directly proportional to the carbonate ion concentration suggesting general base catalysis. Decomposition of the dfh-OCl('-) derived intermediate was found to occur in a biphasic manner. This is indicative of either two parallel first-order reactions (UNFORMATTED TABLE FOLLOWS) k(,1) A (--->) C 1 k(,2) B (--->) C 2 or two first-order reactions in series k(,1) k(,2) A (--->) B (--->) C 3 (TABLE ENDS) Assuming k(,1) > k(,2), values of rate constants were found to be independent of concentrations of deuteroferriheme, hypochlorite and hydrogen ion (k(,1) = 6.65 x 10('-2)sec('-1) and k(,2) = 8.30 x 10('-3)sec('-1)). The reaction of dfh with Me(,3)NO is inversely proportional to hydrogen ion concentration and yields a peroxidatic intermediate which is spectrally similar to that of the dfh-OCl('-) derived inter- mediate. The intermediate was found to N-demethylate (CH(,3))(,3)N, (CH(,3))(,2)NH and (CH(,3))NH(,2) resulting in the formation of CH(,2)O.