Abstract
The properties of the nonheme iron of bromoperoxidase from Corallina pilulifera were studied. The enzyme lost its activity when reduced with formamidine-sulfinic acid and recovered it when oxidized by air. Incubation of the enzyme with ferric or ferrous ion-chelating agents indicated that its nonheme iron was ferric. Analyses of circular dichroism and proton NMR spectra suggested that the ferric ion tightly bound to cysteine, histidine, or tyrosine residues of the enzyme. The enzyme catalyzed Br--dependent catalase reactions to yield 1 mol of O2 from 2 mol of H2O2. No O2 evolution was observed when bromination reaction of monochlorodimedone occurred. From these results, together with previous knowledge of this enzyme, it was concluded that it activated bromide anion (Br-) to bromonium cation (Br+) using one molecule of H2O2, and this Br+OH- formed at the active site then decomposed another H2O2 to yield O2 in the absence of halogen acceptors (substrate). When substrate was present in the reaction mixture, it and H2O2 competitively reacted with the reaction intermediate (Br+OH-) to give brominated products.
Highlights
The detailed properties of the enzyme of C. pilulifera were reported in previous papers [6, 7], but those of the enzyme's nonheme iron were not clear
Enzyme Assay-Bromoperoxidase activity was assayed by measurement of the change in absorbance at 290nm caused by the conversion of monochlorodimedone to monobromomonochlorodimedone [4].' Catalase activity was measured by the modified method of Beers and Sizer [17], whose detailed reaction conditions were described in the previous paper [4]
Bromoperoxidase ofC. pilulifera is an interesting haloperedition of the Journal that is available from Waverly Press
Summary
0 1987 by The American Society for Biochemistry and Molecular Biology, Inc. Vol 262, No 25, Issue of September 5, pp. 11982-11987.1987 Printed in U.S.A. 11982-11987.1987 Printed in U.S.A. Characterization of Nonheme Iron and Reaction Mechanisomf Bromoperoxidase in Corallina pilulifera". No O2 evolution was observed when bromination reaction of monochlorodimedoneoccurred From these results, together with previous knowledge of this enzyme, it was concluded that it activated bromide anion (Br-) to bromonium cation (Br+) using onemolecule of H202a,nd this Br+OH-formed at the active sitethen decomposed another H20z toyield O2in the absence of halogen acceptors (substrate). Bromination by the H-type chloroperoxidase of the fungus C. fumago is caused by the release of molecular halogen into the reaction mixture [10, 14, 15]. This work describes the halogenation mechanism of nonheme bromoperoxidase of C. pilulifera and clarifies the differences between the reaction mechanisms of heme and nonheme haloperoxidases
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