Prosthetic heme modification during halide ion oxidation. Demonstration of chloride oxidation by horseradish peroxidase

摘要

Myeloperoxidase (MPO), eosinophil peroxidase (EPO), and chloroperoxidase can oxidize iodide, bromide, and chloride, but most peroxidases, including the prototypical horseradish peroxidase (HRP), reportedly only oxidize iodide and, in some cases, bromide. We report here that incubation of HRP with Br- and H2O2 at acidic pH results in both bromination of monochlorodimedone and modification of the heme group. Mass spectrometry indicates that the heme 2- and 4-vinyl groups are modified by either replacement of a vinyl hydrogen by a bromide or addition of HOBr to give a bromohydrin. These reactions do not occur if protein-free heme and Br- are co-incubated with H2O2 or if the HRP reaction is carried out at pH 7. Surprisingly, similar prosthetic heme modifications occur in incubations of HRP with H2O2 and Cl-. A mechanism is proposed involving oxidation of Br- or Cl- to give HOBr or HOCl, respectively, followed by addition to a vinyl group. In the reaction with Cl-, a meso-chloro heme adduct is also formed. This first demonstration of Cl- oxidation by HRP, and the finding that prosthetic heme modification occurs when Br- or Cl- is oxidized in the absence of a cosubstrate, show that only modest tuning is required to achieve the unique chloride oxidation activity of MPO and EPO. The results raise the question of how the prosthetic hemes of MPO and EPO, whose function is to produce oxidized halide species, escape modification.