Single turnover studies of oxidative halophenol dehalogenation by horseradish peroxidase reveal a mechanism involving two consecutive one electron steps: Toward a functional halophenol bioremediation catalyst

Horseradish peroxidase (HRP) catalyzes the oxidative para-dechlorination of the environmental pollutant/carcinogen 2,4,6-trichlorophenol (2,4,6-TCP). A possible mechanism for this reaction is a direct oxygen atom transfer from HRP compound I (HRP I) to trichlorophenol to generate 2,6-dichloro 1,4-benzoquinone, a two-electron transfer process. An alternative mechanism involves two consecutive one-electron transfer steps in which HRP I is reduced to compound II (HRP II) and then to the ferric enzyme as first proposed by Wiese et al. [F.W. Wiese, H.C. Chang, R.V. Lloyd, J.P. Freeman, V.M. Samokyszyn, Arch. Environ. Contam. Toxicol. 34 (1998) 217–222]. To probe the mechanism of oxidative halophenol dehalogenation, the reactions between 2,4,6-TCP and HRP compounds I or II have been investigated under single turnover conditions (i.e., without excess H2O2) using rapid scan stopped-flow spectroscopy. Addition of 2,4,6-TCP to HRP I leads rapidly to HRP II and then more slowly to the ferric resting state, consistent with a mechanism involving two consecutive one-electron oxidations of the substrate via a phenoxy radical intermediate. HRP II can also directly dechlorinate 2,4,6-TCP as judged by rapid scan stopped-flow and mass spectrometry. This observation is particularly significant since HRP II can only carry out one-electron oxidations. A more detailed understanding of the mechanism of oxidative halophenol dehalogenation will facilitate the use of HRP as a halophenol bioremediation catalyst.

Graphical abstractSingle turnover experiments reveal that horseradish peroxidase (HRP) catalyzes the oxidative dehalogenation of halophenols by a mechanism involving two consecutive one-electron oxidations of the substrate via a phenoxy radical intermediate. Further development of HRP as a halophenol bioremediation catalyst will benefit from the more detailed understanding of this mechanism described in this paper.Figure optionsDownload full-size imageDownload as PowerPoint slideHighlights► The mechanism of halophenol dehalogenation by horseradish peroxidase is examined. ► Stopped-flow experiments reveal both Compounds I and II to be active catalysts. ► Single turnover data establish a mechanism involving two one-electron steps. ► Detailed mechanistic information will foster use of HRP as a bioremediation catalyst.