北海道支部について

The coelomic hemoglobin, dehaloperoxidase (DHP), from the sea worm Amphitrite

ornata is the first heme-containing globin possessing physiological peroxidase

enzymatic activity. DHP catalyzes the H2O2-dependent dehalogenation of

halophenols by O-atom addition to generate the corresponding quinones. As both

a dioxygen binding globin and a peroxidase, it is a dual function heme protein. With

two different starting oxidation states required for reversible O2 binding (ferrous)

and peroxidase activity (ferric), the question arises as to how DHP manages the two

functions. In our previous study, the co-presence of substrate 2,4,6-trichorophenol

(TCP) along with H2O2 has been found to be essential for the activation of

enzymatically inactive oxyferrous DHP (oxy-DHP). Based on that study, a functional

switching mechanism involving substrate (TCP•) radicals was proposed. To further

support this mechanism, we have further examined the H2O2-mediated conversion

of oxy-DHP to ferric state triggered by both biologically relevant [TCP, 4-

bromophenol (4-BP)] and non-relevant (ferrocyanide, etc.) compounds. We also find

that all of these conversion reactions are completely inhibited by the presence of

ferric heme ligands (KCN, imidazole), indicating an involvement of the ferric

oxidation state. Furthermore, the spin-trapping reagent 5,5-dimethyl-1-pyrroline-Noxide

(DMPO) effectively inhibits the TCP/4-BP (but not ferrocyanide)-triggered oxyto

ferric conversion of DHP. Taken together, the results demonstrate that substrate

TCP triggers the activation of oxy-DHP into a catalytically active peroxidase

through direct oxidation by TCP radicals that are generated and start to accumulate

upon H2O2-oxidation of TCP catalyzed by trace amounts of ferric DHP initially

present in the oxy-DHP sample. The data presented in this study further clarify the

mechanism of how halophenol substrate triggers the activation of hemoglobin DHP

into a peroxidase. (Funding: NSF MCB 0820456).