Substrate binding triggers a switch in the iron coordination in dehaloperoxidase from Amphitrite ornata:: HYSCORE experiments

Dehaloperoxidase (DHP) from the terebellid polychaete Amphitrite omata is the first known hemoglobin to exhibit efficient peroxidase activity in the oxidation of phenolic substrates. Hyperfine sublevel correlation spectroscopic (HYSCORE) analysis of the ferric form of DHP was carried out to characterize effects of the substrate 2,4,6-trifluorophenol (TFP) binding on the iron coordination in order to elucidate the molecular mechanism of the change in protein function from a globin to a peroxidase. Continuous wave EPR spectra show that heme iron of DHP at pH 6.0 exists in the high spin state. HYSCORE spectra recorded at magnetic field corresponding to g = 2 revealed the presence of exchangeable protons with hyperfine coupling of ca. 6 MHz, consistent with a water molecule being the sixth ligand in the iron coordination. These protons' spectral features disappeared upon substrate binding. This observation highlights the proposed role of the substrata as a trigger for the switch from hemoglobin to peroxidase function.