THE UNUSUAL REACTIVITIES OF AMPHIRITE-ORNATA DEHALOPEROXIDASE AND NOTOMASTUS-LOBATUS CHLOROPEROXIDASE DO NOT ARISE FROM A HISTIDINE IMIDAZOLATE PROXIMAL HEME IRON LIGAND

Notomastus lobatus chloroperoxidase (NCPO) and Amphitrite ornata dehal operoxidase (DHP) catalyze the halogenation of phenols and dehalogenat ion of halophenols, respectively. Both enzymes require peroxide for ac tivity and have recently been shown to contain histidine (His) as thei r proximal heme iron ligand, DHP is the only heme enzyme known to cata lyze peroxide-dependent defluorination reactions, and NCPO is the firs t His-Ligated heme-containing peroxidase capable of chlorinating halog en accepters using chloride as the halogen donor. To probe the roles o f the proximal His ligands in the mechanism of action of these two nov el peroxidases, they have been examined by resonance Raman and infrare d spectroscopy to determine the vibrational properties of their deoxyf errous and ferrous-CO derivatives. Previous studies of His-Ligated hem e proteins have revealed characteristic distinctions between dioxygen- binding globins and peroxidases. In the latter, the spectral propertie s are indicative of strong hydrogen bonding of the His imidazole N-1 h ydrogen, giving histidinate character to the His, whereas the globins are thought to contain a predominately neutral His. The partially anio nic peroxidase proximal His is thought to facilitate O-O bond cleavage of bound peroxide en route to the key oxo-ferryl compound I peroxidas e intermediate. Surprisingly, the vibrational data reported herein for DHP and NCPO are similar to those previously reported for neutral His /imidazole-ligated globins and heme complexes. The vibrational data in dicate that these enzymes do not activate bound peroxide through a mec hanism dependent on a push effect imparted by a partially ionized prox imal histidine as proposed for typical heme peroxidases.