Reversible formation of high-valent-iron-oxo-porphyrin intermediate in heme-based catalysis: revisiting the kinetic model for horseradish peroxidase

• Published in: Inorganica Chimica Acta 1998, Vol.276
• Main Authors: Haandel, M.J.H., van, Primus, J.L, Teunis, C, Boersma, M.G, Osman, A.M, Veeger, C, Rietjens, I.M.C.M
• Format: Article
• Subjects: Biochemie; Biochemistry
• ISSN: 0020-1693; 1873-3255

Many heme-containing biocatalysts exert their catalytic action through the initial formation of so-called high-valent-iron-oxo porphyrin intermediates. For horseradish peroxidase the initial intermediate formed has been identified as a high-valent-iron-oxo porphyrin π-radical cation, called compound I. A strongly held concept in the field of peroxidase-type of catalysis is the irreversible character of the reaction leading to formation of this compound I. Results of the present paper, however, point to reversibility of formation of the high-valent-iron-oxo porphyrin intermediate for various heme-containing catalysts, including horseradish peroxidase. This results in heme-catalysed exchange of the oxygens of H2O2 with those of H2O. The existence of this heme-catalysed oxygen exchange follows from the observation that upon incubation of 18O-labelled H182O2 with heme-containing biocatalysts significant loss of the 18O label from the H182O2, accompanied by the formation of unlabelled H2O2, is observed. Thus, for the heme biocatalysts studied, exchange of the oxygen of their high-valent-iron-oxo intermediate with that of water occurs rapidly. This observation implies the need for an update of the kinetic model for horseradish peroxidase. Revaluation and extension of the previous kinetic model showed the necessity to include several additional reaction steps, taking both reversible compound I formation and formation of enzyme-substrate complexes into account.