Thiocyanate Modulates the Catalytic Activity of Mammalian Peroxidases *

We investigated the potential role of the co-substrate, thiocyanate (SCN – ), in modulating the catalytic activity of myeloperoxidase (MPO) and other members of the mammalian peroxidase superfamily (lactoperoxidase (LPO) and eosinophil peroxidase (EPO)). Pre-incubation of SCN – with MPO generates a more complex biological setting, because SCN – serves as either a substrate or inhibitor, causing diverse impacts on the MPO heme iron microenvironment. Consistent with this hypothesis, the relationship between the association rate constant of nitric oxide binding to MPO-Fe(III) as a function of SCN – concentration is bell-shaped, with a trough comparable with normal SCN – plasma levels. Rapid kinetic measurements indicate that MPO, EPO, and LPO Compound I formation occur at rates slower than complex decay, and its formation serves to simultaneously catalyze SCN – via 1e – and 2e – oxidation pathways. For the three enzymes, Compound II formation is a fundamental feature of catalysis and allows the enzymes to operate at a fraction of their possible maximum activities. MPO and EPO Compound II is relatively stable and decays gradually within minutes to ground state upon H 2 O 2 exhaustion. In contrast, LPO Compound II is unstable and decays within seconds to ground state, suggesting that SCN – may serve as a substrate for Compound II. Compound II formation can be partially or completely prevented by increasing SCN – concentration, depending on the experimental conditions. Collectively, these results illustrate for the first time the potential mechanistic differences of these three enzymes. A modified kinetic model, which incorporates our current findings with the mammalian peroxidases classic cycle, is presented.