Hydroperoxidase activity of lipoxygenase: a kinetic study of isoproterenol oxidation

Abstract

The hydroperoxidase activity of soybean lipoxygenase, a non-heme protein, oxidized isoproterenol using H 20 2 at pH 6.0. This oxidation was enzymatic, since neither heat-denaturated enzyme or iron ions in the presence of H 20 2 produced an increase in absorbance. The initial rate was not linear and showed a characteristic lag period whose length depended on the enzyme and substrate concentration. The lag was decreased if the enzyme and isoproterenol concentration were increased, whereas it increased if the H 20 2 concentration was increased. Lipoxygenase showed the typical low specificity for electron donor characteristic of this hydroperoxidase activity (26 mM), but a high affinity for H 20 2 (94 μM), although with substrate inhibition ( k si = 3.6 mM). The chemical intermediates produced during the oxidation of isoproterenol were characterized in order to determine the origin of the lag period. A plausible kinetic mechanism is proposed to explain the observed lag period and inhibition by high concentrations of H 20 2.