Calcium requirement for lipoxygenase catalyzed linoleate oxidation

Abstract

In further studies of the calcium (Ca²⁺) activation of lipoxygenase, it was observed that Ca²⁺ was the most specific cation for this enzyme activation. However, Mn²⁺ and Mg²⁺ were also activators, while Fe³⁺ and Cu²⁺ were competitive inhibitors. The pH activity curves for lipoxygenase showed different pH optima in the presence and absence of Ca²⁺. The difference can be explained by the presence of two enzymes or a single enzyme requiring two ionizable groups for activity. Buffer type and concentration were found to affect the enzyme activity at constant Ca²⁺ concentration. Order of addition of enzyme and Ca²⁺ to reaction mixture was investigated to better understand Ca²⁺ activation. Tween 20 was found to strongly inhibit the Ca²⁺ activated lipoxygenase at pH 7.5, while stimulating non-Ca²⁺-activated lipoxygenase at pH 6.4. These results seem to vie for the possibility of two different enzymes. A cold insoluble material was recovered from navy bean extracts which, when added to the reaction mixture, caused stimulation of lipoxygenase activity when low concentrations of imidazole were used as the buffer. These studies give additional evidence to show that despite previous reports to the contrary, a lipoxygenase does exist which requires the presence of a divalent cation for its activity.