Ca2+Activation of Wheat Peroxidase: A Possible Physiological Mechanism of Control

Peroxidation of substrates such as ascorbic acid, pyrogallol, or ferulic acid, as well as indole acetic acid oxidation catalyzed by wheat germ peroxidase (WGP)22Abbreviations used: WGP, wheat germ peroxidase; HRP, horseradish peroxidase; IAA, indolacetic acid; FA, ferulic acid; P, peroxidase resting state; CMP I, compound I; CMP II, compound II; HOPh, a phenol; HOP[formula], a phenol radical. re found to be activated by Ca2+. This activation is independent of the stabilizing effect of structural Ca2+reported for peroxidases. Steady state kinetics of ferulic acid oxidation catalyzed by WGP C2showed an increase in the rate of compound I formation and of compound II decomposition in the presence of the ion, evidenced as an increase in rate constantsk1, from 8.9 × 105to 4.5 × 106m−1cm−1, andk3, from 4.4 × 105to 1.1 × 106m−1cm−1. The dissociation constantKd, for the cyanide derivative of the enzyme showed a marked decrease from 220 to 34 μmin the presence of Ca2+, thus implying an effect of the ion in the H2O2binding step. In the presence of Ca2+, a conformational change in the protein was revealed by tryptophan fluorescence, providing a basis for the activation mechanism. Other peroxidases such as horseradish peroxidase and WGP C3were not activated by Ca2+. The results suggest the existence of a physiological mechanism of control of peroxidase isozymes activity mediated by Ca2+.