Comparisons between hog intestinal peroxidase and bovine lactoperoxidase—Compound I formation and inhibition by benzhydroxamic acid

Abstract

Absorbance spectra of hog intestinal peroxidase and bovine lactoperoxidase resembled each other in every state of Compounds I, II, and III. The rate constant for Compound I formation was different between the two enzymes, but their pH-rate profiles were alike. The heme-linked ionizations of p K = 4.75 in intestinal peroxidase and p K = 3.5 in lactoperoxidase (S. Kimura and I. Yamazaki, 1978, Arch. Biochem. Biophys. 189, 14) were directly reflected in the pH dependence of the dissociation constant for the enzyme-benzhydroxamic acid complex, but not in that of the rate of Compound I formation. Benzhydroxamic acid inhibited the peroxidative oxidation of guaiacol. The concentration of benzhydroxamic acid causing the half-inhibition was 0.5 μ m for intestinal peroxidase and 100 μ m for lactoperoxidase. A new intermediate compound with an absorbance spectrum similar to that of Compound III was formed during the reduction of intestinal peroxidase Compound II by benzhydroxamic acid. Such a compound was not observed in the reaction of lactoperoxidase. The p K = 7.07 ionization in lactoperoxidase was associated with the reactions between benzhydroxamic acid and the native enzyme or Compound II, while no analogous ionization was detectable in intestinal peroxidase because the ionization of benzhydroxamic acid with p K = 8.8 obscured the analysis.