Purification and properties of carotene 15,15′-dioxygenase of rabbit intestine

Abstract

Carotene 15,15′-dioxygenase, which oxidizes carotenoids to retinal, has been purified up to 200-fold from rabbit intestine by ammonium sulfate fractionation, heat treatment, and acetone precipitation. With β-apo-10′-carotenol as the substrate, the purified enzyme has a pH optimum of 7.8, a Km, of 6.7 × 10–5 m, and a Vmax at 37°C of 9 nmoles of retinal/mg protein/hr. The purified enzyme is inhibited by ferrous ion-chelating agents such as α,α′-dipyridyl and o-phenanthroline, and by sulfhydryl-binding agents such as iodoacetamide, N-ethylmaleimide, and p-chloromercuribenzoate. The latter inhibitory effects are reversed by reduced glutathione. The cleavage of β-apo-10′-carotenol is competitively inhibited by its acetylenic analog, 15,15′-dehydro-β-apo-10′-carotenol. The enzyme is present in the intestinal mucosa of several mammals, the chicken, the tortoise, and a freshwater fish, but it is absent from cat intestinal tissue.

Keywords

• β-apo-10′-carotenol
• β-carotene cleavage
• 15,15′-dehydro-β-apo-10′-carotenol
• retinal
• thiobarbituric acid method
• mammals