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 K(m) of 6.7 x 10 -5 M, and a V(max) 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.