Purification and properties of carotene 15,15'-dioxygenase of rabbit intestine.
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 beta-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 degrees C of 9 nmoles of retinal/mg protein/hr. The purified enzyme is inhibited by ferrous ion-chelating agents such as alpha,alpha'-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 beta-apo-10'-carotenol is competitively inhibited by its acetylenic analog, 15,15'-dehydro-beta-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.
D000096:Acetone; D000818:Animals; D002338:Carotenoids; D002415:Cats; D002614:Chelating Agents; D002645:Chickens; D004586:Electrophoresis; D005399:Fishes; D005978:Glutathione; D006168:Guinea Pigs; D000882:Haplorhini; D006863:Hydrogen-Ion Concentration; D007413:Intestinal Mucosa / Q000201:enzymology*; D007421:Intestine, Small / Q000201:enzymology; D007700:Kinetics; D010105:Oxygenases / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D011817:Rabbits; D013045:Species Specificity; D013329:Structure-Activity Relationship; D013439:Sulfhydryl Reagents; D014426:Turtles; D014461:Ultracentrifugation
D002614:Chelating Agents; D013439:Sulfhydryl Reagents; D000096:Acetone; D002338:Carotenoids; D010105:Oxygenases; D005978:Glutathione