Biochemical properties of purified recombinant human β-carotene 15,15′-monooxygenase

Abstract

β-Carotene 15,15′-monooxygenase (BCO), formerly known as β-carotene 15,15′-dioxygenase, catalyzes the first step in the synthesis of vitamin A from dietary carotenoids. We have biochemically and enzymologically characterized the purified recombinant human BCO enzyme. A highly active BCO enzyme was expressed and purified to homogeneity from baculovirus-infected Spodoptera frugiperda 9 insect cells. The Km and Vmax of the enzyme for β-carotene were 7 μM and 10 nmol retinal/mg x min, respectively, values that corresponded to a turnover number (kcat) of 0.66 min-1 and a catalytic efficiency (kcat/Km) of ∼105 M-1.min-1. The enzyme existed as a tetramer in solution, and substrate specificity analyses suggested that at least one unsubstituted β-ionone ring half-site was imperative for efficient cleavage of the carbon 15,15′-double bond in carotenoid substrates. High levels of BCO mRNA were observed along the whole intestinal tract, in the liver, and in the kidney, whereas lower levels were present in the prostate, testis, ovary, and skeletal muscle. The current data suggest that the human BCO enzyme may, in addition to its well established role in the digestive system, also play a role in peripheral vitamin A synthesis from plasma-borne provitamin A carotenoids.