␤-Carotene 15,15-monooxygenase (BCO), formerly known as ␤-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 baculovirusinfected Spodoptera frugiperda 9 insect cells. The K m and V max of the enzyme for ␤-carotene were 7 M and 10 nmol retinal/mg ؋ min, respectively, values that corresponded to a turnover number (k cat ) of 0.66 min ؊1 and a catalytic efficiency (k cat /K m ) of ϳ10 M ؊1 ⅐min؊1 . The enzyme existed as a tetramer in solution, and substrate specificity analyses suggested that at least one unsubstituted ␤-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.

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We demonstrate that in order to kill cancer cells MTH1 inhibitors must also introduce oxidized nucleotides into DNA. Furthermore, we describe TH1579 as a best-in-class MTH1 inhibitor, which we expect to be useful in order to further validate the MTH1 inhibitor concept.

S U M M A R YThe symmetrically cleaving ␤ -carotene 15,15 Ј -monooxygenase (BCO1) catalyzes the first step in the conversion of provitamin A carotenoids to vitamin A in the mucosa of the small intestine. This enzyme is also expressed in epithelia in a variety of extraintestinal tissues. The newly discovered ␤ -carotene 9 Ј ,10 Ј -monooxygenase (BCO2) catalyzes asymmetric cleavage of carotenoids. To gain some insight into the physiological role of BCO2, we determined the expression pattern of BCO2 mRNA and protein in human tissues. By immunohistochemical analysis it was revealed that BCO2 was detected in cell types that are known to express BCO1, such as epithelial cells in the mucosa of small intestine and stomach, parenchymal cells in liver, Leydig and Sertoli cells in testis, kidney tubules, adrenal gland, exocrine pancreas, and retinal pigment epithelium and ciliary body pigment epithelia in the eye. BCO2 was uniquely detected in cardiac and skeletal muscle cells, prostate and endometrial connective tissue, and endocrine pancreas. The finding that the BCO2 enzyme was expressed in some tissues and cell types that are not sensitive to vitamin A deficiency and where no BCO1 has been detected suggests that BCO2 may also be involved in biological processes other than vitamin A synthesis. T he enzymatic action of ␤ -carotene 15,15 Ј -monooxygenases (BCO1) is crucial for the conversion of provitamin A carotenoids to retinol (vitamin A) in the epithelial cells of the small intestine mucosa. The enzyme cleaves the ␤ -ionone ring containing carotenoids centrally, which results in two aldehyde molecules with polyene chains of identical length. Thus cleavage of the most common carotenoid ␤ -carotene results in two molecules of retinaldehyde (retinal), which will be further converted to retinol by a reductive retinal reductase enzyme present in the same cell as BCO1. Previously, we studied the cell type-specific expression of BCO1 and found that the enzyme is also expressed in epithelia of several extraintestinal tissues, which suggested that these tissues have the capacity to directly convert locally stored carotenoids to vitamin A. Thus this may serve as a backup pathway of vitamin A synthesis during times of insufficient dietary intake of preformed vitamin A and provitamin A carotenoids.Recently, Kiefer et al. (2001) cloned and characterized a second carotenoid-cleaving enzyme termed ␤ -carotene 9 Ј ,10 Ј -monooxygenase (BCO2) because it catalyzes asymmetric cleavage of carotenoids, thus yielding one molecule of ␤ -apo-10 Ј -carotenal and one molecule of ␤ -ionone when ␤ -carotene is used as substrate. Interestingly, the BCO2 enzyme appears to accept a wider variety of substrates as compared with BCO1, including the acyclic carotenoid lycopene, which implies that this enzyme may have physiological roles other than providing precursors for vitamin A synthesis.BCO1 and BCO2 belong to a superfamily of nonheme iron-containing oxygenases, many of whose functions are unknown. The physiological role of BCO1 is now well establ...