The carotenase AtCCD1 from Arabidopsis thaliana is a dioxygenase.
Apocarotenoids resulting from the oxidative cleavage of carotenoids serve as important signaling and accessory molecules in a variety of biological processes. The enzymes catalyzing these reactions are referred to as carotenases or carotenoid oxygenases. Whether they act according to a monooxygenase mechanism, requiring two oxygens from different sources, or a dioxygenase mechanism is still a topic of controversy. In this study, we utilized the readily available beta-apo-8'-carotenal as a substrate for the heterologously expressed AtCCD1 protein from Arabidopsis thaliana to investigate the oxidative cleavage mechanism of the 9,10 double bond of carotenoids. Beta-ionone and a C(17)-dialdehyde were detected as products by gas and liquid chromatography-mass spectrometry as well as NMR analysis. Labeling experiments using H(2)(18)O or (18) O(2) showed that the oxygen in the keto-group of beta-ionone is derived solely from molecular dioxygen. When experiments were performed in an (18)O(2)-enriched atmosphere, a substantial fraction of the C(17)-dialdehyde contained labeled oxygen. The results unambiguously demonstrate a dioxygenase mechanism for the carotenase AtCCD1 from A. thaliana.
Schmidt
Holger
H
Foundation for Biomolecular Food Technology, Technische Universität München, Lise-Meitner-Strasse 34, D-84354 Freising, Germany. holger.schmidt@mytum.de
Kurtzer
Robert
R
Eisenreich
Wolfgang
W
Schwab
Wilfried
W
D017360:Arabidopsis / Q000201:enzymology*; D001665:Binding Sites; D002849:Chromatography, Gas; D002853:Chromatography, Liquid; D002855:Chromatography, Thin Layer; D003001:Cloning, Molecular; D049308:Dioxygenases / Q000737:chemistry* / Q000235:genetics; D018506:Gene Expression Regulation, Plant; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008956:Models, Chemical; D010100:Oxygen / Q000737:chemistry* / Q000378:metabolism; D010105:Oxygenases / Q000737:chemistry*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry
D011994:Recombinant Proteins; D010105:Oxygenases; D049308:Dioxygenases; C431939:carotenoid cleavage dioxygenase 1; D010100:Oxygen