title: Characterization of cyanobacterial carotenoid ketolase CrtW and hydroxylase CrtR by complementation analysis in Escherichia coli.
abstract: The pathway from beta-carotene to astaxanthin is a crucial step in the synthesis of astaxanthin, a red antioxidative ketocarotenoid that confers beneficial effects on human health. Two enzymes, a beta-carotene ketolase (carotenoid 4,4'-oxygenase) and a beta-carotene hydroxylase (carotenoid 3,3'-hydroxylase), are involved in this pathway. Cyanobacteria are known to utilize the carotenoid ketolase CrtW and/or CrtO, and the carotenoid hydroxylase CrtR. Here, we compared the catalytic functions of CrtW ketolases, which originated from Gloeobacter violaceus PCC 7421, Anabaena (also known as Nostoc) sp. PCC 7120 and Nostoc punctiforme PCC 73102, and CrtR from Synechocystis sp. PCC 6803, Anabaena sp. PCC 7120 and Anabaena variabilis ATCC 29413 by complementation analysis using recombinant Escherichia coli cells that synthesized various carotenoid substrates. The results demonstrated that the CrtW proteins derived from Anabaena sp. PCC 7120 as well as N. punctiforme PCC 73102 (CrtW148) can convert not only beta-carotene but also zeaxanthin into their 4,4'-ketolated products, canthaxanthin and astaxanthin, respectively. In contrast, the Anabaena CrtR enzymes were very poor in accepting either beta-carotene or canthaxanthin as substrates. By comparison, the Synechocystis sp. PCC 6803 CrtR converted beta-carotene into zeaxanthin efficiently. We could assign the catalytic functions of the gene products involved in ketocarotenoid biosynthetic pathways in Synechocystis sp. PCC 6803, Anabaena sp. PCC 7120 and N. punctiforme PCC 73102, based on the present and previous findings. This explains why these cyanobacteria cannot produce astaxanthin and why only Synechocystis sp. PCC 6803 can produce zeaxanthin.
authors.affiliation: School of Fisheries Sciences, Kitasato University, Sanriku-cho, Ofunato, 022-0101 Japan.
authors.forename: Takuya
authors.initials: T
authors.lastname: Makino
authors.forename: Hisashi
authors.initials: H
authors.lastname: Harada
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Ikenaga
authors.forename: Satoru
authors.initials: S
authors.lastname: Matsuda
authors.forename: Shinichi
authors.initials: S
authors.lastname: Takaichi
authors.forename: Kazutoshi
authors.initials: K
authors.lastname: Shindo
authors.forename: Gerhard
authors.initials: G
authors.lastname: Sandmann
authors.forename: Takehiko
authors.initials: T
authors.lastname: Ogata
authors.forename: Norihiko
authors.initials: N
authors.lastname: Misawa
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D016644:Canthaxanthin / Q000378:metabolism; D000458:Cyanobacteria / Q000201:enzymology / Q000235:genetics*; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D006899:Mixed Function Oxygenases / Q000235:genetics / Q000378:metabolism*; D010105:Oxygenases / Q000235:genetics / Q000378:metabolism*; D013045:Species Specificity; D013379:Substrate Specificity; D024341:Xanthophylls / Q000096:biosynthesis / Q000378:metabolism; D065146:Zeaxanthins; D019207:beta Carotene / Q000096:biosynthesis
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D024341:Xanthophylls; D065146:Zeaxanthins; D019207:beta Carotene; D016644:Canthaxanthin; C005948:astaxanthine; D006899:Mixed Function Oxygenases; D010105:Oxygenases; C093259:beta-carotene ketolase; C079232:beta-carotene hydroxylase