Publications

• Cloning of the astaxanthin synthase gene from Xanthophyllomyces dendrorhous (Phaffia rhodozyma) and its assignment as a beta-carotene 3-hydroxylase/4-ketolase.

  Ojima K., Breitenbach J., Visser H., Setoguchi Y., Tabata K., Hoshino T., van den Berg J., Sandmann G.

  A gene has been cloned from Xanthophyllomyces dendrorhous by complementation of astaxanthin formation in a beta-carotene accumulating mutant. It consists of 3,166 bp and contains 17 introns. For the beta-carotene mutant ATCC 96815, a single point mutation in the splicing sequence of intron 8 was f ... >> More

  A gene has been cloned from Xanthophyllomyces dendrorhous by complementation of astaxanthin formation in a beta-carotene accumulating mutant. It consists of 3,166 bp and contains 17 introns. For the beta-carotene mutant ATCC 96815, a single point mutation in the splicing sequence of intron 8 was found. The resulting improper splicing of the mRNA results in an inactive protein. The cDNA of this beta-carotene oxygenase encodes a cytochrome P450 monooxygenase belonging to the 3A subfamily. P450-specific domains were identified including a cytochrome P450 and an oxygen binding motif. Electrons are provided by a cytochrome P450 reductase. Functional characterization of the enzyme by genetic modification of X. dendrorhous demonstrated that this P450 monooxygenase is multifunctional catalyzing all steps from beta-carotene to astaxanthin formation by oxygenation of carbon 3 and 4. The reaction sequence is first 4-ketolation of beta-carotene followed by 3-hydroxylation. A hydroxylation mechanism at allylic carbon atoms has been proposed for the generation of 4-keto and 3-hydroxy groups at both beta-ionone ends. << Less

  Mol Genet Genomics 275:148-158(2006) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• A carotenoid synthesis gene cluster from Algoriphagus sp. KK10202C with a novel fusion-type lycopene beta-cyclase gene.

  Tao L., Yao H., Kasai H., Misawa N., Cheng Q.

  A carotenoid synthesis gene cluster was isolated from a marine bacterium Algoriphagus sp. strain KK10202C that synthesized flexixanthin. Seven genes were transcribed in the same direction, among which five of them were involved in carotenoid synthesis. This cluster had a unique gene organization, ... >> More

  A carotenoid synthesis gene cluster was isolated from a marine bacterium Algoriphagus sp. strain KK10202C that synthesized flexixanthin. Seven genes were transcribed in the same direction, among which five of them were involved in carotenoid synthesis. This cluster had a unique gene organization, with an isoprenoid gene, ispH (previously named lytB), being present among the carotenoid synthesis genes. The lycopene beta-cyclase encoded by the crtY ( cd ) gene appeared to be a fusion of bacterial heterodimeric lycopene cyclase CrtY(c) and CrtY(d). This was the first time that a fusion-type of lycopene beta-cyclase was reported in eubacteria. Heterologous expression of the Algoriphagus crtY ( cd ) gene in lycopene-accumulating Escherichia coli produced bicyclic beta-carotene. A biosynthesis pathway for monocyclic flexixanthin was proposed in Algoriphagus sp. strain KK10202C, though several of the carotenoid synthesis genes not linked with the cluster have not yet been cloned. << Less

  Mol Genet Genomics 276:79-86(2006) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Cloning and expression in Escherichia coli of the gene encoding beta-C-4-oxygenase, that converts beta-carotene to the ketocarotenoid canthaxanthin in Haematococcus pluvialis.

  Lotan T., Hirschberg J.

  In the green alga Haematococcus pluvialis the ketocarotenoid astaxanthin accumulates under stress conditions. Astaxanthin is a red carotenoid pigment which possess antioxidative activity. We have cloned the gene for beta-C-4 oxygenase (beta-carotene ketolase) from the green algae H. pluvialis. The ... >> More

  In the green alga Haematococcus pluvialis the ketocarotenoid astaxanthin accumulates under stress conditions. Astaxanthin is a red carotenoid pigment which possess antioxidative activity. We have cloned the gene for beta-C-4 oxygenase (beta-carotene ketolase) from the green algae H. pluvialis. The cloning method took advantage of a strain of E. coli which was genetically engineered to produce beta-carotene. An expression cDNA library of H. pluvialis was transfected to cells of this strain and visually screened for brown-red pigmented colonies. One colony out of 100,000 transformants showed color change due to accumulation of canthaxanthin. The cDNA clone in this transformant colony encodes the enzyme beta-C-4 oxygenase that catalyzes the conversion of beta carotene to canthaxanthin via echinenone. This enzyme does not convert zeaxanthin to astaxanthin. It is concluded that in H. pluvialis astaxanthin is synthesized via canthaxanthin and therefore an additional enzyme is predicted, which converts canthaxanthin to astaxanthin. << Less

  FEBS Lett 364:125-128(1995) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Cloning of two carotenoid ketolase genes from Nostoc punctiforme for the heterologous production of canthaxanthin and astaxanthin.

  Steiger S., Sandmann G.

  For the heterologous synthesis of keto-carotenoids such as astaxanthin, two carotenoid ketolase genes crtW38 and crtW148, were cloned from the cyanobacterium, Nostoc punctiforme PCC 73102 and functionally characterized. Upon expression in Escherichia coli, both genes mediated the conversion of bet ... >> More

  For the heterologous synthesis of keto-carotenoids such as astaxanthin, two carotenoid ketolase genes crtW38 and crtW148, were cloned from the cyanobacterium, Nostoc punctiforme PCC 73102 and functionally characterized. Upon expression in Escherichia coli, both genes mediated the conversion of beta-carotene to canthaxanthin. However in a zeaxanthin-producing E. coli, only the gene product of crtW148 introduced 4-keto groups into the 3,3'-dihydroxy carotenoid zeaxanthin yielding astaxanthin. The gene product of crtW38 was unable to catalyze this reaction. Both ketolases differ in their interaction with a hydroxylase in the biosynthetic pathway from beta-carotene to astaxanthin. << Less

  Biotechnol Lett 26:813-817(2004) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Regulation of astaxanthin and its intermediates through cloning and genetic transformation of beta-carotene ketolase in Haematococcus pluvialis.

  Kathiresan S., Chandrashekar A., Ravishankar G.A., Sarada R.

  Astaxanthin, a high-value ketocarotenoid used in the pharmaceutical and nutraceutical industries is mainly produced from green alga, Haematococcus pluvialis. It is biosynthesized by the action of key enzyme, β-carotene ketolase (BKT) on β-carotene through intermediates echinenone and canthaxanthin ... >> More

  Astaxanthin, a high-value ketocarotenoid used in the pharmaceutical and nutraceutical industries is mainly produced from green alga, Haematococcus pluvialis. It is biosynthesized by the action of key enzyme, β-carotene ketolase (BKT) on β-carotene through intermediates echinenone and canthaxanthin. In this study, the β-carotene ketolase (bkt) gene was isolated from H. pluvialis and cloned in a vector pRT100 and further mobilized to a binary vector pCAMBIA 1304. The T-DNA of pCAMBIA 1304, which consists of cloned bkt, was successfully transformed to H. pluvialis through Agrobacterium mediation. The cloning and transformation of bkt in H. pluvialis was confirmed by Southern blotting and also by PCR analysis. Total carotenoids and astaxanthin content in the transformed cells were found to be 2-3-fold higher, while the intermediates like echinenone and canthaxanthin were found to be 8-10-fold higher than in the control cells. The expression level of carotenogenic genes like phytoene synthase (psy), phytoene desaturase (pds), lycopene cyclase (lcy), bkt, and β-carotene hydroxylase (bkh) were found to be higher in transformed cells compared to the non-transformed (NT) H. pluvialis. << Less

  J Biotechnol 196-197:33-41(2015) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Expression in Escherichia coli and properties of the carotene ketolase from Haematococcus pluvialis.

  Breitenbach J., Misawa N., Kajiwara S., Sandmann G.

  High level expression of the functional beta-carotene ketolase gene bkt from Haematococcus pluvialis occurred in Escherichia coli transformants producing beta-carotene or zeaxanthin as a result of the presence of additional carotenoid genes from Erwinia uredovora. Requirement of molecular oxygen f ... >> More

  High level expression of the functional beta-carotene ketolase gene bkt from Haematococcus pluvialis occurred in Escherichia coli transformants producing beta-carotene or zeaxanthin as a result of the presence of additional carotenoid genes from Erwinia uredovora. Requirement of molecular oxygen for the insertion of the keto group was demonstrated. The final product of this two-step ketolase reaction from beta-carotene is canthaxanthin (4,4'-diketo-beta-carotene) with the 4-monoketo derivative echinenone as an intermediate. A reaction sequence for the formation of astaxanthin from beta-carotene was established based on kinetic data on astaxanthin formation in E. coli transformants carrying the hydroxylase gene crtZ from Erwinia along with bkt. We conclude that the carotenoids zeaxanthin and adonixanthin which accumulate in addition to astaxanthin in this transformant are products of side reactions rather than direct precursors of astaxanthin. The possible mechanisms for the formation of the keto derivatives are discussed. << Less

  FEMS Microbiol Lett 140:241-246(1996) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.