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

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

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Abstract

A gene has been cloned from Xanthophyllomyces dendrorhous by complementation of astaxanthin formation in a ß-carotene accumulating mutant. It consists of 3,166 bp and contains 17 introns. For the ß-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 ß-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 ß-carotene to astaxanthin formation by oxygenation of carbon 3 and 4. The reaction sequence is first 4-ketolation of ß-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 ß-ionone ends
Original languageEnglish
Pages (from-to)148-158
JournalMolecular Genetics and Genomics
Volume275
Issue number2
DOIs
Publication statusPublished - 2006

Keywords

  • escherichia-coli
  • functional-characterization
  • haematococcus-pluvialis
  • biosynthetic-pathway
  • yeast
  • family
  • host
  • canthaxanthin
  • hydroxylase
  • expression

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