Metabolic engineering of the carotenoid biosynthetic pathway in the yeast Xanthophyllomyces dendrorhous (Phaffia rhodozyma).
The crtYB locus was used as an integrative platform for the construction of specific carotenoid biosynthetic mutants in the astaxanthin-producing yeast Xanthophyllomyces dendrorhous. The crtYB gene of X. dendrorhous, encoding a chimeric carotenoid biosynthetic enzyme, could be inactivated by both single and double crossover events, resulting in non-carotenoid-producing transformants. In addition, the crtYB gene, linked to either its homologous or a glyceraldehyde-3-phosphate dehydrogenase promoter, was overexpressed in the wild type and a beta-carotene-accumulating mutant of X. dendrorhous. In several transformants containing multiple copies of the crtYB gene, the total carotenoid content was higher than in the control strain. This increase was mainly due to an increase of the beta-carotene and echinone content, whereas the total content of astaxanthin was unaffected or even lower. Overexpression of the phytoene synthase-encoding gene (crtI) had a large impact on the ratio between mono- and bicyclic carotenoids. Furthermore, we showed that in metabolic engineered X. dendrorhous strains, the competition between the enzymes phytoene desaturase and lycopene cyclase for lycopene governs the metabolic flux either via beta-carotene to astaxanthin or via 3,4-didehydrolycopene to 3-hydroxy-3'-4'-didehydro-beta-psi-caroten-4-one (HDCO). The monocylic carotenoid torulene and HDCO, normally produced as minority carotenoids, were the main carotenoids produced in these strains.
Verdoes
Jan C
JC
Division of Industrial Microbiology, Department of Agrotechnology and Food Sciences, Wageningen University, Dreijenlaan 2, 6703 HA Wageningen, The Netherlands.
Sandmann
Gerhard
G
Visser
Hans
H
Diaz
Maria
M
van Mossel
Minca
M
van Ooyen
Albert J J
AJ
D019883:Alkyl and Aryl Transferases / Q000235:genetics / Q000378:metabolism; D001487:Basidiomycota / Q000235:genetics / Q000378:metabolism; D002338:Carotenoids / Q000096:biosynthesis; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D005818:Genetic Engineering / Q000379:methods*; D051232:Geranylgeranyl-Diphosphate Geranylgeranyltransferase; D019753:Intramolecular Lyases / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D024341:Xanthophylls; D019207:beta Carotene / Q000031:analogs & derivatives* / Q000096:biosynthesis
D005656:Fungal Proteins; D011994:Recombinant Proteins; D024341:Xanthophylls; D019207:beta Carotene; D002338:Carotenoids; C005948:astaxanthine; D010088:Oxidoreductases; C022398:phytoene dehydrogenase; D019883:Alkyl and Aryl Transferases; D051232:Geranylgeranyl-Diphosphate Geranylgeranyltransferase; D019753:Intramolecular Lyases; C070925:lycopene cyclase-isomerase