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Arabidopsis thaliana CYP77A4 is the first cytochrome P450 able to catalyze the epoxidation of free fatty acids in plants
Author(s) -
Sauveplane Vincent,
Kandel Sylvie,
Kastner PierreEdouard,
Ehlting Jürgen,
Compag Vincent,
WerckReichhart Danièle,
Pinot Franck
Publication year - 2009
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/j.1742-4658.2008.06819.x
Subject(s) - arabidopsis thaliana , cytochrome p450 , chemistry , cytochrome , biochemistry , arabidopsis , botany , organic chemistry , enzyme , biology , gene , mutant
An approach based on an in silico analysis predicted that CYP77A4, a cytochrome P450 that so far has no identified function, might be a fatty acid‐metabolizing enzyme. CYP77A4 was heterologously expressed in a Saccharomyces cerevisiae strain (WAT11) engineered for cytochrome P450 expression. Lauric acid (C 12:0 ) was converted into a mixture of hydroxylauric acids when incubated with microsomes from yeast expressing CYP77A4 . A variety of physiological C 18 fatty acids were tested as potential substrates. Oleic acid ( cis ‐Δ 9 C 18:1 ) was converted into a mixture of ω‐4‐ to ω‐7‐hydroxyoleic acids (75%) and 9,10‐epoxystearic acid (25%). Linoleic acid ( cis , cis ‐Δ 9 ,Δ 12 C 18:2 ) was exclusively converted into 12,13‐epoxyoctadeca‐9‐enoic acid, which was then converted into diepoxide after epoxidation of the Δ 9 unsaturation. Chiral analysis showed that 9,10‐epoxystearic acid was a mixture of 9 S/ 10 R and 9 R/ 10 S in the ratio 33 : 77, whereas 12,13‐epoxyoctadeca‐9‐enoic acid presented a strong enantiomeric excess in favor of 12 S/ 13 R , which represented 90% of the epoxide. Neither stearic acid (C 18:0 ) nor linolelaidic acid ( trans , trans ‐Δ 9 ,Δ 12 C 18:2 ) was metabolized, showing that CYP77A4 requires a double bond, in the cis configuration, to metabolize C 18 fatty acids. CYP77A4 was also able to catalyze the in vitro formation of the three mono‐epoxides of α‐linolenic acid ( cis , cis , cis ‐Δ 9 ,Δ 12 ,Δ 15 C 18:3 ), previously described as antifungal compounds. Epoxides generated by CYP77A4 are further metabolized to the corresponding diols by epoxide hydrolases located in microsomal and cytosolic subcellular fractions from Arabidopsis thaliana . The concerted action of CYP77A4 with epoxide hydrolases and hydroxylases allows the production of compounds involved in plant–pathogen interactions, suggesting a possible role for CYP77A4 in plant defense.