A peroxygenase pathway involved in the biosynthesis of epoxy fatty acids in oat

  1. Get@NRC: A peroxygenase pathway involved in the biosynthesis of epoxy fatty acids in oat (Opens in a new window)
DOIResolve DOI: http://doi.org/10.1104/pp.111.178822
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Journal titlePlant Physiology
Pages454463; # of pages: 10
Subjectcomplementary DNA; fatty acid; lipoxygenase; mixed function oxidase; peroxygenase; primer DNA; article; enzyme specificity; Escherichia coli; genetics; mass fragmentography; metabolism; molecular cloning; molecular genetics; nucleotide sequence; oat; Pichia; plasmid; Avena sativa; Base Sequence; Cloning, Molecular; DNA Primers; DNA, Complementary; Escherichia coli; Fatty Acids; Gas Chromatography-Mass Spectrometry; Lipoxygenases; Mixed Function Oxygenases; Molecular Sequence Data; Pichia; Plasmids; Substrate Specificity; Avena sativa; Escherichia coli; Pichia pastoris
AbstractWhile oat (Avena sativa) has long been known to produce epoxy fatty acids in seeds, synthesized by a peroxygenase pathway, the gene encoding the peroxygenase remains to be determined. Here we report identification of a peroxygenase cDNA AsPXG1 from developing seeds of oat. AsPXG1 is a small protein with 249 amino acids in length and contains conserved heme-binding residues and a calcium-binding motif. When expressed in Pichia pastoris and Escherichia coli, AsPXG1 catalyzes the strictly hydroperoxide-dependent epoxidation of unsaturated fatty acids. It prefers hydroperoxy-trienoic acids over hydroperoxydienoic acids as oxygen donors to oxidize a wide range of unsaturated fatty acids with cis double bonds. Oleic acid is the most preferred substrate. The acyl carrier substrate specificity assay showed phospholipid and acyl-CoA were not effective substrate forms for AsPXG1 and it could only use free fatty acid or fatty acid methyl esters as substrates. A second gene, AsLOX2, cloned from oat codes for a 9-lipoxygenase catalyzing the synthesis of 9-hydroperoxy-dienoic and 9-hydroperoxy-trienoic acids, respectively, when linoleic (18:2-9c,12c) and linolenic (18:3-9c,12c,15c) acids were used as substrates. The peroxygenase pathway was reconstituted in vitro using a mixture of AsPXG1 and AsLOX2 extracts from E. coli. Incubation of methyl oleate and linoleic acid or linolenic acid with the enzyme mixture produced methyl 9,10-epoxy stearate. Incubation of linoleic acid alone with a mixture of AsPXG1 and AsLOX2 produced two major epoxy fatty acids, 9,10-epoxy-12-cis-octadecenoic acid and 12,13-epoxy-9-cis-octadecenoic acid, and a minor epoxy fatty acid, probably 12,13-epoxy-9-hydroxy-10-transoctadecenoic acid. AsPXG1 predominately catalyzes intermolecular peroxygenation. © 2011 American Society of Plant Biologists. All Rights Reserved.
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AffiliationNational Research Council Canada (NRC-CNRC); NRC Plant Biotechnology Institute (PBI-IBP)
Peer reviewedYes
NPARC number21271137
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Record identifier9d0a71e9-da8e-4f0e-9049-bc4c95058816
Record created2014-03-24
Record modified2016-05-09
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