title: Molecular cloning of a Pinguiochrysis pyriformis oleate-specific microsomal Δ12-fatty acid desaturase and functional analysis in yeasts and thraustochytrids.
abstract: We isolated a putative desaturase gene from a marine alga, Pinguiochrysis pyriformis MBIC 10872, which is capable of accumulating eicosapentaenoic acid (C20:5(Δ5,8,11,14,17)). The gene possessed an open reading frame of 1,314 bp encoding a putative 437 amino acid residues showing high sequence identity (37-48%) with fungal and nematode Δ12-fatty acid desaturases. Yeast cells transformed with the gene converted endogenous oleic acid (C18:1(Δ9)) to linoleic acid (C18:2(Δ9,12)). However, no double bonds were introduced into other endogenous fatty acids or exogenously added fatty acids. Flag-tagged enzyme was recovered in the micosome fraction when expressed in yeast cells. To express the gene in thraustochytrids, a construct driven by the thraustochytrid-derived ubiquitin promoter was used. Interestingly, exogenously added oleic acid was converted to linoleic acid in the gene transformants but not mock transformants of Aurantiochytrium limacinum mh0186. These results clearly indicate that the gene encodes a microsomal Δ12-fatty acid desaturase and was expressed functionally in not only yeasts but also thraustochytrids. This is the first report describing the heterozygous expression of a fatty acid desaturase in thraustochytrids, and could facilitate a genetic approach towards fatty acid synthesis in thraustochytrids which are expected to be an alternative source of polyunsaturated fatty acids.
authors.affiliation: Department of Bioscience and Biotechnology, Graduate School of Bioresource and Bioenvironmental Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan.
authors.forename: Takanori
authors.initials: T
authors.lastname: Matsuda
authors.forename: Keishi
authors.initials: K
authors.lastname: Sakaguchi
authors.forename: Takumi
authors.initials: T
authors.lastname: Kobayashi
authors.forename: Eriko
authors.initials: E
authors.lastname: Abe
authors.forename: Norihide
authors.initials: N
authors.lastname: Kurano
authors.forename: Akira
authors.initials: A
authors.lastname: Sato
authors.forename: Yuji
authors.initials: Y
authors.lastname: Okita
authors.forename: Shinichi
authors.initials: S
authors.lastname: Sugimoto
authors.forename: Yoichiro
authors.initials: Y
authors.lastname: Hama
authors.forename: Masahiro
authors.initials: M
authors.lastname: Hayashi
authors.forename: Nozomu
authors.initials: N
authors.lastname: Okino
authors.forename: Makoto
authors.initials: M
authors.lastname: Ito
mesh_terms: D003001:Cloning, Molecular; D044943:Fatty Acid Desaturases / Q000235:genetics* / Q000378:metabolism*; D058086:Microalgae / Q000201:enzymology* / Q000235:genetics*; D008969:Molecular Sequence Data; D044925:Oxidoreductases Acting on CH-CH Group Donors / Q000235:genetics* / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D058009:Stramenopiles / Q000201:enzymology* / Q000235:genetics*
chemical_list: D044943:Fatty Acid Desaturases; C021904:Phosphatidylcholine desaturase; D044925:Oxidoreductases Acting on CH-CH Group Donors