title: Enzymatic modification by point mutation and functional analysis of an omega-6 fatty acid desaturase from Arctic Chlamydomonas sp.
abstract: Arctic Chlamydomonas sp. is a dominant microalgal strain in cold or frozen freshwater in the Arctic region. The full-length open reading frame of the omega-6 fatty acid desaturase gene (AChFAD6) was obtained from the transcriptomic database of Arctic Chlamydomonas sp. from the KOPRI culture collection of polar micro-organisms. Amino acid sequence analysis indicated the presence of three conserved histidine-rich segments as unique characteristics of omega-6 fatty acid desaturases, and three transmembrane regions transported to plastidic membranes by chloroplast transit peptides in the N-terminal region. The AChFAD6 desaturase activity was examined by expressing wild-type and V254A mutant (Mut-AChFAD6) heterologous recombinant proteins. Quantitative gas chromatography indicated that the concentration of linoleic acids in AChFAD6-transformed cells increased more than 3-fold [6.73 ± 0.13 mg g-1dry cell weight (DCW)] compared with cells transformed with vector alone. In contrast, transformation with Mut-AChFAD6 increased the concentration of oleic acid to 9.23 ± 0.18 mg g-1DCW, indicating a change in enzymatic activity to mimic that of stearoyl-CoA desaturase. These results demonstrate that AChFAD6 of Arctic Chlamydomonas sp. increases membrane fluidity by enhancing denaturation of C18 fatty acids and facilitates production of large quantities of linoleic fatty acids in prokaryotic expression systems.
authors.affiliation: a Division of Polar Life Sciences, Korea Polar Research Institute , Korea Institute of Ocean Science and Technology , Incheon , Republic of Korea.
authors.forename: Woongsic
authors.initials: W
authors.lastname: Jung
authors.affiliation: a Division of Polar Life Sciences, Korea Polar Research Institute , Korea Institute of Ocean Science and Technology , Incheon , Republic of Korea.|b Department of Polar Life Sciences , University of Science and Technology , Incheon , Republic of Korea.
authors.forename: Eun Jae
authors.initials: EJ
authors.lastname: Kim
authors.affiliation: a Division of Polar Life Sciences, Korea Polar Research Institute , Korea Institute of Ocean Science and Technology , Incheon , Republic of Korea.|b Department of Polar Life Sciences , University of Science and Technology , Incheon , Republic of Korea.
authors.forename: Se Jong
authors.initials: SJ
authors.lastname: Han
authors.affiliation: c Division of Polar Ocean Sciences, Korea Polar Research Institute , Korea Institute of Ocean Science and Technology , Incheon , Republic of Korea.
authors.forename: Sung-Ho
authors.initials: SH
authors.lastname: Kang
authors.affiliation: a Division of Polar Life Sciences, Korea Polar Research Institute , Korea Institute of Ocean Science and Technology , Incheon , Republic of Korea.
authors.forename: Han-Gu
authors.initials: HG
authors.lastname: Choi
authors.affiliation: a Division of Polar Life Sciences, Korea Polar Research Institute , Korea Institute of Ocean Science and Technology , Incheon , Republic of Korea.
authors.forename: Sanghee
authors.initials: S
authors.lastname: Kim
mesh_terms: D000595:Amino Acid Sequence; D001110:Arctic Regions; D002696:Chlamydomonas / Q000201:enzymology*; D004926:Escherichia coli / Q000235:genetics; D044943:Fatty Acid Desaturases / Q000737:chemistry / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D017354:Point Mutation; D017386:Sequence Homology, Amino Acid
chemical_list: D044943:Fatty Acid Desaturases; C040448:delta-12 fatty acid desaturase
keywords: Arctic green microalga; Chlamydomonas sp; fatty acid methyl ester; linoleic acid methyl ester; oleic acid methyl ester; psychrophilic