title: A Δ-9 Fatty Acid Desaturase Gene in the Microalga Myrmecia incisa Reisigl: Cloning and Functional Analysis.
abstract: The green alga Myrmecia incisa is one of the richest natural sources of arachidonic acid (ArA). To better understand the regulation of ArA biosynthesis in M. incisa, a novel gene putatively encoding the Δ9 fatty acid desaturase (FAD) was cloned and characterized for the first time. Rapid-amplification of cDNA ends (RACE) was employed to yield a full length cDNA designated as MiΔ9FAD, which is 2442 bp long in sequence. Comparing cDNA open reading frame (ORF) sequence to genomic sequence indicated that there are 8 introns interrupting the coding region. The deduced MiΔ9FAD protein is composed of 432 amino acids. It is soluble and localized in the chloroplast, as evidenced by the absence of transmembrane domains as well as the presence of a 61-amino acid chloroplast transit peptide. Multiple sequence alignment of amino acids revealed two conserved histidine-rich motifs, typical for Δ9 acyl-acyl carrier protein (ACP) desaturases. To determine the function of MiΔ9FAD, the gene was heterologously expressed in a Saccharomyces cerevisiae mutant strain with impaired desaturase activity. Results of GC-MS analysis indicated that MiΔ9FAD was able to restore the synthesis of monounsaturated fatty acids, generating palmitoleic acid and oleic acid through the addition of a double bond in the Δ9 position of palmitic acid and stearic acid, respectively.
authors.affiliation: College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China. xwb989510@gmail.com.
authors.forename: Wen-Bin
authors.initials: WB
authors.lastname: Xue
authors.affiliation: College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China. liufanlflf@gmail.com.
authors.forename: Fan
authors.initials: F
authors.lastname: Liu
authors.affiliation: College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China. zsun@shou.edu.cn.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Sun
authors.affiliation: College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China. zgzhou@shou.edu.cn.
authors.forename: Zhi-Gang
authors.initials: ZG
authors.lastname: Zhou
mesh_terms: D000595:Amino Acid Sequence; D016718:Arachidonic Acid / Q000378:metabolism*; D003001:Cloning, Molecular; D044943:Fatty Acid Desaturases / Q000235:genetics / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D058086:Microalgae / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D010802:Phylogeny; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013230:Stearoyl-CoA Desaturase
chemical_list: D016718:Arachidonic Acid; D044943:Fatty Acid Desaturases; D013230:Stearoyl-CoA Desaturase; C029627:delta-9 fatty acid desaturase
keywords: Acyl-ACP desaturase; RACE; Saccharomyces cerevisiae; fatty acid composition