Delta 9-fatty acid desaturase from arachidonic acid-producing fungus. Unique gene sequence and its heterologous expression in a fungus, Aspergillus.
Based on the sequence information for delta 9-desaturase genes (from rat, mouse and yeast), which are involved in the desaturation of palmitic acid and stearic acid to palmitoleic acid and oleic acid, respectively, the corresponding cDNA and genomic gene were cloned from the fungal strain, Mortierella alpina 1S-4, which industrially produces arachidonic acid. There was a cytochrome b5-like domain linked to the carboxyl terminus of this Mortierella desaturase, as also seen in the yeast delta 9-desaturase. The Mortierella delta 9-desaturase genomic gene had only one intron, in which a novel phenomenon was observed: there was a GC-end at the 5'-terminus instead of a GT-end that is, in general, found in introns of eukaryotic genes. The full-length cDNA clone was expressed under the control of an amyB promoter in a filamentous fungus, Aspergillus oryzae, resulting in drastic changes in the fatty acid composition in the transformant cells; the contents of palmitoleic acid (16:1) and oleic acid (18:1) increased significantly, with accompanying decreases in palmitic acid (16:0) and stearic acid (18:0). These changes were controlled by the addition of maltose as a carbon source to the medium. Also, the expression of the gene caused a significant change in the lipid composition in the Aspergillus transformant. Genomic Southern blot analysis of the transformant with the Mortierella delta 9-desaturase gene as a probe confirmed the integration of this gene into the genome of A. oryzae.
Sakuradani
E
E
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Japan.
Kobayashi
M
M
Shimizu
S
S
D000595:Amino Acid Sequence; D016718:Arachidonic Acid / Q000378:metabolism*; D001236:Aspergillus oryzae / Q000201:enzymology*; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D044943:Fatty Acid Desaturases / Q000235:genetics*; D005227:Fatty Acids / Q000032:analysis; D005656:Fungal Proteins / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics; D015966:Gene Expression Regulation, Fungal / Q000235:genetics; D008055:Lipids / Q000032:analysis; D008969:Molecular Sequence Data; D020094:Mortierella / Q000201:enzymology*; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013230:Stearoyl-CoA Desaturase
D005227:Fatty Acids; D005656:Fungal Proteins; D008055:Lipids; D016718:Arachidonic Acid; D044943:Fatty Acid Desaturases; D013230:Stearoyl-CoA Desaturase; C029627:delta-9 fatty acid desaturase