A novel fungal omega3-desaturase with wide substrate specificity from arachidonic acid-producing Mortierella alpina 1S-4.
A filamentous fungus, Mortierella alpina 1S-4, is capable of producing not only arachidonic acid (AA; 20:4n-6) but also eicosapentaenoic acid (EPA; 20:5n-3) below a cultural temperature of 20 degrees C. Here, we describe the isolation and characterization of a gene (maw3) that encodes a novel omega3-desaturase from M. alpina 1S-4. Based on the conserved sequence information for M. alpina 1S-4 Delta12-desaturase and Saccharomyces kluyveri omega3-desaturase, the omega3-desaturase gene from M. alpina 1S-4 was cloned. Homology analysis of protein databases revealed that the amino acid sequence showed 51% identity, at the highest, with M. alpina 1S-4 Delta12-desaturase, whereas it exhibited 36% identity with Sac. kluyveri omega3-desaturase. The cloned cDNA was confirmed to encode the omega3-desaturase by its expression in the yeast Sac. cerevisiae. Analysis of the fatty acid composition of the yeast transformant demonstrated that 18-carbon and 20-carbon n-3 polyunsaturated fatty acids (PUFAs) were accumulated through conversion of exogenous 18-carbon and 20-carbon n-6 PUFAs. The substrate specificity of the M. alpina 1S-4 omega3-desaturase differs from those of the known fungal omega3-desaturases from Sac. kluyveri and Saprolegnia diclina. Plant, cyanobacterial and Sac. kluyveri omega3-desaturases desaturate 18-carbon n-6 PUFAs, Spr. diclina omega3-desaturase desaturates 20-carbon n-6 PUFAs and Caenorhabditis elegans omega3-desaturase prefers 18-carbon n-6 PUFAs as substrates rather than 20-carbon n-6 PUFAs. The substrate specificity of M. alpina 1S-4 omega3-desaturase is rather similar to that of C. elegans omega3-desaturase, but the M. alpina omega3-desaturase can more effectively convert AA into EPA when expressed in yeast. The M. alpina 1S-4 omega3-desaturase is the first known fungal desaturase that uses both 18-carbon and 20-carbon n-6 PUFAs as substrates.
Sakuradani
Eiji
E
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan.
Abe
Takahiro
T
Iguchi
Keita
K
Shimizu
Sakayu
S
D000595:Amino Acid Sequence; D000818:Animals; D017173:Caenorhabditis elegans / Q000201:enzymology; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000737:chemistry; D044943:Fatty Acid Desaturases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D005227:Fatty Acids / Q000032:analysis / Q000302:isolation & purification; D005231:Fatty Acids, Unsaturated / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D005800:Genes, Fungal; D008969:Molecular Sequence Data; D020094:Mortierella / Q000201:enzymology*; D010802:Phylogeny; D012440:Saccharomyces / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D044762:Saprolegnia / Q000201:enzymology; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D004271:DNA, Fungal; D005227:Fatty Acids; D005231:Fatty Acids, Unsaturated; D005656:Fungal Proteins; D044943:Fatty Acid Desaturases; C078565:omega-3 fatty acid desaturase