1887

Abstract

Genes encoding two distinct fatty acid Δ9-desaturases were isolated from strains of the oleaginous fungus . Two genomic sequences, Δ9-1 and Δ9-2, each containing a single intron, were cloned from strain CBS 528.72 while one cDNA clone, LM9, was isolated from strain CBS 210.32. The Δ9-1 gene encoded a protein of 445 aa which shared 99% identity with the LM9 gene product. These proteins also showed 40–60% identity to the Δ9-desaturases (Ole1p) of other fungi and contained the three conserved histidine boxes, C-terminal cytochrome fusion and transmembrane domains characteristic of endoplasmic reticulum membrane-bound Δ9-desaturases. LM9 and Δ9-1 are therefore considered to represent the same gene (). The gene was transcriptionally active in all strains tested and its function was confirmed by complementation of the mutation. Fatty acid analysis of yeast transformants expressing the CBS 210.32 gene showed an elevated level of oleic acid (18:1) compared to palmitoleic acid (16:1), the major fatty acid component of wild-type . This indicated that the Δ9-desaturase had a substrate preference for stearic acid (18:0) rather than palmitic acid (16:0). Genomic clone Δ9-2 () also encoded a protein of 445 aa which had 86% identity to the Δ9-1 and LM9 proteins and whose ORF also complemented the yeast mutation. The transcript from this gene could only be detected in one of the six strains tested, suggesting that its expression may be strain-specific or induced under certain physiological conditions.

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1999-10-01
2019-10-24
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References

  1. Anamnart, S., Tomita, T., Fukui, F., Fujimori, K., Harashima, S., Yamada, Y. & Oshima, Y. ( 1997; ). The P-OLE1 gene of Pichia angusta encodes a Δ9-fatty acid desaturase and complements the ole1 mutation of Saccharomyces cerevisiae. Gene 184, 299-306.[CrossRef]
    [Google Scholar]
  2. Certik, M., Sakuradani, E. & Shimizu, S. ( 1998; ). Desaturase-defective fungal mutants: useful tools for the regulation and overproduction of polyunsaturated fatty acids. Trends Biotechnol 16, 500-505.[CrossRef]
    [Google Scholar]
  3. Choi, J.-Y., Stukey, J., Hwang, S.-Y. & Martin, C. E. ( 1996; ). Regulatory elements that control transcription activation and unsaturated fatty acid-mediated repression of the Saccharomyces cerevisiae OLE1 gene. J Biol Chem 271, 3581-3589.[CrossRef]
    [Google Scholar]
  4. Fukuchi-Mizutani, M., Savin, K., Cornish, E., Tanaka, Y., Ashikari, T., Kusumi, T. & Murata, N. ( 1995; ). Senescence-induced expression of a homologue of Δ9 desaturase in rose petals. Plant Mol Biol 29, 627-635.[CrossRef]
    [Google Scholar]
  5. Fukuchi-Mizutani, M., Tasaka, Y., Tanaka, Y., Ashikari, T., Kusumi, T. & Murata, N. ( 1998; ). Characterization of Δ9 acyl-lipid desaturase homologues from Arabidopsis thaliana. Plant Cell Physiol 39, 247-253.[CrossRef]
    [Google Scholar]
  6. Gargano, S., Di Lallo, G., Kobayashi, G. S. & Maresca, B. ( 1995; ). A temperature-sensitive strain of Histoplasma capsulatum has an altered Δ9-fatty acid desaturase gene. Lipids 30, 899-906.[CrossRef]
    [Google Scholar]
  7. Gietz, R. D., Schiestl, R. H., Willems, A. R. & Woods, R. A. ( 1995; ). Studies on the transformation of intact yeast cells by the LiAc/ss-DNA/PEG procedure. Yeast 11, 355-360.[CrossRef]
    [Google Scholar]
  8. Gonzalez, C. I. & Martin, C. E. ( 1996; ). Fatty acid-responsive control of mRNA stability. J Biol Chem 271, 25801-25809.[CrossRef]
    [Google Scholar]
  9. Gurr, S. J., Unkles, S. E. & Kinghorn, J. R. ( 1987; ). The structure and organization of nuclear genes of filamentous fungi. In Gene Structure in Eukaryotic Microbes, pp. 93-139. Edited by J. R. Kinghorn. Oxford: IRL Press.
  10. Hamilton, R., Watanabe, C. K. & de Boer, H. A. ( 1987; ). Compilation and comparison of the sequence context around the AUG start codons in Saccharomyces cerevisiae mRNAs. Nucleic Acids Res 15, 3581-3593.[CrossRef]
    [Google Scholar]
  11. Harwood, J. L. ( 1997; ). Plant lipid metabolism. In Plant Biochemistry, pp. 237-272. Edited by P. M. Dey & J. B. Harborne. London: Academic Press.
  12. Hempenius, R. A., van Delft, J. M. H., Prinsen, M. & Lina, B. A. R. ( 1997; ). Preliminary safety assessment of an arachidonic acid-enriched oil derived from Mortierella alpina: summary of toxicological data. Food Chem Toxicol 35, 573-581.[CrossRef]
    [Google Scholar]
  13. Jackson, F. M., Fraser, T. C. M., Smith, M. A., Lazarus, C., Stobart, A. K. & Griffiths, G. ( 1998; ). Biosynthesis of C18 polyunsaturated fatty acids in microsomal membrane preparations from the filamentous fungus Mucor circinelloides. Eur J Biochem 252, 513-519.[CrossRef]
    [Google Scholar]
  14. Jareonkitmongkol, S., Sakuradani, E. & Shimizu, S. ( 1994; ). Isolation and characterization of an ω3-desaturation-defective mutant of an arachidonic acid-producing fungus, Mortierella alpina 1S-4. Arch Microbiol 161, 316-319.[CrossRef]
    [Google Scholar]
  15. Katayama, S. & Lee, J. B. ( 1993; ). Prostaglandins and leukotrienes. In Encyclopaedia of Food Science, Food Technology and Nutrition, pp. 3775-3781. Edited by R. Macrae, R. K. Robinson & M. J. Sadler. London: Academic Press.
  16. Kawashima, H., Nishihara, M., Hirano, Y., Kamada, N., Akimoto, K., Konishi, K. & Shimizu, S. ( 1997; ). Production of 5,8,11-eicosatrienoic acid (Mead acid) by a Δ6 desaturation activity-enhanced mutant derived from a Δ12 desaturase-defective mutant of an arachidonic acid-producing fungus, Mortierella alpina 1S-4. Appl Environ Microbiol 63, 1820-1825.
    [Google Scholar]
  17. Knutzon, D. S., Thurmond, J. M., Huang, Y.-S., Chaudhary, S., Bobik, E. G.Jr, Chan, G. M., Kirchner, S. J. & Mukerji, P. ( 1998; ). Identification of Δ5-desaturase from Mortierella alpina by heterologous expression in bakers’ yeast and canola. J Biol Chem 273, 29360-29366.[CrossRef]
    [Google Scholar]
  18. Leman, J. ( 1997; ). Oleaginous microorganisms: an assessment of the potential. Adv Appl Microbiol 43, 195-243.
    [Google Scholar]
  19. Meesters, P. A. E. P. & Eggink, G. ( 1996; ). Isolation and characterization of a Δ9-fatty acid desaturase gene from the oleaginous yeast Cryptococcus curvatus CBS 570. Yeast 12, 723-730.[CrossRef]
    [Google Scholar]
  20. Michaelson, L. V., Lazarus, C. M., Griffiths, G., Napier, J. A. & Stobart, A. K. ( 1998; ). Isolation of a Δ5-fatty acid desaturase gene from Mortierella alpina. J Biol Chem 273, 19055-19059.[CrossRef]
    [Google Scholar]
  21. Morrice, J., MacKenzie, D. A., Parr, A. J. & Archer, D. B. ( 1998; ). Isolation and characterisation of the acetyl-CoA carboxylase gene from Aspergillus nidulans. Curr Genet 34, 379-385.[CrossRef]
    [Google Scholar]
  22. Ratledge, C. ( 1993; ). Single cell oils – have they a biotechnological future? Trends Biotechnol 11, 278-284.[CrossRef]
    [Google Scholar]
  23. Sakuradani, E., Kobayashi, M. & Shimizu, S. ( 1999; ). Δ9-Fatty acid desaturase from arachidonic acid-producing fungus: unique gene sequence and its heterologous expression in a fungus, Aspergillus. Eur J Biochem 260, 208-216.[CrossRef]
    [Google Scholar]
  24. Sancholle, M. & Lösel, D. ( 1995; ). Lipids in fungal biotechnology. In The Mycota II: Genetics and Biotechnology, pp. 339-367. Edited by U. Kück. Berlin: Springer.
  25. Shanklin, J., Whittle, E. & Fox, B. G. ( 1994; ). Eight histidine residues are catalytically essential in a membrane-associated iron enzyme, stearoyl-CoA desaturase, and are conserved in alkane hydroxylase and xylene monooxygenase. Biochemistry 33, 12787-12794.[CrossRef]
    [Google Scholar]
  26. Stukey, J. E., McDonough, V. M. & Martin, C. E. ( 1989; ). Isolation and characterization of OLE1, a gene affecting fatty acid desaturation from Saccharomyces cerevisiae. J Biol Chem 264, 16537-16544.
    [Google Scholar]
  27. Stukey, J. E., McDonough, V. M. & Martin, C. E. ( 1990; ). The OLE1 gene of Saccharomyces cerevisiae encodes the Δ9 fatty acid desaturase and can be functionally replaced by the rat stearoyl-CoA desaturase gene. J Biol Chem 265, 20144-20149.
    [Google Scholar]
  28. Tabor, D. E., Xia, Y.-R., Mehrabian, M., Edwards, P. A. & Lusis, A. J. ( 1998; ). A cluster of stearoyl CoA desaturase genes, Scd 1 and Scd 2, on mouse chromosome 19. Mamm Genome 9, 341-342.[CrossRef]
    [Google Scholar]
  29. Vernet, T., Dignard, D. & Thomas, D. Y. ( 1987; ). A family of yeast expression vectors containing the phage f1 intergenic region. Gene 52, 225-233.[CrossRef]
    [Google Scholar]
  30. Wicker-Thomas, C., Henriet, C. & Dallerac, R. ( 1997; ). Partial characterization of a fatty acid desaturase gene in Drosophila melanogaster. Insect Biochem Mol Biol 27, 963-972.[CrossRef]
    [Google Scholar]
  31. Willatts, P., Forsyth, J. S., DiModugno, M. K., Varma, S. & Colvin, M. ( 1998; ). Influence of long-chain polyunsaturated fatty acids on infant cognitive function. Lipids 33, 973-980.[CrossRef]
    [Google Scholar]
  32. Yukawa, Y., Takaiwa, F., Shoji, K., Masuda, K. & Yamada, K. ( 1996; ). Structure and expression of two seed-specific cDNA clones encoding stearoyl-acyl carrier protein desaturase from sesame, Sesamum indicum L. Plant Cell Physiol 37, 201-205.[CrossRef]
    [Google Scholar]
  33. Zhang, L., Ge, L., Parimoo, S., Stenn, K. & Prouty, S. ( 1998; ). Characterization of the human stearoyl-CoA desaturase (SCD) gene and a processed SCD pseudogene. Mol Biol Cell 9, 191a (abstract).[CrossRef]
    [Google Scholar]
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