Isolation and characterization of genes from the marine microalga Pavlova salina encoding three front-end desaturases involved in docosahexaenoic acid biosynthesis

The fatty acid Δ4, Δ5 and Δ8-desaturase genes were cloned from marine microalga Pavlova salina, and the encoded enzymes produced omega-3 long chain polyunsaturated fatty acids (LC-PUFA) eicosatetraenoic acid (ETA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in yeast cells and/or tran...

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Bibliographic Details
Published in:Phytochemistry (Oxford) 2007-03, Vol.68 (6), p.785-796
Main Authors: Zhou, Xue-Rong, Robert, Stanley S., Petrie, James R., Frampton, Dion M.F., Mansour, Maged P., Blackburn, Susan I., Nichols, Peter D., Green, Allan G., Singh, Surinder P.
Format: Article
Language:English
Subjects:
Algal Proteins - genetics
Algal Proteins - metabolism
Base Sequence
Biological and medical sciences
Blotting, Southern
Chemical constitution
Chromatography, Gas
Cloning, Molecular
complementary DNA
conserved sequences
DNA, Complementary - chemistry
DNA, Complementary - genetics
docosahexaenoic acid
Docosahexaenoic Acids - metabolism
eicosapentaenoic acid
eicosatetraenoic acid
Eukaryota - enzymology
Eukaryota - genetics
fatty acid composition
Fatty Acid Desaturases - genetics
Fatty Acid Desaturases - metabolism
Fatty Acids, Unsaturated - metabolism
Fundamental and applied biological sciences. Psychology
Gas Chromatography-Mass Spectrometry
gene expression regulation
Gene Expression Regulation, Enzymologic
Gene isolation
Isoenzymes - genetics
Isoenzymes - metabolism
long chain fatty acids
Long chain polyunsaturated fatty acids
Metabolism
Metabolism. Physicochemical requirements
Microalgae
Models, Biological
molecular cloning
Molecular Sequence Data
nucleotide sequences
Pavlova
Pavlova salina
Phylogeny
Plant physiology and development
Polymerase Chain Reaction
polyunsaturated fatty acids
Reverse Transcriptase Polymerase Chain Reaction
sequence analysis
Sequence Analysis, DNA
stearoyl-CoA desaturase
Δ4-desaturase
Δ5-desaturase
Δ8-desaturase
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Description
Summary:The fatty acid Δ4, Δ5 and Δ8-desaturase genes were cloned from marine microalga Pavlova salina, and the encoded enzymes produced omega-3 long chain polyunsaturated fatty acids (LC-PUFA) eicosatetraenoic acid (ETA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in yeast cells and/or transgenic seeds, respectively, with appropriate substrates. The marine microalga Pavlova salina produces lipids containing approximately 50% omega-3 long chain polyunsaturated fatty acids (LC-PUFA) such as docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA). Three cDNA sequences, designated PsD4Des, PsD5Des, PsD8Des, were isolated from P. salina and shown to encode three front-end desaturases with Δ4, Δ5 and Δ8 specificity, respectively. Southern analysis indicated that the P. salina genome contained single copies of all three front-end fatty acid desaturase genes. When grown at three different temperatures, analysis of fatty acid profiles indicated P. salina desaturation conversions occurred with greater than 95% efficiency. Real-Time PCR revealed that expression of PsD8Des was higher than for the other two genes under normal growth conditions, while PsD5Des had the lowest expression level. The deduced amino acid sequences from all three genes contained three conserved histidine boxes and a cytochrome b 5 domain. Sequence alignment showed that the three genes were homologous to corresponding desaturases from other microalgae and fungi. The predicted activities of these three front-end desaturases leading to the synthesis of LC-PUFA were also confirmed in yeast and in higher plants.
ISSN:0031-9422
1873-3700
DOI:10.1016/j.phytochem.2006.12.016