Characterization of the Brassica napus Extraplastidial Linoleate Desaturase by Expression in Saccharomyces cerevisiae

The substrate specificity and regioselectivity of the Brassica napus extraplastidial linoleate desaturase (FAD3) was investigated in vivo in a heterologous expression system. A strain of the yeast Saccharomyces cerevisiae producing the plant enzyme was constructed and cultured in media containing a...

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Bibliographic Details
Published in:Plant physiology (Bethesda) 2000-03, Vol.122 (3), p.715-720
Main Authors: REED, D. W, SCHÄFER, U. A, COVELLO, P. S
Format: Article
Language:English
Subjects:
Agronomy. Soil science and plant productions
Base Sequence
Biochemistry and Macromolecular Structure
Biological and medical sciences
Brassica - enzymology
Brassica - genetics
Chemical desaturation
DNA Primers - genetics
Economic plant physiology
Enzyme substrates
Enzymes
Fatty Acid Desaturases - genetics
Fatty Acid Desaturases - metabolism
Fatty acids
Fatty Acids - chemistry
Fatty Acids - metabolism
Fundamental and applied biological sciences. Psychology
Gas Chromatography-Mass Spectrometry
Gene Expression
Genes, Plant
Lipids
Metabolism
Nutrition. Photosynthesis. Respiration. Metabolism
Plant physiology and development
Plastids
Saccharomyces cerevisiae
Saccharomyces cerevisiae - genetics
Substrate Specificity
Substrates
Transformation, Genetic
Unsaturated fatty acids
Yeasts
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Summary:The substrate specificity and regioselectivity of the Brassica napus extraplastidial linoleate desaturase (FAD3) was investigated in vivo in a heterologous expression system. A strain of the yeast Saccharomyces cerevisiae producing the plant enzyme was constructed and cultured in media containing a variety of fatty acids. The products of desaturation of these potential substrates were determined by gas chromatographic and mass spectrometric analysis of the yeast cultures. The results indicate that the enzyme has: (a) ω-3, as opposed to Δ-15 or double-bond-related regioselectivity, (b) the ability to desaturate substrates in the 16 to 22 carbon range, (c) a preference for substrates with ω-6 double bonds, but the ability to desaturate substrates with ω-6 hydroxyl groups or ω-9 or ω-5 double bonds, and (d) a relative insensitivity to double bonds proximal to the carboxyl end of the substrate.
ISSN:0032-0889
1532-2548
DOI:10.1104/pp.122.3.715