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 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) omega-3, as opposed to Delta-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 omega-6 double bonds, but the ability to desaturate substrates with omega-6 hydroxyl groups or omega-9 or omega-5 double bonds, and (d) a relative insensitivity to double bonds proximal to the carboxyl end of the substrate.
Reed
D W
DW
National Research Council of Canada, Plant Biotechnology Institute, 110 Gymnasium Place, Saskatoon, Saskatchewan, Canada.
Schäfer
U A
UA
Covello
P S
PS
D001483:Base Sequence; D001937:Brassica / Q000201:enzymology* / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D044943:Fatty Acid Desaturases / Q000235:genetics* / Q000378:metabolism*; D005227:Fatty Acids / Q000737:chemistry / Q000378:metabolism; D008401:Gas Chromatography-Mass Spectrometry; D015870:Gene Expression; D017343:Genes, Plant; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013379:Substrate Specificity; D014170:Transformation, Genetic
D017931:DNA Primers; D005227:Fatty Acids; D044943:Fatty Acid Desaturases; C078565:omega-3 fatty acid desaturase