Oleate 6-hydroxylase activity in the membrane fraction from an oleaginous fungus, Mortierella ramanniana var. angulispora.
Conversion of [1-14C]oleoyl-CoA (OA-CoA) in subcellular fractions of an oleaginous fungus, Mortierella ramanniana var. angulispora (IFO 8187) was investigated. The membrane fraction actively catalyzed the transformation of OA-CoA into one metabolite, which was identified as 6-hydroxy oleic acid (6-HOOA) by GC-MS analysis. The enzyme activity to form 6-HOOA was dependent on OA-CoA concentrations and increased linearly with incubation time and protein concentrations under appropriate experimental conditions. Since the enzyme activity required molecular oxygen and reduced pyridine nucleotides (NADH and NADPH), it was most probable that the formation of 6-HOOA was ascribed to direct hydroxylation to the C6 position, oleate 6-hydroxylase. 6-HOOA was formed from free oleic acid (OA) and OA-NH4 salt as substrate as well as OA-CoA, but not from OA covalently bound to phosphatidylcholine or triacylglycerol. Since CoA stimulated the hydroxylation of free OA and OA-NH4 salt but not that of OA-CoA, OA may be converted to OA-CoA and then hydroxylated. The enzyme activity was distinctly reduced by addition of lysophosphatidic acid, phosphatidic acid and dithiothreitol. Since hydroxy fatty acids are very minor components in this fungus, it may raise the possibility that the 6-hydroxylase activity works for other functions such as comprising one step for desaturation as well as forming the hydroxy fatty acids.
Certik
M
M
Applied Microbiology Department, National Institute of Bioscience and Human-Technology, Ibaraki-ken, Japan.
Nakahara
T
T
Kamisaka
Y
Y
D000214:Acyl Coenzyme A / Q000378:metabolism*; D006900:Hydroxylation; D008566:Membranes / Q000201:enzymology; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D009090:Mucorales / Q000201:enzymology*; D009829:Oleic Acids / Q000096:biosynthesis*; D013347:Subcellular Fractions / Q000201:enzymology; D013379:Substrate Specificity
C102523:6-hydroxyoleic acid; D000214:Acyl Coenzyme A; D009829:Oleic Acids; C017585:oleoyl-coenzyme A; D006899:Mixed Function Oxygenases