BIOSYNTHESIS OF C-18 POLYUNSATURATED FATTY-ACIDS IN MICROSOMAL MEMBRANE PREPARATIONS FROM THE FILAMENTOUS FUNGUS MUCOR-CIRCINELLOIDES

The biosynthesis of C-18 polyunsaturated fatty acids has been studied in the fungus Mucor circinelloides. Microsomal membrane preparations c ontained Delta 9, Delta 12 and Delta 6 desaturase activities. The Delt a 9 desaturase exhibited characteristics similar to those of the anima l and yeast Delta 9 desaturases in being membrane bound and utilising stearoyl-CoA as substrate. Cytochrome b(5) (a soluble form lacking the 20-amino-acid hydrophobic C-terminus) stimulated desaturation and was identified as a major cytochrome component of the membranes. A high f erricyanide reductase activity (indicative of NADH:cytochrome b, reduc tase activity) coupled to inhibition by cyanide further supported the similarity with the mammalian and yeast enzymes. Time-course studies w ith radiolabelled oleoyl-CoA showed that the oleate [18:1(9)] was tran sferred to position sn-2 of phosphatidylcholine (PtdCho) and was desat urated to linoleoyl-PtdCho. Removal of the excess oleoyl-CoA from the membranes prior to addition of reductant confirmed that oleoyl-PtdCho is a substrate for the Delta 12 desaturase. The entry of oleate at thi s position of the phospholipid was facilitated by the activity of lyso -PtdCho:acyl-CoA acyltransferase (LPCAT), which readily transferred ol eate from oleoyl-CoA to lyso-PtdCho. Desaturation of oleate at the sn- 1 position of PtdCho was also demonstrated after the entry of oleate i n to the phospholipid by the enzymes of the Kennedy pathway. Thus olea te at sn-1 and sn-2 positions served as substrate for the Delta 12 des aturase and is consistent with observations in oil seed tissues. LPCAT activity was substantially higher than that observed with lysophospha tidylethanolamine : acyl-CoA acyltransferase (LPEAT) indicating that o leate is less effectively channelled to phosphatidylethanolamine for l inoleate synthesis. No desaturation on phosphatidylinositol could be d emonstrated. Delta 6 desaturase utilised linoleate at the sn-2 positio n of exogenously supplied PtdCho presented to the membranes in the pre sence of reductant. Thus, the entry of substrates into PtdCho via LPCA T and the synthesis of linoleate [18:2(9,12)] and gamma-linolenate [18 :3(6,9,12)] on this phospholipid is similar to that reported for oil s eed membranes.