Differential effect of N-ethyl maleimide on delta6-desaturase activity in human fetal liver toward fatty acids of the n-6 and n-3 series.
The effect of N-ethyl-maleimide (NEM) on delta5- and delta6-desaturase activities and the incorporation of substrates and products into different microsomal lipid classes and phospholipid (PL) subclasses were studied in human fetal liver microsomes, obtained after legally approved therapeutic abortion. Desaturase activities were measured by a radiochemical method using reversed-phase high-performance liquid chromatography (HPLC). After nonphospholipid (NPL) and PL separation on silica cartridges, the radioactivity in different lipids of the NPL group was assessed by two-dimensional thin-layer chromatography, and their fatty acid (FA) composition by gas-liquid chromatography. The PL subclasses were separated, and the distribution of radioactivity between products and substrates was determined in PL subclasses. NEM inhibited the delta5- and delta6-desaturase activities in the n-6 series of FA but not the delta6-desaturase activity in the n-3 series, which suggests the existence of two distinct delta6-desaturases, one for the n-6 series and another for the n-3 series. Whether NEM was present or absent, most of the radioactivity was recovered in the free FA form (about 80%). The desaturation products, obtained in the presence or absence of NEM, were preferentially incorporated into PL, suggesting a channeling of the newly synthesized FA toward microsomal PL. The comparison of the distribution of substrates and products incorporated into the different PL classes showed that most of the labeled FA were incorporated into phosphatidylcholine and to a lesser degree into phosphatidylethanolamine.