Biotransformation of Polyunsaturated Fatty Acids to Trioxilins by Lipoxygenase from Pleurotus sajor‐caju

Abstract

A lipoxygenase from Pleurotus sajor‐caju (PsLOX) was cloned, expressed in Escherichia coli, and purified as a soluble protein with a specific activity of 629 μmol/min/mg for arachidonic acid (AA). The native PsLOX exhibited a molecular mass of 146 kDa, including a 73‐kDa homodimer, as estimated by gel‐filtration chromatography. The major products converted from polyunsaturated fatty acids (PUFAs), including AA, eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA), were identified as trioxilins (TrXs), namely 13,14,15‐TrXB3, 13,14,15‐TrXB4, and 15,16,17‐TrXB5, respectively, through high‐performance liquid chromatography (HPLC) and liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) analyses. The enzyme displayed its maximum activity at pH 8.0 and 20 °C. Under these conditions, the specific activity and catalytic efficiency of PsLOX for PUFAs exhibited the following order: AA>EPA>DHA. Based on HPLC analysis and substrate specificity, PsLOX was identified as an arachidonate 15‐LOX. PsLOX efficiently converted 10 mM of AA, EPA, and DHA to 8.7 mM of 13,14,15‐TrXB3 (conversion rate: 87 %), 7.9 mM of 13,14,15‐TrXB4 (79 %), and 7.2 mM of 15,16,17‐TrXB5 (72 %) in 15, 20, and 20 min, respectively, marking the highest conversion rates reported to date. Collectively, our results demonstrate that PsLOX is an efficient TrXs‐producing enzyme.