Die Enzymologie der C8‐Oxylipinbiosynthese

Abstract

Based on transcriptomic, volatilomic and metabolomic data of the model organism Cyclocybe aegerita, the endogenous and enzymatic production of the C8‐oxylipins oct‐2‐en‐1‐ol, octan‐ 1‐ol, oct‐2‐enal, octanal, octan‐3‐ol, oct‐1‐en‐3‐ol, oct‐1‐en‐3‐one and octan‐3‐ one via biotransformations with [U‐13C]linoleic acid was compared to the developmental state depending gene expression patterns of putative oxylipin‐associated enzyme classes like lipoxygenases, dioxygenases, ene‐reductases and alcohol dehydrogenases. On closer consideration of these data sets, the C8‐oxylipin production seemed to occur cluster‐like.While one cluster (ketonic‐cycle) consisted of octan‐3‐ol, oct‐1‐en‐3‐one, octan‐3‐one and oct‐1‐en‐3‐ol, which were detected through out all developmental stages, a second cluster (aldehydic‐cycle) primarily active at early developmental stages involved oct‐2‐enal, octanal, oct‐2‐en‐1‐ol and octan‐1‐ol. Studying these clusters revealed that all C8‐oxylipins of the aldehydic‐cycle and ketonic‐cycle can be derived from oct‐2‐enal and oct‐1‐en‐3‐one, respectively. A detailled consideration of the gene expression patterns revealed several enzyme classes putatively involved in the C8‐oxylipin biosynthesis. Especially the two genes AAE3 04864 and AAE3 13549 encoding the lox4‐ and enr1‐gene showed great similarities between their strongly increasing transcript counts with maturation. Subsequent recombinant production of CaeLOX4 and CaeEnR1 revealed that CaeLOX4 showed a preference for C18‐ fatty acids with the highest affinity towards linoleic acid. Analysis of the reaction product revealed that this LOX selectively oxygenated linoleic acid at position 13 to 13‐ hydroperoxyoctadecadienoic acid (13‐HPOD). CaeEnR1 showed high affinities towards the reduction of oct‐1‐en‐3‐one and oct‐2‐enal to their saturated counterparts octan‐3‐one and octanal, respectively. Due to the strong similarities between the expression patterns of CaeLOX4 and CaeEnR1 a coherence between their main reaction products 13‐HPOD and the precursor of the ketonic‐cycle oct‐1‐en‐3‐one seems very likely. This suggests a biosynthetic route from linoleic acid to 13‐HPOD, followed by a subsequent cleavage step to oct‐1‐en‐3‐ one and therefore to C8‐oxylipins of the ketonic‐cycle.