Discovery of α-Linolenic Acid 16 Vegetative Lipoxygenase 3

• Published in: International journal of molecular sciences 2023-08, Vol.24 (16)
• Main Authors: Gorina, Svetlana S, Egorova, Alevtina M, Lantsova, Natalia V, Toporkova, Yana Y, Grechkin, Alexander N
• Format: Article
• Language: English
• Subjects: Enzymes; High performance liquid chromatography; Linolenic acids; Physiological aspects
• ISSN: 1422-0067
• DOI: 10.3390/ijms241612977

The GC-MS profiling of the endogenous oxylipins (Me/TMS) from cucumber (Cucumis sativus L.) leaves, flowers, and fruit peels revealed a remarkable abundance of 16-hydroxy-9,12,14-octadecatrienoic acid (16-HOT). Incubations of homogenates from these organs with α-linolenic acid yielded 16(S)-hydroperoxide (16-HPOT) as a predominant product. Targeted proteomic analyses of these tissues revealed the presence of several highly homologous isoforms of the putative “9S-lipoxygenase type 6”. One of these isoenzymes (CsLOX3, an 877 amino acid polypeptide) was prepared by heterologous expression in E. coli and exhibited 16(S)- and 13(S)-lipoxygenase activity toward α-linolenic and linoleic acids, respectively. Furthermore, α-linolenate was a preferred substrate. The molecular structures of 16(S)-HOT and 16(S)-HPOT (Me or Me/TMS) were unequivocally confirmed by the mass spectral data, [sup.1]H-NMR, 2D [sup.1]H-[sup.1]H-COSY, TOCSY, HMBC, and HSQC spectra, as well as enantiomeric HPLC analyses. Thus, the vegetative CsLOX3, biosynthesizing 16(S)-HPOT, is the first 16(S)-LOX and ω3-LOX ever discovered. Eicosapentaenoic and hexadecatrienoic acids were also specifically transformed to the corresponding ω3(S)-hydroperoxides by CsLOX3.