期刊
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
卷 24, 期 16, 页码 -出版社
MDPI
DOI: 10.3390/ijms241612977
关键词
lipoxygenase; molecular cloning; & omega;3 fatty acids; & omega;3(S) dioxygenation; fatty acid hydroperoxides; cucumber (Cucumis sativus L.)
The profiling of endogenous oxylipins in cucumber revealed the abundance of 16-hydroxy-9,12,14-octadecatrienoic acid. The homogenates of cucumber organs yielded 16(S)-hydroperoxide as a predominant product when incubated with a-linolenic acid. The targeted proteomic analysis identified CsLOX3 as the first 16(S)-LOX and w3-LOX that synthesizes 16(S)-HPOT.
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-9,12,14octadecatrienoic acid (16-HOT). Incubations of homogenates from these organs with a-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 a-linolenic and linoleic acids, respectively. Furthermore, a-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, 1H-NMR, 2D 1H -1H-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 w3-LOX ever discovered. Eicosapentaenoic and hexadecatrienoic acids were also specifically transformed to the corresponding w3(S)-hydroperoxides by CsLOX3.
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