1. Synthesis of 13R,20-dihydroxy-docosahexaenoic acid by site-directed mutagenesis of lipoxygenase derived from Oscillatoria nigro-viridis PCC 7112
- Author
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Jeong-Woo Seo, Baek-Rock Oh, Jong-Jae Yi, Sun-Yeon Heo, Woo Sung Son, and Jung-Hyun Ju
- Subjects
0301 basic medicine ,Docosahexaenoic Acids ,Linoleic acid ,Biophysics ,Biochemistry ,03 medical and health sciences ,Lipoxygenase ,chemistry.chemical_compound ,0302 clinical medicine ,Biosynthesis ,Humans ,Molecular Biology ,chemistry.chemical_classification ,Alanine ,integumentary system ,biology ,food and beverages ,Stereoisomerism ,Cell Biology ,Lipoxygenases ,Amino acid ,030104 developmental biology ,Enzyme ,chemistry ,Oscillatoria ,Docosahexaenoic acid ,030220 oncology & carcinogenesis ,Mutagenesis, Site-Directed ,biology.protein ,lipids (amino acids, peptides, and proteins) ,Polyunsaturated fatty acid - Abstract
Lipoxygenases (LOXs) are implicated in the biosynthesis of pro- and anti-inflammatory lipid mediators involved in immune cell signaling, most of which catalyze peroxidation of polyunsaturated fatty acids by distinct regio- and stereoselectivity. Current reports suggested that conserved amino acid, Gly in R-LOXs and Ala in S-LOXs, in the catalytic domain play an important role in determining the position as well as the stereochemistry of the functional group. Recently, we have confirmed that the catalytic specificity of cyanobacterial lipoxygenase, named Osc-LOX, with alanine at 296 was 13S-type toward linoleic acid, and producing a 17S- hydroxy-docosahexaenoic acid from docosahexaenoic acid (DHA). Here, we aimed to change the catalytic property of LOX from13S-LOX to 9R-LOX by replacing Ala with Gly and to produce a lipid mediators different from the wild-type using DHA. Finally, we succeeded in generating human endogenous a 13R-hydroxy-docosahexaenoic acid and a 13R,20-dihydroxy-docosahexaenoic acid from DHA through an enzymatic reaction using the Osc-LOX-A296G. Our study could enable physiological studies and pharmaceutical research for the 13R,20-dihydroxy-docosahexaenoic acid.
- Published
- 2020