Abstract
Objective
To quantitatively hydroxylate 8S- and 10S-positions on polyunsaturated fatty acids by recombinant Escherichia coli cells expressing mouse arachidonate 8S-lipoxygenase (8S-LOX).
Results
Hydroxylated products gained from the conversion of arachidonic acid (20:4Δ5Z,8Z,11Z,14Z, AA), eicosapentanoic acid (20:5Δ5Z,8Z,11Z,14Z,17Z, EPA), and (22:6Δ4Z,7Z,10Z,13Z,16Z,19Z, DHA) by recombinant E. coli cells containing 8S-LOX from mouse were identified as 8S-hydroxy-5,9,11,14(Z,E,Z,Z)-eicosatetranoic acid (8S-HETE), 8S-hydroxy-5,9,11,14,17(Z,E,Z,Z,Z)-eicosapentanoic acid (8S-HEPE), and 10S-hydroxy-4,8,12,14,16,19(Z,E,Z,Z,Z,Z)-docosahexaenoic acid (10S-HDoHE), respectively. Under the optimal hydroxylation conditions of pH 7.5, 30 °C, 5% (v/v) ethanol, 15 g cells l−1, and 5 mM substrate, AA, EPA, and DHA were hydroxylated into 4.37 mM 8S-HETE, 3.77 mM 8S-HEPE, and 3.13 mM 10S-HDoHE for 60, 90, and 60 min, with 87, 75, and 63% molar conversions, respectively.
Conclusion
To the best of our knowledge, this is the first quantitatively biotechnological production of 8S-HETE, 8S-HEPE, and 10S-HDoHE.
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Acknowledgements
This paper was written as part of Konkuk University’s research support program for its faculty on sabbatical leave in 2018.
Supporting information
Supplementary Fig. 1—SDS-PAGE analysis of mouse 8S-LOX at each purification step.
Supplementary Fig. 2—Chirality analysis of 8-HETE and 8-HEPE produced by recombinant E. coli cells expressing mouse 8S-LOX.
Supplementary Fig. 3—Investigation of pH and temperature to optimize the production of 8S-HETE from AA by recombinant E. coli cells expressing mouse 8S-LOX.
Supplementary Fig. 4—Investigation of enzyme stability to optimize the production of 8S-HETE from AA by recombinant E. coli cells expressing mouse 8S-LOX.
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Shin, KC., Kang, WR., Seo, MJ. et al. Production of 8S- and 10S-hydroxy polyunsaturated fatty acids by recombinant Escherichia coli cells expressing mouse arachidonate 8S-lipoxygenase. Biotechnol Lett 41, 575–582 (2019). https://doi.org/10.1007/s10529-019-02659-5
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DOI: https://doi.org/10.1007/s10529-019-02659-5