Whole-Cell Production of Resolvin E2 and 17R-Resolvin D5 by a Double-Oxygenating 15S-Lipoxygenase With...

Resolvin E2 (RvE2; 5S,18R-dihydroxyeicosapentaenoic acid) and 17R-resolvin D5 (17R-RvD5; 7S,17R-dihydroxydocosahexaenoic acid) are specialized pro-resolving mediators (SPMs) that actively promote the resolution of inflammation. However, their efficient biocatalytic production remains challenging because of the limited catalytic performance and poor stability of previously reported 5S-lipoxygenases (5S-LOXs). Here, we identified and characterized a novel double-oxygenating 15S-LOX from Pseudobdellovibrionaceae bacterium exhibiting 5S-LOX activity and evaluated its potential as a whole-cell biocatalyst for SPM production. Recombinant Escherichia coli cells expressing this enzyme stereoselectively converted 18R-hydroxyeicosapentaenoic acid (18R-HEPE) and 17R-hydroxydocosahexaenoic acid (17R-HDHA), prepared using engineered 18R- and 15R-LOXs from Sorangium cellulosum, into RvE2 and 17R-RvD5, respectively. Product identities were confirmed by chiral-phase HPLC and LC-MS/MS analyses. Reaction conditions, including pH, temperature, and DMSO, cell, and substrate concentrations, were optimized to enhance biotransformation performance. Under optimized conditions, 4.0 mM 18R-HEPE and 17R-HDHA were converted into 0.57 mM (190.61 mg/L) RvE2 and 1.48 mM (533.5 mg/L) 17R-RvD5 within 60 and 30 min, respectively. RvE2 production was 2.4-fold higher than that obtained using a previously reported purified 5S-LOX, and this study provides the first quantitative report of 17R-RvD5 production. These findings establish a double-oxygenating 15S-LOX with 5S-LOX activity as an efficient whole-cell platform for SPM production.