Abstract
Soluble epoxide hydrolase (sEH) is a bifunctional enzyme that has a C-terminus epoxide hydrolase domain and an N-terminus phosphatase domain. The endogenous substrates of epoxide hydrolase are known to be epoxyeicosatrienoic acids, but the endogenous substrates of the phosphatase activity are not well understood. In this study, to explore the substrates of sEH, we investigated the inhibition of the phosphatase activity of sEH toward 4-methylumbelliferyl phosphate by using lecithin and its hydrolyzed products. Although lecithin itself did not inhibit the phosphatase activity, the hydrolyzed lecithin significantly inhibited it, suggesting that lysophospholipid or fatty acid can inhibit it. Next, we investigated the inhibition of phosphatase activity by lysophosphatidyl choline, palmitoyl lysophosphatidic acid, monopalmitoyl glycerol, and palmitic acid. Palmitoyl lysophosphatidic acid and fatty acid efficiently inhibited phosphatase activity, suggesting that lysophosphatidic acids (LPAs) are substrates for the phosphatase activity of sEH. As expected, palmitoyl, stearoyl, oleoyl, and arachidonoyl LPAs were efficiently dephosphorylated by sEH (Km, 3-7 μM; Vmax, 150-193 nmol/min/mg). These results suggest that LPAs are substrates of sEH, which may regulate physiological functions of cells via their metabolism.
Highlights
Soluble epoxide hydrolase is a bifunctional enzyme that has a C-terminus epoxide hydrolase domain and an N-terminus phosphatase domain
To explore the substrates for the phosphatase activity of Soluble epoxide hydrolase (sEH), we investigated the inhibitory effect of lecithin from egg yolk and its hydrolyzed products with a synthetic substrate, 4-methylumbelliferyl phosphate (4-MUP)
Because lipid phosphates can be substrates of phosphatase, p-lysophosphatidic acid (LPA) was used in this assay. palmitoyl LPA (p-LPA) inhibited the phosphatase activity of sEH, but the dephosphorylated product, palmitoyl glycerol, did not (Fig. 1B)
Summary
Soluble epoxide hydrolase (sEH) is a bifunctional enzyme that has a C-terminus epoxide hydrolase domain and an N-terminus phosphatase domain. To explore the substrates of sEH, we investigated the inhibition of the phosphatase activity of sEH toward 4-methylumbelliferyl phosphate by using lecithin and its hydrolyzed products. Palmitoyl lysophosphatidic acid and fatty acid efficiently inhibited phosphatase activity, suggesting that lysophosphatidic acids (LPAs) are substrates for the phosphatase activity of sEH. Palmitoyl, stearoyl, oleoyl, and arachidonoyl LPAs were efficiently dephosphorylated by sEH (Km, 3–7 M; Vmax, 150–193 nmol/min/mg) These results suggest that LPAs are substrates of sEH, which may regulate physiological functions of cells via their metabolism.—Oguro, A., and S. Newman et al [7] hypothesized that dihydroxy fatty acid phosphates are substrates for the phosphatase of sEH because sEH metabolizes the EETs or linoleic acid epoxide to form the corresponding diols.
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