Purification and Properties of an NADPH-Aldose Reductase (Aldehyde Reductase) from Euonymus japonica Leaves

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Abstract
The enzyme aldose (aldehyde) reductase was partially purified (142-fold) and characterized from Euonymus japonica leaves. The reductase, a dimer, had an average molecular weight of 67,000 as determined by gel filtration on Sephadex G-100. The enzyme was NADPH specific and reduced a broad range of substrates including aldoses, aliphatic aldehydes, and aromatic aldehydes. Maximum activity was observed at pH 8 in phosphate and Tris-HCl buffers and at pH 8.6 to 9.0 in glycine-NaOH buffer using DL-glyceraldehyde or 3-pyridinecarboxaldehyde as substrate. NADP was a competitive inhibitor with respect to NADPH with a Ki of 60 micromolar. Glycerol was an uncompetitive inhibitor to DL-glyceraldehyde (K''i = 460 millimolar). The Euonymus enzyme was inhibited by sulfhydryl inhibitor, phenobarbital, and high concentrations of Li2SO4. Pyrazol and metal chelating agents inhibited the enzyme slightly. Enzyme activity was detected in the leaves and berries of Celastrus orbiculatus and several species of Euonymus. Probable function of this enzyme is to reduce D-galactose to galactitol, a characteristic metabolite in phloem sap of members of the Celastraceae family.