Production of Daidzein and Genistein from Seed and Root Extracts of Korean Wild Soybean (Glycine soja) by Thermostable β-Galactosidase from Thermoproteus uzoniensis

Abstract

Isoflavone glycosides are commonly biotransformed into isoflavone aglycones due to the superior biological activities of the latter. Wild soybeans contain a higher isoflavone content than domesticated soybeans due to their high level of genetic diversity. In this study, we cloned and characterized a thermostable β-galactosidase from the extreme thermophile Thermoproteus uzoniensis for potential application in isoflavone conversion in Korean wild soybeans. The purified recombinant enzyme exhibited a maximum specific activity of 1103 µmol/min/mg at pH 5.0 and 90◦C with a half-life of 46 h and exists as a homodimer of 113 kDa. The enzyme exhibited the highest activity for p-nitrophenyl (pNP)-β-D-galactopyranoside among aryl glycosides and it hydrolyzed isoflavone glycosides in the order genistin > daidzin > ononin > glycitin. The enzyme completely hydrolyzed 2.77 mM daidzin and 3.85 mM genistin in the seed extract of wild soybean after 80 and 70 min with productivities of 1.86 and 3.30 mM/h, respectively, and 9.89 mM daidzin and 1.67 mM genistin in the root extract after 180 and 30 min, with the highest productivities of 3.30 and 3.36 mM/h, respectively, compared to other glycosidases. Our results will contribute to the industrial production of isoflavone aglycone using wild soybean and this is the first report on the enzymatic production of isoflavone aglycones from isoflavone glycosides in wild soybeans.