Purification of 2-hydroxyisoflavanone dehydratase from the cell cultures of Pueraria lobata

2-Hydroxyisoflavanone dehydratase, which catalyzes the final step of the formation of the isoflavonoid skeleton, was purified and characterized from yeast extract-elicited cell suspension cultures of Pueraria lobata. 2-Hydroxyisoflavanone, the substrate of the dehydratase, is the product of 2-hydroxyisoflavanone synthase, as cytochrome P-450 which catalyzes the hydroxylation step associated with aryl migration of flavanone. The dehydratase was purified to apparent homogeneity for the first time by a seven-step purification procedure. It is a single polypeptide with a molecular weight of 38 kDa, and has an isoelectric point at pH 5.1 and a pH optimum at 6.8. It required no co-factor, and the apparent Michaelis constant for 2,7,4'-trihydroxyisoflavanone was 7.0 mM. (C) 1998 Published by Elsevier Science Ltd. All rights reserved.