Yutaka Ebizuka

An inducible hydrolase from Mortierella isabellina (basidiomycetes). The deacylation of (+)-usnic acid

Abstract An inducible enzyme catalysing the hydrolysis of (+)-usnic acid to (+)-2-desacetylusnic acid and acetic acid has been purified 150-fold from the mycelium of Mortierella isabellina grown in the presence of (+)-usnic acid. Purification was achieved by treatment with protamine sulfate, (NH 4 ) 2 SO 4 fractionation, negative adsorption on alumina Cγ gel and hydroxylapatite followed by chromatography on DEAE-cellulose and Sephadex G-200. The elution pattern from a Sephadex G-200 column indicated a MW of ca 7.6 × 10 4 for the enzyme. The apparent K m value for (+)-usnic acid at the pH optimum (pH 7) was 4.0 × 10 −5 M. The enzyme was specific for (+)-usnic acid and inactive towards (−)-usnic acid, (+)-isousnic acid or certain phloracetophenone derivatives. Its activity was enhanced in the presence of divalent metal ions such as Co 2+ , Ni 2+ , Mn 2+ , Mg 2+ and Zn 2+ .

An inducible lactone hydrolase yielding 2,5-diphenyl-3-hydroxy-4-oxo-2-hexendioic acid from pulvinic acid

Abstract The metabolism of vulpinic acid by an unclassified soil micro-organism was studied. A new compound, 2,5-diphenyl-3-hydroxy-4-oxo-2-hexendioic acid (DHOHA) was isolated from the reaction mixture of a cell-free preparation and pulvinic acid. The existence of a hydrolase which catalyses the conversion of vulpinic acid to pulvinic acid was detected in cell-free preparation, and an inducible lactone hydrolase capable of converting pulvinic acid to DHOHA was purified 130-fold and characterized. This enzyme had a MW of ca 34 000, a K m for pulvinic acid at pH optimum (pH 7.0) less than 10 − 6 M, p I = 5.0, and was inhibited by p -chloromercuriphenylsulfonate and diethylpyrocarbonate. The enzyme was highly specific for pulvinic acid. The initial degradative steps proposed for this organism are vulpinic acid → pulvinic acid → DHOHA.