Purification and Properties of Secondary Alcohol Oxidase from a Strain of Pseudomonas

Abstract

An enzyme which catalyzes the oxidation of poly(vinyl alcohol) has been purified to a homogeneous state from an enzyme preparation obtained from a culture broth of a strain of Pseudomonas and described previously as a polyvinyl alcohol)-degrading enzyme (Y. Wata-nabe et al., Agric. Biol. Chem., 39, 2447 (1975)). When the purified enzyme reacts on poly (vinyl alcohol), hydroxyl groups seem to be oxidized to keto groups and 1 mol of hydrogen peroxide is produced as 1 mol of molecular oxygen is consumed. Essentially no decrease is observed in the viscosity of poly(vinyl alcohol) solution. The enzyme is active not only to poly(vinyl alcohol) but also to a variety of low molecular weight secondary alcohols and 4-heptanone is formed from 4-heptanol. A name of secondary alcohol oxidase has been pro-posed to the enzyme. The enzyme is a single polypeptide with a molecular weight of 50, 000 with an isoelectric point at pH 10.3. Alanine and valine has been identified as N- and C-terminal residues, respectively. The enzyme is pink, has absorption maxima at 277, 364 and 466nm, and contains 1g atom of non-heme iron per molecule. No flavin has been detected. The enzyme is most active at pH 7 and at 50°C, and is stable between pH 4.5 and 9 and below 50°C. Among the compounds examined, Hg²⁺, Pb²⁺ Zn²⁺, o-phenanthroline and ethylene-diaminetetraacetate have shown weak inhibitions of the activity. The following two succes-sive reactions, the first is catalyzed by the secondary alcohol oxidase and the second by a pro-bable hydrolase have been postulated as a mechanism of bacterial degradation of poly (vinyl alcohol):
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