Purification and Properties of Oxidized Poly(vinyl a1cohol)-Degrading Enzyme

Abstract

An enzyme catalyzing the degradation of secondary alcohol oxidase-oxidized poly(vinyl alcohol), in which hydroxyl groups of poly(vinyl alcohol) are partially converted to keto groups, was purified to an electrophoretically homogeneous state from a mixed culture broth of at least three different soil bacteria. The enzyme was active to the oxidized poly(vinyl alcohol), but not to intact poly(vinyl alcohol) and to a variety of examined low molecular weight keto compounds. The enzyme was, therefore, tentatively called oxidized poly(vinyl alcohol)-degrading enzyme. The enzyme was a single polypeptide having a molecular weight of 38,000. The N- and C-terminal amino acids were alanine and threonine, respectively. The isoelectric point was pH 10.0. The optimum pH for activity was 6.5 and the optimum temperature 45°C. The enzyme activity was inhibited by Hg2+ and recovered by reduced glutathione, although p-chloromercuribenzoate had no effect. The enzyme reaction on oxidized poly(vinyl alcohol) required neither oxygen nor other electron acceptors, and resulted in a rapid decrease in viscosity, a fall of pH and an increase in compounds that are positive to a color reaction specific to carboxylic acids. The infrared spectrum of reaction products also showed the presence of carboxylic acids. Based on these results, the reaction catalyzed by the enzyme has been suggested to be hydrolytic cleavages of the main chain of oxidized poly(vinyl alcohol) as in the equation: An enzymatic system of poly(vinyl alcohol) degradation was reconstructed by use of purified samples of secondary alcohol oxidase and oxidized poly(vinyl alcohol)-degrading enzyme.