Kinetics and mechanism of heterogeneous hydrolysis of poly[(R)-3-hydroxybutyrate]fielm by PHA depolymerases

The kinetics and mechanism of enzymatic degradation on the surface of poly[(R)-3-hydroxybutyrate](PC(R)-3HB]) jilm have been studied using three types of extracellular poly(hydroxyalkanoate)(PHA)depolymerases from Alcaligenes faecalis, Pseudomonas pickettii and Comamonas testosteroni. The monomer and dimer of 3-hydroxybutyric acid were produced during the course of the enzymatic degradation of PC(R)-3HB]film, and the rate of production was determined by monitoring the increase in absorbance at 210 nm on a spectrophotometer. The rate of enzymatic degradation increased to a maximum value with the concentration of PHA depolymerase, followed by a gradual decrease. The kinetic data were accountedfor in terms of a heterogeneous enzymatic reaction, involving enzymatic degradation on the surface of PC(R)-3HB]jilm via two steps of adsorption and hydrolysis by a PHA depolymerase with binding and catalytic domains. The kinetic results suggest that the properties oj-the catalytic domains are very similar among the three PHA depolymerases, but that those of the binding domains are strongly dependent on the type of depolymerase.