The role of enzyme adsorption in the enzymatic degradation of an aliphatic polyester

Abstract

The enzyme catalyzed degradation of poly(3-hydroxybutyrate) (PHB) is a two-step process consisting of the adsorption of the enzyme on the surface of a PHB substrate and the cleavage of ester bonds. A deactivated enzyme was prepared by point mutagenesis to separate the two steps from each other. Measurements carried out with active and inactive enzymes on PHB particles proved that mutagenesis was successful and the modified enzyme did not catalyze degradation. Based on the Michaelis-Menten approach a kinetic model was proposed which could describe the processes quantitatively, the agreement between prediction and the measured data was excellent. The separation of the two processes allowed the determination of the adsorption kinetics of the enzyme; the rate constants of the adsorption and desorption process were determined for the first time. Comparison of these constants to reaction rates showed that adsorption is not instantaneous and can be the rate-determining step. The area occupied by an enzyme molecule was also determined (13.1 nm2) and it was found to be smaller than the value published in the literature (17 ± 8 nm2). The separation of the two steps makes possible the prediction and control of the degradation process.