Pulsed light inactivation of polygalacturonase

Abstract

Pulsed light is a non-thermal technology capable to inactivate enzymes. This study investigated the effect of pulsed light on the activity of polygalacturonase, and on the structure of the enzyme by means of fluorescence emission spectra, free sulphydryl detection and analysis of changes in parameter A and phase-diagram. The results showed that pulsed light is able to inactivate polygalacturonase in buffer, with > 90 % reduction of enzyme activity after applying 128 J/cm2 and a first-order kinetic constant of 0.0426 cm2/J under the experimental conditions. The free sulfhydryl detection revealed the rupture of sulfhydryl bridges. Fluorescence spectra analysis showed that the tertiary structure of polygalacturonase was changed. Phase diagram analysis shows the existence of only two populated states. It is suggested that the inactivation of polygalacturonase by pulsed light is an all-or-none process where disulfide bridges are broken and the enzyme is unfolded.