DETECTION AND DIFFERENTIATION OF PECTIC ENZYME-ACTIVITY IN-VITRO AND IN-VIVO BY CAPILLARY ELECTROPHORESIS OF PRODUCTS FROM FLUORESCENT-LABELED SUBSTRATE

A sensitive assay is described for the detection of pectate-depolymeri zing enzymes using capillary electrophoresis of a fluorescent end-labe led pectate oligomer. The labeled oligomer is allowed to react with th e enzyme either in vitro or in vivo, such as inside the intercellular spaces of a cotton cotyledon, and after an appropriate incubation time the products are analyzed by capillary electrophoresis. The site and mode of action of the pectate-depolymerizing activity can be inferred from the products. Both endo- and exopolygalacturonase activity, and l yase activity, were distinguished. Since only the fluorescent oligomer and products from its labeled reducing end are detected, there is no interference from other compounds; only pectic enzyme activity is dete cted. By this type of analysis we can show that there is considerable endo- and exopolygalacturonase activity in the intercellular spaces of cotton cotyledons.