Partial deglycosylation of an anionic isoperoxidase from peach seeds – effect on enzyme activity, stability and antigenicity

Abstract

An anionic isoperoxidase (EC 1.11.1.7) purified from peach seeds (Prunus persica L. Batsch cv. Merry) was partially deglycosylated by glycopeptidase F (EC 3.2.2.18) treatment. A 40% deglycosylation resulted in an activity loss of 50% when assayed with o‐dianisidine. 60% with guaiacol and 78% with 2,2′‐azino‐bis(3‐ethyl)benzethiozoline‐6‐sulfonic acid (ABTS) as substrate. The indole‐3‐acetic acid oxidase activity loss was close to 55%. The partially deglycosylated isoperoxidase also showed a higher Km value for H2O2 and higher values for Arrhenius activation energy and enthalpy of activation. There was a decrease in enzyme stability at 4°C after deglycosylation. Native and partially deglycosylated isoperoxidase reacted equally well in an enzyme‐linked immunosorbent assay (ELISA) with rabbit polyclonal antibodies raised against the native enzyme. The carbohydrate moiety of this peach seed isoperoxidase appears to be important for enzyme activity and stability.