Role of the Carbohydrate Moiety in Phospholipase B from Torulaspora delbrueckii

Abstract

Water-soluble phospholipase B containing approximately 50% carbohydrate, purified to homogeneity from yeast cell washings of Torulaspora delbrueckii, was treated with endoglycosidase H (Endo-H), and about 75% of the carbohydrate associated with this enzyme was removed. The native and carbohydrate-depleted enzymes were compared. The phospholipase B activity was higher in the carbohydrate-depleted enzyme than in the native form. The enzymatic activation by the removal of carbohydrate was due to the increased affinity of the enzyme for the substrate. The carbohydrate-depleted phospholipase B was less stable upon incubation at 50°C and more susceptible to proteolysis by V8 protease. These results suggested that the carbohydrate of phospholipase B in yeast cells stabilizes the enzyme conformation and protects the enzyme from proteolysis.