COMPARATIVE-STUDY OF EXTERNAL AND INTERNAL BETA-GLUCOSIDASES AND GLUCOAMYLASE OF ARXULA-ADENINIVORANS

Properties of external and internal beta-glucosidases and glucoamylase of Arxula adeninivorans were compared. For the purification of enzymes, the C-catabolite derepression mutant SBUG 724/41 was used. The following K(m)-values for beta-glucosidases with cellobiose as substrate were measured: 14.3 mM (internal beta-glucosidase I), 4.1 mM (external beta-glucosidase I), 5.0 mM (internal beta-glucosidase II), 3.0 mM (external beta-glucosidase II). The K(m)-values of the glucoamylase for maltose were 4.6 mM (internal enzyme), 11.1 mM (external enzyme), and for starch 0.32 g/l and 1.2 g/l, respectively. Both external beta-glucosidases showed a relatively sharp pH-optimum in a range from 4.5 to 5.5 while that of internal enzymes was between 5.5 and 6.0. No differences were obtained between pH-optima (4.5-5.5) of internal and external glucoamylase. The temperature optimum values of internal beta-glucosidase I (40-degrees-C) and of internal beta-glucosidase II (50-degrees-C) were remarkably low compared with those of the external beta-glucosidases (60-63-degrees-C) and both glucoamylases (60-degrees-C). Molecular mass of 570 kD for the external and of 290 kD for the internal beta-glucosidase I, on the one hand, and of 525 kD and 180 kD for the internal beta-glucosidase II on the other hand were determined. The external glucoamylase consists of two subunits with a molecular mass of 110 kD. After deglycosylation a molecular mass of 76 kD was measured, indicating a carbohydrate content of 30%, whereas for the internal enzyme, a molecular mass of 95 kD was estimated.