Effects of carbohydrate depletion on the structure, stability and activity of glucose oxidase from Aspergillus niger.
Glucose oxidase from Aspergillus niger was purified to homogeneity by hydrophobic interaction and ion-exchange chromatography. Approx. 95% of the carbohydrate moiety was cleaved from the protein by incubation of glucose oxidase with endoglycosidase H and alpha-mannosidase. Cleavage of the carbohydrate moiety effected a 24-30% decrease in the molecular weight and a reduction in the number of isoforms of glucose oxidase. No significant changes were observed in the circular dichroism spectra of the deglycosylated enzyme. Other properties, such as thermal stability, pH and temperature optima of glucose oxidase activity and substrate specificity were not affected. However, removal of the carbohydrate moiety marginally affected the kinetics of glucose oxidation and stability at low pH. From these results it appears that the carbohydrate chain of glucose oxidase does not contribute significantly to the structure, stability and activity of glucose oxidase.
Department of Enzyme Technology, GBF Gesellscaft für Biotechnologische Forschung, Braunschweig, F.R.G.
H M
HM
Kalisz
H J
HJ
Hecht
D
D
Schomburg
R D
RD
Schmid
D000595:Amino Acid Sequence; D001234:Aspergillus niger / Q000201:enzymology*; D002241:Carbohydrates / Q000032:analysis; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005949:Glucose Oxidase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D006023:Glycoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D006031:Glycosylation; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D011487:Protein Conformation; D013816:Thermodynamics
D002241:Carbohydrates; D006023:Glycoproteins; D005949:Glucose Oxidase