Oxidation of lactose to lactobionic acid by a Microdochium nivale carbohydrate oxidase: kinetics and operational stability.
Oxidation of lactose to lactobionic acid by a Microdochium nivale carbohydrate oxidase was studied. The K(m)-value for lactose, obtained by a traditional enzymatic assay, was 0.066 mM at pH 6.4 and 38 degrees C. The effect of oxygen on the enzymatic rate of reaction as well as the operational stability of the enzyme was studied by performing reactions at constant pH and temperature in a stirred tank reactor. Catalase was included in all reactions to avoid inhibition and deactivation of the oxidase by hydrogen peroxide. At pH 6.4 and 38 degrees C, K(m) for oxygen was 0.97 mM, while the catalytical rate constant, k(cat), was 94 s(-1). Furthermore, we found that the operational stability of the oxidase was dependent on the type of base used for neutralization of the acid produced. Thus, when 2 M NaOH was used for neutralization of a reaction medium containing 50 mM phosphate buffer, significant deactivation of the oxidase was observed. Also, we found that the oxidase was protected against deactivation by base at high lactose concentrations. A simple model is proposed to explain the obtained results.
Nordkvist
Mikkel
M
Center for Microbial Biotechnology, BioCentrum-DTU, Technical University of Denmark, DK-2800 Kgs. Lyngby, Denmark.
Nielsen
Per Munk
PM
Villadsen
John
J
D001203:Ascomycota / Q000201:enzymology* / Q000378:metabolism; D019149:Bioreactors / Q000382:microbiology; D001711:Biotransformation; D002237:Carbohydrate Dehydrogenases / Q000378:metabolism*; D004187:Disaccharides / Q000378:metabolism*; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007785:Lactose / Q000378:metabolism*; D010084:Oxidation-Reduction; D013696:Temperature
D004187:Disaccharides; C005608:lactobionic acid; D002237:Carbohydrate Dehydrogenases; D007785:Lactose