Covalant coupling of β-fructofuranosidase on microbial cells

Abstract

β-Fructofuranosidase, an hydrolyzing β-1,6-fructosidic linkage, was immobilized covalently on the oxidized microbial cell wall of a Penicillium species (PS-8), which is totally different from the conventional whole cell immobilization. The microbial cells of the PS-8 strain, which has a fluffy characteristic during cultivation, was treated with sodium metaperiodate to form active aldehyde groups on them. The periodate treated cells were reacted with the β-fructofuranosidase. This mixture of cell and the enzyme was again treated with 0.5% glutaraldehyde for 1 hr and finally spinned with a syringe (diameter 1 mm) to form pellets, and dried at air. By determining the effects of periodate concentration on the aldehyde formation of microbial cells, the optimal treating concentration of periodate was 1.2 grams per gram of the dry weight and the formation of the aldehyde was 35% as the % dialdehyde units. The reaction temperature did not give any effect on the formation of aldehyde of the microbial cells at this conditions. The concentrations and treating time of glutaraldehyde on the immobilization of the β-fructofuranosidase on dialdehyde microbial cells were very important factors in the enzyme immobilization. The highest enzyme activity was observed to be immobilized for 30 minute at 0.5% (w/v) concentration of glutaraldehyde. The final pellet products (1×2mm) of the immobilized β-fructofuranosidase showed good mechanical strength and was not powderized in shape when they were soaked even for long periods of time. The flow rate was also satisfied when the pellets were packed in the column. Overall recovery of β-fructofuranosidase activity of the final immobilized product showed 26% of the total activity applied. Kinetic parameters of the soluble and the immobilized β-fructofuranosidase were compared. The results showed that the optimal pH of the immobilized β-fructofuranosidase was PH5, which was same as that of the soluble enzyme. The optimal temperature of ...