Immobilization of a soluble chemically thermostabilized enzyme

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Abstract
Cellobiase [from Aspergillus niger] was coupled to a dialdehyde dextran by reductive alkylation in the presence of sodium cyanoborohydride. The resulting conjugate, obtained without loss of enzymic activity, presents properties of thermoresistance largely superior to those of native enzyme; the rate of inactivation is reduced compared to that of native enzyme and its optimal temperature of activity is 70.degree.-75.degree. C instead of 65.degree. C. The conjugate presents increased longevity when subjected to experiments of operational stability; its hydrolytic activity is maintained at 60.degree. C in a 10% (wt/vol) cellobiose solution for more than 100 h whereas the native enzyme is inactivated after 45 h. The cellobiase-dextran conjugate was immobilized by covalent coupling on aminated silica by reductive alkylation in the presence of NaBH3CN. The characteristics of thermoresistance of this stabilized and immobilized conjugate were studied and compared to those of a preparation of native cellobiase immobilized on a silica support activated with glutaraldehyde. Analysis of the thermoresistance of these 2 cellobiase preparations clearly shows that immobilization maintained and even enhanced their properties. In particular, the operational stability, measured at 68.degree. C on 10% (wt/vol) cellobiose shows an increased longevity of the stabilized and immobilized enzyme for 120 h compared to 60 h for the native immobilized enzyme. Two successive incubations of these cellobiase derivatives show that it is possible to obtain 2.5 times more glucose with the stabilized-immobilized enzyme than with the immobilized preparation. The procedure described above enables us to prepare a thermostabilized immobilized cellobiase.

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