SEQUENTIAL PRECIPITATION WITH REVERSIBLY SOLUBLE INSOLUBLE POLYMERS AS A BIOSEPARATION STRATEGY - PURIFICATION OF BETA-GLUCOSIDASE FROM TRICHODERMA-LONGIBRACHIATUM

Precipitation of proteins/enzymes with reversibly soluble-insoluble po lymers is a simple approach which can be easily scaled up. it is shown here that; considerable purification may be achieved if more than one polymer (chosen judiciously) is used in sequence for purification. Th e principle is illustrated with purification of beta-glucosidase from a commercial preparation of Trichoderma longibrachiatum. Precipitation with chitosan removed cellulase activity from the preparation with 4- fold purification and 99% recovery of beta-glucosidase activity in the supernatant. The treatment of the supernatant with Eudragit S-100 pre cipitated 98% of the enzyme activity. The two different elution proced ures gave 82% recovery (with 14-fold purification) and 86% recovery (w ith 12-fold purification) of the enzyme activity from the precipitate. A low molecular weight protein still present in these preparations co uld be removed by gel filtration. This finally yielded an electrophore tically homogeneous enzyme with 74% recovery and 29-fold purification. (C) 1996 Academic Press, Inc.