DISSOCIATION OF THE CELLULOSOME OF CLOSTRIDIUM-THERMOCELLUM UNDER NONDENATURING CONDITIONS

The cellulosome from the anaerobic, thermophilic, cellulolytic bacteri um, Clostridium thermocellum, was dissociated under nondenaturing cond itions by incubation at 60 degrees C in the combined presence of EDTA and cellulose. This approach differs from previous strategies in that detergents were not necessary for dissociation of the subunits. Select ed enzymatic subunits could be isolated with relative ease. One of the se is the major cellulosomal cellobiohydrolase, CelS (previously desig nated subunit S8 or S-s). Both the intact cellulosome complex and the combined dissociated subunits exhibited similar levels of activities o n soluble and acid-swollen cellulosic substrates. In contrast, the mix ture of the free enzymatic subunits was markedly deficient in its degr adation of crystalline cellulose compared with that of the intact cell ulosome. The results support the critical role of divalent cations. e. g. calcium, in the stability of the cellulosome complex. In addition, the results reinforce previous indications that the cellulosome underg oes conformational changes upon binding to cellulose. Copyright (C) 19 96 Elsevier Science B.V.