ENHANCEMENT OF THE ENDO-BETA-1,4-GLUCANASE ACTIVITY OF AN EXOCELLOBIOHYDROLASE BY DELETION OF A SURFACE LOOP

In the commonly accepted mechanism for enzymatic hydrolysis of cellulo se, endo-beta-1,4-glucanases randomly cleave glucosidic bonds within g lucan polymers, providing sites for attack by exo-cellobiohydrolases ( EC 3.2.1.91), It has been proposed that hydrolysis by Trichoderma rees ei cellobiohydrolase II is restricted to the ends of cellulose polymer s because two surface loops cover its active site to form a tunnel, In a closely related endoglucanase, E2 from Thermomonospora fusca, acces s to the substrate appears to be relatively unhindered because the car boxyl-proximal loop is shortened, and the amino-proximal loop is displ aced, The hypothesis was examined by deletion of a region in Cellulomo nas fimi cellobiohydrolase A corresponding to part of the carboxyl-pro ximal loop of T. reesei cellobiohydrolase II, The mutation enhanced th e endoglucanase activity of the enzyme on soluble O-(carboxymethyl)cel lulose and altered its activities on 2',4'-dinitrophenyl-beta-D-cellob ioside, insoluble cellulose, and cellotetraose,