A newly described cellulosomal cellobiohydrolase, CelO, from Clostridium thermocellum: Investigation of the exo-mode of hydrolysis, and binding capacity to crystalline cellulose

Abstract

The sequence of the ceIO gene from Clostridium thermocellum F7 was determined. The gene product, cellulase CeIO (Ct-CeI5F), had a modular structure consisting of a carbohydrate-binding module of the CBM3 family and a catalytic domain of the glycosyl hydrolase family 5. The presence of the dockerin module indicated that the enzyme was a component of the cellulosome complex. The thermostable recombinant gene product was active on cellodextrins, barley β-glucan, carboxymethylcellulose and insoluble cellulose. Cellobiose was the only product released from amorphic and crystalline cellulose, cellotetraose and higher cello-oligosaccharides, identifying CeIO as a cellobiohydrolase. The cleavage pattern of p-nitrophenyl β-D-cellotetraoside, blockage of the hydrolysis of NaBH4-reduced cellopentaose and the reduction in substrate viscosity suggested activity from the reducing end in a processive mode after making random cuts. Binding to insoluble, i.e. amorphous, and crystalline cellulose was mediated by the carbohydrate-binding module CBM3b, with a preference for the crystalline substrate.