Effects of cellobiose on cellulose colonization by a mesophilic, cellulolytic Clostridium (strain C401)

Abstract

When cultured on a mixture of cellobiose and cellulose, Clostridium C401 did not initially attach to cellulose but remained in the liquid phase. After cellobiose exhaustion, bacterial cells grew in association with the insoluble cellulose. Carboxymethylcellulase (CMCase) production on Avicel cellulose was four- to fivefold greater than on cellobiose, and cellulose-grown cells adhered to filter paper with an initial adhesion rate about four- to fivefold greater than did cellobiose-grown cells. Using tritiated thymidine incorporation as a measure of growth, it appeared that transfer of strain C401 from cellobiose to cellulose required an adaptation phase. An extracellular cellulase complex was isolated by affinity chromatography. This enzyme system is a multicomponent aggregate (molecular mass above 5 MDa), and yielded two major polypeptide bands by SDS-PAGE having molecular masses of 130 and 70 kDa. Cellobiose strongly inhibited Avicelase activity and slightly inhibited p-nitrophenylcellobiose hydrolysis (pNPCbase), but had no effect on the CMCase activity of the cellulase complex. In addition, polyclonal antibodies, raised against the purified 130 kDa protein inhibited Avicelase activity, but not CMCase and pNPCbase activities.