Analysis of the Surfaces of Wood Tissues and Pulp Fibers Using Carbohydrate-Binding Modules Specific for Crystalline Cellulose and Mannan

Carbohydrate binding modules (CBMs) are noncatalytic substrate binding domains of many enzymes involved in carbohydrate metabolism. Here we used fluorescent labeled recombinant CBMs specific for crystalline cellulose (CBM1 Hj Cel7A) and mannans (CBM27 Tm Man5 and CBM35 Cj Man5C) to analyze the complex surfaces of wood tissues and pulp fibers. The crystalline cellulose CBM1 Hj Cel7A was found as a reliable marker of both bacterially produced and plant G-layer cellulose, and labeling of spruce pulp fibers with CBM1 Hj Cel7A revealed a signal that increased with degree of fiber damage. The mannan-specific CBM27 Tm Man5 and CBM35 Cj Man5C CBMs were found to be more specific reagents than a monoclonal antibody specific for (1→4)-β-mannan/galacto-(1→4)-β-mannan for mapping carbohydrates on native substrates. We have developed a quantitative fluorometric method for analysis of crystalline cellulose accumulation on fiber surfaces and shown a quantitative difference in crystalline cellulose binding sites in differently processed pulp fibers. Our results indicated that CBMs provide useful, novel tools for monitoring changes in carbohydrate content of nonuniform substrate surfaces, for example, during wood or pulping processes and possibly fiber biosynthesis.