Reducing end-specific fluorescence labeled celluloses for cellulase mode of action

Cellobiohydrolases (CBH-s) attack primarily from the cellulose chain ends. It is known that CBH-s can differ in their chain end preference. Here the cellulose reducing end-specific fluorescence labeling was studied with the aim to find the most suitable labeled cellulose for easy determination of CBH-s chain end preference. Anthranilic acid (AA) was used as fluorescence label. Bacterial cellulose, amorphous Avicel and filter paper were subjected to the labeling. Suitability of modified bicincoic acid method for determination of reducing groups on cellulose was also confirmed. AA labeled celluloses were hydrolysed with family 7 and 6 CBH-s from Trichoderma reesei and Phanerochaete chrysosporium. Hydrolysis data were plotted as released label (%) versus total degradation (%) (progress curves). Most characteristic progress curves were obtained with AA labeled regenerated amorphous Avicel where the family 6 CBH-s produced clearly upward curved progress curves confirming their preference for non-reducing ends.