Adsorption and synergism of cellobiohydrolase I and II of Trichoderma reesei during hydrolysis of microcrystalline cellulose

• Published in: Biotechnology and bioengineering 1994-11, Vol.44 (9), p.1064-1073
• Main Authors: Medve, J. (University of Lund, Lund, Sweden.), Stahlberg, J, Tjerneld, F
• Format: Article
• Language: English
• Subjects: 09 BIOMASS FUELS; ADSORCION; ADSORPTION; Biological and medical sciences; Biotechnology; CARBOHYDRATES; CELLOBIOSE; CELLULASE; CELLULOSE; cellulose hydrolysis; CELOBIOSA; CELULASA; CELULOSA; CHEMICAL REACTIONS; CHROMATOGRAPHY; DECOMPOSITION; ENZYMATIC HYDROLYSIS; ENZYME ACTIVITY; Enzyme engineering; ENZYMES; EUMYCOTA; FPLC; Fundamental and applied biological sciences. Psychology; FUNGI; HIDROLASAS; HIDROLISIS; HYDROLASE; HYDROLASES; HYDROLYSE; HYDROLYSIS; LIQUID COLUMN CHROMATOGRAPHY; LYSIS; Methods. Procedures. Technologies; Miscellaneous; ORGANIC COMPOUNDS; PLANTS; POLYSACCHARIDES; PROTEINS; SACCHARIDES; SEPARATION PROCESSES; SINERGISMO; SOLVOLYSIS; SORPTION; SYNERGIE; SYNERGISM; TRICHODERMA 090900 > Biomass Fuels-- Processing-- (1990-); TRICHODERMA LONGIBRACHIATUM; Trichoderma reesei; TRICHODERMA VIRIDE
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.260440907

Hydrolysis of microcrystalline cellulose (Avicel) by cellobiohydrolase I and II (CBH I and II) from Trichoderma reesei has been studied. Adsorption and synergism of the enzymes were investigated. Experiments were performed at different temperatures and enzyme/substrate ratios using CBH I and CBH II alone and in reconstituted equimolar mixtures. Fast protein liquid chromatography (FPLC) analysis was found to be an accurate and reproducible method to follow the enzyme adsorption. A linear correlation was found between the conversion and the amount of adsorbed enzyme when Avicel was hydrolyzed by increasing amounts of CBH I and/or CBH II. CBH I had lower specific activity compared to CBH II although, over a wide concentration range, more CBH I was adsorbed than CBH II. Synergism between the cellobiohydrolases during hydrolysis of the amorphous fraction of Avicel showed a maximum as a function of total enzyme concentration. Synergism measured as a function of bound enzyme showed a continuous increase, which indicates that by decreasing the distance between the two enzymes the synergism is enhanced. The adsorption process for both enzymes was slow. Depending on the enzyme/substrate ratio it took 30-90 min to reach 95% of the equilibrium binding. The amount of bound enzyme decreased with increasing temperature. The two enzymes compete for the adsorption sites but also bind to specific sites. Stronger competition for adsorption sites was shown by CBH I