Practical screening of purified cellobiohydrolases and endoglucanases with α-cellulose and specification of hydrodynamics
Author(s)
Type
Journal Article
Abstract
Background: It is important to generate biofuels and society must be weaned from its dependency on fossil fuels.
In order to produce biofuels, lignocellulose is pretreated and the resulting cellulose is hydrolyzed by cellulases such
as cellobiohydrolases (CBH) and endoglucanases (EG). Until now, the biofuel industry has usually applied
impractical celluloses to screen for cellulases capable of degrading naturally occurring, insoluble cellulose. This
study investigates how these cellulases adsorb and hydrolyze insoluble a-cellulose − considered to be a more
practical substrate which mimics the alkaline-pretreated biomass used in biorefineries. Moreover, this study
investigates how hydrodynamics affects cellulase adsorption and activity onto a-cellulose.
Results: First, the cellulases CBH I, CBH II, EG I and EG II were purified from Trichoderma reesei and CBH I and EG I
were utilized in order to study and model the adsorption isotherms (Langmuir) and kinetics (pseudo-first-order).
Second, the adsorption kinetics and cellulase activities were studied under different hydrodynamic conditions,
including liquid mixing and particle suspension. Third, in order to compare a-cellulose with three typically used
celluloses, the exact cellulase activities towards all four substrates were measured.
It was found that, using a-cellulose, the adsorption models fitted to the experimental data and yielded parameters
comparable to those for filter paper. Moreover, it was determined that higher shaking frequencies clearly improved
the adsorption of cellulases onto a-cellulose and thus bolstered their activity. Complete suspension of a-cellulose
particles was the optimal operating condition in order to ensure efficient cellulase adsorption and activity. Finally,
all four purified cellulases displayed comparable activities only on insoluble a-cellulose.
Conclusions: a-Cellulose is an excellent substrate to screen for CBHs and EGs. This current investigation shows in
detail, for the first time, the adsorption of purified cellulases onto a-cellulose, the effect of hydrodynamics on
cellulase adsorption and the correlation between the adsorption and the activity of cellulases at different
hydrodynamic conditions. Complete suspension of the substrate has to be ensured in order to optimize the
cellulase attack. In the future, screenings should be conducted with a-cellulose so that proper cellulases are
selected to best hydrolyze the real alkaline-pretreated biomass used in biorefineries.
In order to produce biofuels, lignocellulose is pretreated and the resulting cellulose is hydrolyzed by cellulases such
as cellobiohydrolases (CBH) and endoglucanases (EG). Until now, the biofuel industry has usually applied
impractical celluloses to screen for cellulases capable of degrading naturally occurring, insoluble cellulose. This
study investigates how these cellulases adsorb and hydrolyze insoluble a-cellulose − considered to be a more
practical substrate which mimics the alkaline-pretreated biomass used in biorefineries. Moreover, this study
investigates how hydrodynamics affects cellulase adsorption and activity onto a-cellulose.
Results: First, the cellulases CBH I, CBH II, EG I and EG II were purified from Trichoderma reesei and CBH I and EG I
were utilized in order to study and model the adsorption isotherms (Langmuir) and kinetics (pseudo-first-order).
Second, the adsorption kinetics and cellulase activities were studied under different hydrodynamic conditions,
including liquid mixing and particle suspension. Third, in order to compare a-cellulose with three typically used
celluloses, the exact cellulase activities towards all four substrates were measured.
It was found that, using a-cellulose, the adsorption models fitted to the experimental data and yielded parameters
comparable to those for filter paper. Moreover, it was determined that higher shaking frequencies clearly improved
the adsorption of cellulases onto a-cellulose and thus bolstered their activity. Complete suspension of a-cellulose
particles was the optimal operating condition in order to ensure efficient cellulase adsorption and activity. Finally,
all four purified cellulases displayed comparable activities only on insoluble a-cellulose.
Conclusions: a-Cellulose is an excellent substrate to screen for CBHs and EGs. This current investigation shows in
detail, for the first time, the adsorption of purified cellulases onto a-cellulose, the effect of hydrodynamics on
cellulase adsorption and the correlation between the adsorption and the activity of cellulases at different
hydrodynamic conditions. Complete suspension of the substrate has to be ensured in order to optimize the
cellulase attack. In the future, screenings should be conducted with a-cellulose so that proper cellulases are
selected to best hydrolyze the real alkaline-pretreated biomass used in biorefineries.
Date Issued
2010-08-18
Date Acceptance
2010-08-18
Citation
Biotechnology for Biofuels, 2010, 3
ISSN
1754-6834
Publisher
BioMed Central
Journal / Book Title
Biotechnology for Biofuels
Volume
3
Copyright Statement
© 2010 Jäger et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons
Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in
any medium, provided the original work is properly cited.
Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in
any medium, provided the original work is properly cited.
License URL
Subjects
Science & Technology
Life Sciences & Biomedicine
Technology
Biotechnology & Applied Microbiology
Energy & Fuels
BIOTECHNOLOGY & APPLIED MICROBIOLOGY
TRICHODERMA-REESEI CELLULASES
PROTEIN LIQUID-CHROMATOGRAPHY
ENZYMATIC-HYDROLYSIS RATE
STEAM-EXPLODED WOOD
FILTER-PAPER ASSAY
MICROCRYSTALLINE CELLULOSE
ETHANOL-PRODUCTION
CRYSTALLINE CELLULOSE
DIFFERENTIAL-EQUATIONS
ADSORPTION-KINETICS
0904 Chemical Engineering
1003 Industrial Biotechnology
Notes
Cited By :27 Export Date: 6 May 2015
Publication Status
Published
Article Number
18