Cellulases without carbohydrate-binding modules in high consistency ethanol production process

Abstract

BackgroundEnzymes still comprise a major part of ethanol production costs from lignocellulose raw materials. Irreversible binding of enzymes to the residual substrate prevents their reuse and no efficient methods for recycling of enzymes have so far been presented. Cellulases without a carbohydrate-binding module (CBM) have been found to act efficiently at high substrate consistencies and to remain non-bound after the hydrolysis.ResultsHigh hydrolysis yields could be obtained with thermostable enzymes of Thermoascus aurantiacus containing only two main cellulases: cellobiohydrolase I (CBH I), Cel7A and endoglucanase II (EG II), Cel5A. The yields were decreased by only about 10% when using these cellulases without CBM. A major part of enzymes lacking CBM was non-bound during the most active stage of hydrolysis and in spite of this, produced high sugar yields. Complementation of the two cellulases lacking CBM with CBH II (CtCel6A) improved the hydrolysis. Cellulases without CBM were more sensitive during exposure to high ethanol concentration than the enzymes containing CBM. Enzymes lacking CBM could be efficiently reused leading to a sugar yield of 90% of that with fresh enzymes. The applicability of cellulases without CBM was confirmed under industrial ethanol production conditions at high (25% dry matter (DM)) consistency.ConclusionsThe results clearly show that cellulases without CBM can be successfully used in the hydrolysis of lignocellulose at high consistency, and that this approach could provide new means for better recyclability of enzymes. This paper provides new insight into the efficient action of CBM-lacking cellulases. The relationship of binding and action of cellulases without CBM at high DM consistency should, however, be studied in more detail.

Exo‐exo synergy between Cel6A and Cel7A from Hypocrea jecorina: Role of carbohydrate binding module and the endo‐lytic character of the enzymes

“…Looking, for example, at the data in Figure 1B and D, we find that at high substrate loads, the pair of core variants had a comparable or higher activity than the pair of wild types with CBMs. High activity of CBM-free enzymes in concentrated substrate suspensions, as observed here, has been reported earlier (Le Costaouec et al, 2013;Pakarinen et al, 2014;V arnai et al, 2013), and interpreted as a sign of an off-rate controlled reaction (Sørensen et al, 2015a,b). 2).…”

“…2). High activity of CBM-free enzymes in concentrated substrate suspensions, as observed here, has been reported earlier (Le Costaouec et al, 2013;Pakarinen et al, 2014;V arnai et al, 2013), and interpreted as a sign of an off-rate controlled reaction (Sørensen et al, 2015a,b). Thus, if enzymesubstrate dissociation is the rate limiting step, the weaker association of core-variants will speed up the overall reaction at high loads of substrate (increase V max ) (Sørensen et al, 2015a,b).…”

Distinct Roles for Carbohydrate-Binding Modules of Glycoside Hydrolase 10 (GH10) and GH11 Xylanases from Caldicellulosiruptor sp. Strain F32 in Thermostability and Catalytic Efficiency

“…This makes it an attractive candidate for use in the hydrolysis of lignocellulose with high consistency in industrial applications (56). The optimum temperatures of XynA and XynB both were at 75°C, and they also showed high thermal stability at this temperature.…”

Predominant Nonproductive Substrate Binding by Fungal Cellobiohydrolase I and Implications for Activity Improvement

“…Dual role of CBM in the catalytic activity of different cellulases towards insoluble substrates is well elucidated. The phenomenon of greater activity of cellulases without carbohydrate‐binding modules in the real high‐solids enzymatic hydrolysis of partially decrystallized lignocellulosic substrates is well recognized . Enzyme recovery from residual solids is simplified for cellulase without CBM compared with the CD‐CBM construct.…”