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The synergistic action of accessory enzymes enhances the hydrolytic potential of a "cellulase mixture" but is highly substrate specific.

Hu J, Arantes V, Pribowo A, Saddler JN - Biotechnol Biofuels (2013)

Bottom Line: Currently, the amount of protein/enzyme required to achieve effective cellulose hydrolysis is still too high.The GH10 EX was more effective on steam pretreated agriculture residues and hardwood substrates whereas GH5 XG addition was more effective on softwood substrates.However, the extent of improved hydrolysis was highly substrate dependent.

View Article: PubMed Central - HTML - PubMed

Affiliation: Forestry Products Biotechnology/Bioenergy Group, Wood Science Department, University of British Columbia, 2424 Main Mall, Vancouver, BC V6T 1Z4, Canada. jack.saddler@ubc.ca.

ABSTRACT

Background: Currently, the amount of protein/enzyme required to achieve effective cellulose hydrolysis is still too high. One way to reduce the amount of protein/enzyme required is to formulate a more efficient enzyme cocktail by adding so-called accessory enzymes such as xylanase, lytic polysaccharide monooxygenase (AA9, formerly known as GH61), etc., to the cellulase mixture. Previous work has shown the strong synergism that can occur between cellulase and xylanase mixtures during the hydrolysis of steam pretreated corn stover, requiring lower protein loading to achieve effective hydrolysis. However, relatively high loadings of xylanases were required. When family 10 and 11 endo-xylanases and family 5 xyloglucanase were supplemented to a commercial cellulase mixture varying degrees of improved hydrolysis over a range of pretreated, lignocellulosic substrates were observed.

Results: The potential synergistic interactions between cellulase monocomponents and hemicellulases from family 10 and 11 endo-xylanases (GH10 EX and GH11 EX) and family 5 xyloglucanase (GH5 XG), during hydrolysis of various steam pretreated lignocellulosic substrates, were assessed. It was apparent that the hydrolytic activity of cellulase monocomponents was enhanced by the addition of accessory enzymes although the "boosting" effect was highly substrate specific. The GH10 EX and GH5 XG both exhibited broad substrate specificity and showed strong synergistic interaction with the cellulases when added individually. The GH10 EX was more effective on steam pretreated agriculture residues and hardwood substrates whereas GH5 XG addition was more effective on softwood substrates. The synergistic interaction between GH10 EX and GH5 XG when added together further enhanced the hydrolytic activity of the cellulase enzymes over a range of pretreated lignocellulosic substrates. GH10 EX addition could also stimulate further cellulose hydrolysis when added to the hydrolysis reactions when the rate of hydrolysis had levelled off.

Conclusions: Endo-xylanases and xyloglucanases interacted synergistically with cellulases to improve the hydrolysis of a range of pretreated lignocellulosic substrates. However, the extent of improved hydrolysis was highly substrate dependent. It appears that those accessory enzymes, such as GH10 EX and GH5 XG, with broader substrate specificities promoted the greatest improvements in the hydrolytic performance of the cellulase mixture on all of the pretreated biomass substrates.

No MeSH data available.


Related in: MedlinePlus

SDS-PAGE of purified enzymes: GH11 EX (lane 1), GH10 EX (lane 2), GH5 XG (lane 3), Cel7A (lane 4), Cel6A (lane 5), Cel7B (lane 6), Cel5A (lane 7), GH3 BG (lane 8) and marker (lane M). Proteins were identified by LC-MS/MS. Proteins are named according to their glycoside hydrolase family.
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Figure 1: SDS-PAGE of purified enzymes: GH11 EX (lane 1), GH10 EX (lane 2), GH5 XG (lane 3), Cel7A (lane 4), Cel6A (lane 5), Cel7B (lane 6), Cel5A (lane 7), GH3 BG (lane 8) and marker (lane M). Proteins were identified by LC-MS/MS. Proteins are named according to their glycoside hydrolase family.

Mentions: The major protein within the Multifect Xylanase was found to be GH11 EX, as shown by tandem mass spectrometry, with this protein constituting more than 80% of the total protein present in the mixture. As expected, it had a relatively low molecular weight of about 20 kDa (Figure 1). The major cellulase monocomponents within Celluclast 1.5L, on a protein weight basis, were T. reesei Cel7A, Cel6A, Cel7B, and Cel5A (Figure 1), which comprise about 56%, 12%, 5%, and 6% of the total protein respectively. These values were similar to the proportion of protein concentrations reported earlier by other workers [27,28].


The synergistic action of accessory enzymes enhances the hydrolytic potential of a "cellulase mixture" but is highly substrate specific.

Hu J, Arantes V, Pribowo A, Saddler JN - Biotechnol Biofuels (2013)

SDS-PAGE of purified enzymes: GH11 EX (lane 1), GH10 EX (lane 2), GH5 XG (lane 3), Cel7A (lane 4), Cel6A (lane 5), Cel7B (lane 6), Cel5A (lane 7), GH3 BG (lane 8) and marker (lane M). Proteins were identified by LC-MS/MS. Proteins are named according to their glycoside hydrolase family.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3750293&req=5

Figure 1: SDS-PAGE of purified enzymes: GH11 EX (lane 1), GH10 EX (lane 2), GH5 XG (lane 3), Cel7A (lane 4), Cel6A (lane 5), Cel7B (lane 6), Cel5A (lane 7), GH3 BG (lane 8) and marker (lane M). Proteins were identified by LC-MS/MS. Proteins are named according to their glycoside hydrolase family.
Mentions: The major protein within the Multifect Xylanase was found to be GH11 EX, as shown by tandem mass spectrometry, with this protein constituting more than 80% of the total protein present in the mixture. As expected, it had a relatively low molecular weight of about 20 kDa (Figure 1). The major cellulase monocomponents within Celluclast 1.5L, on a protein weight basis, were T. reesei Cel7A, Cel6A, Cel7B, and Cel5A (Figure 1), which comprise about 56%, 12%, 5%, and 6% of the total protein respectively. These values were similar to the proportion of protein concentrations reported earlier by other workers [27,28].

Bottom Line: Currently, the amount of protein/enzyme required to achieve effective cellulose hydrolysis is still too high.The GH10 EX was more effective on steam pretreated agriculture residues and hardwood substrates whereas GH5 XG addition was more effective on softwood substrates.However, the extent of improved hydrolysis was highly substrate dependent.

View Article: PubMed Central - HTML - PubMed

Affiliation: Forestry Products Biotechnology/Bioenergy Group, Wood Science Department, University of British Columbia, 2424 Main Mall, Vancouver, BC V6T 1Z4, Canada. jack.saddler@ubc.ca.

ABSTRACT

Background: Currently, the amount of protein/enzyme required to achieve effective cellulose hydrolysis is still too high. One way to reduce the amount of protein/enzyme required is to formulate a more efficient enzyme cocktail by adding so-called accessory enzymes such as xylanase, lytic polysaccharide monooxygenase (AA9, formerly known as GH61), etc., to the cellulase mixture. Previous work has shown the strong synergism that can occur between cellulase and xylanase mixtures during the hydrolysis of steam pretreated corn stover, requiring lower protein loading to achieve effective hydrolysis. However, relatively high loadings of xylanases were required. When family 10 and 11 endo-xylanases and family 5 xyloglucanase were supplemented to a commercial cellulase mixture varying degrees of improved hydrolysis over a range of pretreated, lignocellulosic substrates were observed.

Results: The potential synergistic interactions between cellulase monocomponents and hemicellulases from family 10 and 11 endo-xylanases (GH10 EX and GH11 EX) and family 5 xyloglucanase (GH5 XG), during hydrolysis of various steam pretreated lignocellulosic substrates, were assessed. It was apparent that the hydrolytic activity of cellulase monocomponents was enhanced by the addition of accessory enzymes although the "boosting" effect was highly substrate specific. The GH10 EX and GH5 XG both exhibited broad substrate specificity and showed strong synergistic interaction with the cellulases when added individually. The GH10 EX was more effective on steam pretreated agriculture residues and hardwood substrates whereas GH5 XG addition was more effective on softwood substrates. The synergistic interaction between GH10 EX and GH5 XG when added together further enhanced the hydrolytic activity of the cellulase enzymes over a range of pretreated lignocellulosic substrates. GH10 EX addition could also stimulate further cellulose hydrolysis when added to the hydrolysis reactions when the rate of hydrolysis had levelled off.

Conclusions: Endo-xylanases and xyloglucanases interacted synergistically with cellulases to improve the hydrolysis of a range of pretreated lignocellulosic substrates. However, the extent of improved hydrolysis was highly substrate dependent. It appears that those accessory enzymes, such as GH10 EX and GH5 XG, with broader substrate specificities promoted the greatest improvements in the hydrolytic performance of the cellulase mixture on all of the pretreated biomass substrates.

No MeSH data available.


Related in: MedlinePlus