The enhancement of the cellulolytic activity of cellobiohydrolase I and endoglucanase by the addition of cellulose binding domains derived from Trichoderma reesei

TLDR: Hydlysis of filter paper by intact cellobiohydrolase I in the presence of additional CBDs was found to have a synergistic effect, leading to an increase of the sugar production of up to 30%, less pronounced using microcrystalline cellulose, where an increase up to 16% was observed.

About: This article is published in Enzyme and Microbial Technology.The article was published on 2003-01-02 and is currently open access. It has received 45 citations till now. The article focuses on the topics: Cellulose binding & Trichoderma reesei.

Figures

Fig. 2. Data and fit for the hydrolysis of filter paper by endoglucanase and cellobiohydrolase I (in the presence and absence of additional isolated CBDs) using the ratio 25.

Fig. 5. Overall increase of sugar production by endoglucanase and cellobiohydrolase, comparing different substrate/enzyme ratios and type of substrate, when CBDs were added to the enzyme solution.

Fig. 1. Mass spectra of the peptide mixture.

Fig. 4. Differences in the two rates ( , k1-CBD/k1 and, , k2-CBD/k2) obtained with and without the presence of CBDs for the systems of: (a) endoglucanase, Sigmacel; (b) endoglucanase, filter paper; (c) cellobiohydrolase I, Sigmacel and (d) cellobiohydrolase I, filter paper.

Fig. 3. Data and fit for the hydrolysis of Sigmacel by endoglucanase and cellobiohydrolase I (in the presence and absence of additional isolated CBDs) using the ratio 200.

Frequently asked questions

Q1. What are the contributions mentioned in the paper "The enhancement of the cellulolytic activity of cellobiohydrolase i and endoglucanase by the addition of cellulose binding domains derived from trichoderma reesei" ?

The effect of isolated cellulose binding domains ( CBDs ) on the hydrolysis of filter paper and microcrystalline cellulose by both cellobiohydrolase I and endoglucanase, was studied. 

Q2. What are the future works in "The enhancement of the cellulolytic activity of cellobiohydrolase i and endoglucanase by the addition of cellulose binding domains derived from trichoderma reesei" ?

However, the exact mechanism by which the synergistic effect is manifest is not yet clear and further work related to structural changes in celluloses induced by CBDs, using techniques such as scanning electron microscopy, X-ray diffraction and IR spectroscopy, could prove fruitful. 

Q3. What is the role of cellulases in the biotechnology industry?

Cellulases are enzymes able to hydrolyse cellulose and the hydrolysis of crystalline cellulose can be performed by a fungal cellulase system, where three main enzymes play a pivotal role. 

Q4. What is the effect of CBDs on the enzyme activity?

The enhancement of enzyme activity by the presence of additional isolated CBDs is also dependent on the hydrolysis mechanism of the enzyme. 

Q5. How much protein is in the core binding domain of fungal cellulases?

The molecular weight of the core binding domain of fungal cellulases is usually close to 5 kDa, however, if it remains heavily glycosylated a molecular weight of 10 kDa can be expected [16]. 

Q6. How much sugar production was found to increase when using a combination of CBD and enzyme?

Hydrolysis of filter paper by native cellobiohydrolase The authorin the presence of CBDs was found to lead to an increase of the sugar production of up to 30%. 

Q7. What are the main applications of hemicellulases in biotechnology?

Cellulases and hemicellulases have a wide range of application in biotechnology, such as in the food industry, pulp and paper industry, textile and laundry [1]. 

Q8. What are the main enzymes that can hydrolyze cellulose?

These are (i) cellobiohydrolases (CBH), which act as an exoglucanase releasing cellobiose as the main product, (ii) endoglucanases (EG) that act randomly along cellulose chains producing new sites for the cellobiohydrolases, and (iii) -glucosidase, which hydrolyses cellobiose, a potential inhibitor of CBH. 

Q9. What was the main synergistic effect of the hydrolysis?

The hydrolysis was found to be adequately modelled by a two stage process and the major synergism was found in the second phase, after “easy” substrate has been hydrolysed by enzyme with little or no assistance by the CBD. 

Q10. How long did the incubation time be used?

In their work an incubation time of 48 h was used and as in some cases the synergistic effect is more apparent in the k2 rate controlled (longer timescale) process and therefore, care must be taken to ensure that no effect has been overlooked (or not detected). 

Q11. What is the main mechanism of the exo–endo synergism?

The so-called exo–endo synergism has been described in the literature [2–5] and is normally explained by the fact that EG can provide free end chains on the cellulose surface for the CBH to act on. 

Q12. What is the effect of CBDs on the structure of cellulose?

In this study, the authors have shown that unlinked, isolated CBDs from Trichoderma reesei (obtained by proteolysis), when they are added to the solution produce an enhancement in the hydrolysis of cellulose (in both microcrystalline and filterpaper) by intact CBH The authorand EG. 

Q13. What is the effect of the control experiment on the amount of sugar released?

should not in fact display any hydrolytic activity, but the control experiment also helps to rule out any contaminants, that could exhibit activity. 

Q14. What is the effect of the combination of enzyme and CBD?

These data strongly indicate that the enhancement of sugar production can be attributed to the combination of enzyme and CBD and not to any residual activity in the CBD solution. 

Q15. What is the role of the CBD in the cellulose hydrolysis?

For instance, Din et al. [12] observed that the isolated CBD of endoglucanase A (CenA) from Cellulomonas fimi disrupted the structure of cellulose fibres and released small particles.