Construction and characterization of bifunctional cellulases: Caldicellulosiruptor-sourced endoglucanase, CBM, and exoglucanase for efficient degradation of lignocellulose

Li, Xiangqian; Xia, Jilin; Zhu, Xiaoyan; Bilal, Muhammad; Tan, Zhiyong; Shi, Hao

doi:10.1016/j.bej.2019.107363

Cited by 33 publications

(12 citation statements)

References 34 publications

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“…However, the enzyme activity increased rapidly after 48 h heat treatment, especially at 50°C. Similar results were obtained on fused enzymes from C. saccharolyticus [33]. Collectively, our results suggested that the enzyme activity may be stimulated at a certain high temperature and was stable at 50°C, which is beneficial for enzyme stability after heat treatment and reducing enzyme storage costs.…”

Section: Characteristics Of Recombinant Cela1805supporting

confidence: 87%

Characterization of a GH8 β-1,4-Glucanase from Bacillus subtilis B111 and Its Saccharification Potential for Agricultural Straws

Huang

Zhao

et al. 2021

J. Microbiol. Biotechnol.

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Herein, we cloned and expressed an endo-β-1,4-glucanase gene (celA1805) from Bacillus subtilis B111 in Escherichia coli. The recombinant celA1805 contains a glycosyl hydrolase (GH) family 8 domain and shared 76.8% identity with endo-1,4-β-glucanase from Bacillus sp. KSM-330. Results showed that the optimal pH and temperature of celA1805 were 6.0 and 50 o C, respectively, and it was stable at pH 3-9 and temperature ≤50°C. Metal ions slightly affected enzyme activity, but chemical agents generally inhibited enzyme activity. Moreover, celA1805 showed a wide substrate specificity to CMC, barley β-glucan, lichenin, chitosan, PASC and avicel. The K m and V max values of celA1805 were 1.78 mg/ml and 50.09 μmol/min/mg. When incubated with cellooligosaccharides ranging from cellotriose to cellopentose, celA1805 mainly hydrolyzed cellotetrose (G4) and cellopentose (G5) to cellose (G2) and cellotriose (G3), but hardly hydrolyzed cellotriose. The concentrations of reducing sugars saccharified by celA1805 from wheat straw, rape straw, rice straw, peanut straw, and corn straw were increased by 0.21, 0.51, 0.26, 0.36, and 0.66 mg/ml, respectively. The results obtained in this study suggest potential applications of celA1805 in biomass saccharification.

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Section: Characteristics Of Recombinant Cela1805supporting

confidence: 87%

Characterization of a GH8 β-1,4-Glucanase from Bacillus subtilis B111 and Its Saccharification Potential for Agricultural Straws

Huang

Zhao

et al. 2021

J. Microbiol. Biotechnol.

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“…Cellulose degradation is attributed to the synergistic action of three complementary enzyme activities: (1) endoglucanases (EGs); (2) cellobiohydrolases (CBHs); (3) β-glucosidases (BGs) [ 21 ]. In this study, a cellulase high-producer MEA-12 was obtained by compound mutagenesis combined with microbial ALE method.…”

Section: Resultsmentioning

confidence: 99%

Cellulase production and efficient saccharification of biomass by a new mutant Trichoderma afroharzianum MEA-12

Peng

Lin

et al. 2021

Biotechnol Biofuels

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Background Cellulase plays a key role in converting cellulosic biomass into fermentable sugar to produce chemicals and fuels, which is generally produced by filamentous fungi. However, most of the filamentous fungi obtained by natural breeding have low secretory capacity in cellulase production, which are far from meeting the requirements of industrial production. Random mutagenesis combined with adaptive laboratory evolution (ALE) strategy is an effective method to increase the production of fungal enzymes. Results This study obtained a mutant of Trichoderma afroharzianum by exposures to N-methyl-N’-nitro-N-nitrosoguanidine (MNNG), Ethyl Methanesulfonate (EMS), Atmospheric and Room Temperature Plasma (ARTP) and ALE with high sugar stress. The T. afroharzianum mutant MEA-12 produced 0.60, 5.47, 0.31 and 2.17 IU/mL FPase, CMCase, pNPCase and pNPGase, respectively. These levels were 4.33, 6.37, 4.92 and 4.15 times higher than those of the parental strain, respectively. Also, it was found that T. afroharzianum had the same carbon catabolite repression (CCR) effect as other Trichoderma in liquid submerged fermentation. In contrast, the mutant MEA-12 can tolerate the inhibition of glucose (up to 20 mM) without affecting enzyme production under inducing conditions. Interestingly, crude enzyme from MEA-12 showed high enzymatic hydrolysis efficiency against three different biomasses (cornstalk, bamboo and reed), when combined with cellulase from T. reesei Rut-C30. In addition, the factors that improved cellulase production by MEA-12 were clarified. Conclusions Overall, compound mutagenesis combined with ALE effectively increased the production of fungal cellulase. A super-producing mutant MEA-12 was obtained, and its cellulase could hydrolyze common biomasses efficiently, in combination with enzymes derived from model strain T. reesei, which provides a new choice for processing of bioresources in the future.

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“…It is an important structural components of plant’s primary cell wall and is also abundant in the cell walls of algae and the fungal class of oomycetes. Moreover, it has been discovered that some bacteria can secret cellulose biomers to form biofilms [ 39 ]. Cellulose appears as a crystalline white structure, has a melting point of 260–270 °C with 1.50 g/cm 3 density and 162.1406 g/mol molar mass, makes 90% of cotton fibre, about 57% in hemp, and constitutes 40–50% of wood [ 39 ].…”

Section: Lignin Modifying Fungi and Ligninolytic Consortiummentioning

confidence: 99%

“…Moreover, it has been discovered that some bacteria can secret cellulose biomers to form biofilms [ 39 ]. Cellulose appears as a crystalline white structure, has a melting point of 260–270 °C with 1.50 g/cm 3 density and 162.1406 g/mol molar mass, makes 90% of cotton fibre, about 57% in hemp, and constitutes 40–50% of wood [ 39 ]. Hemicellulose, on the other hand, exists as an heteropolymer of organic polysaccharide like arabinoxylans.…”

Section: Lignin Modifying Fungi and Ligninolytic Consortiummentioning

confidence: 99%

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Genome-based engineering of ligninolytic enzymes in fungi

et al. 2021

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Background Many fungi grow as saprobic organisms and obtain nutrients from a wide range of dead organic materials. Among saprobes, fungal species that grow on wood or in polluted environments have evolved prolific mechanisms for the production of degrading compounds, such as ligninolytic enzymes. These enzymes include arrays of intense redox-potential oxidoreductase, such as laccase, catalase, and peroxidases. The ability to produce ligninolytic enzymes makes a variety of fungal species suitable for application in many industries, including the production of biofuels and antibiotics, bioremediation, and biomedical application as biosensors. However, fungal ligninolytic enzymes are produced naturally in small quantities that may not meet the industrial or market demands. Over the last decade, combined synthetic biology and computational designs have yielded significant results in enhancing the synthesis of natural compounds in fungi. Main body of the abstract In this review, we gave insights into different protein engineering methods, including rational, semi-rational, and directed evolution approaches that have been employed to enhance the production of some important ligninolytic enzymes in fungi. We described the role of metabolic pathway engineering to optimize the synthesis of chemical compounds of interest in various fields. We highlighted synthetic biology novel techniques for biosynthetic gene cluster (BGC) activation in fungo and heterologous reconstruction of BGC in microbial cells. We also discussed in detail some recombinant ligninolytic enzymes that have been successfully enhanced and expressed in different heterologous hosts. Finally, we described recent advance in CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)-Cas (CRISPR associated) protein systems as the most promising biotechnology for large-scale production of ligninolytic enzymes. Short conclusion Aggregation, expression, and regulation of ligninolytic enzymes in fungi require very complex procedures with many interfering factors. Synthetic and computational biology strategies, as explained in this review, are powerful tools that can be combined to solve these puzzles. These integrated strategies can lead to the production of enzymes with special abilities, such as wide substrate specifications, thermo-stability, tolerance to long time storage, and stability in different substrate conditions, such as pH and nutrients.

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Construction and characterization of bifunctional cellulases: Caldicellulosiruptor-sourced endoglucanase, CBM, and exoglucanase for efficient degradation of lignocellulose

Cited by 33 publications

References 34 publications

Characterization of a GH8 β-1,4-Glucanase from Bacillus subtilis B111 and Its Saccharification Potential for Agricultural Straws

Characterization of a GH8 β-1,4-Glucanase from Bacillus subtilis B111 and Its Saccharification Potential for Agricultural Straws

Cellulase production and efficient saccharification of biomass by a new mutant Trichoderma afroharzianum MEA-12

Genome-based engineering of ligninolytic enzymes in fungi

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