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

Abstract 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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Arcopilus aureus MaC7A as a New Source of Resveratrol: Assessment of Amino Acid Precursors, Volatiles, and Fungal Enzymes for Boosting Resveratrol Production in Batch Cultures

Applied Sciences ◽

10.3390/app11104583 ◽

2021 ◽

Vol 11 (10) ◽

pp. 4583

Author(s):

Nemesio Villa-Ruano ◽

Luis Ángel Morales-Mora ◽

Jenaro Leocadio Varela-Caselis ◽

Antonio Rivera ◽

María de los Ángeles Valencia de Ita ◽

...

Keyword(s):

Amino Acid ◽

Filamentous Fungi ◽

Hydrolytic Enzymes ◽

Glucanase Activity ◽

Fungal Enzymes ◽

Activity Increase ◽

Batch Cultures ◽

Low Concentrations ◽

Chemical Factors ◽

Amino Acid Precursors

The chemical factors that regulate the synthesis of resveratrol (RV) in filamentous fungi are still unknown. This work reports on the RV production by Arcopilus aureus MaC7A under controlled conditions and the effect of amino acid precursors (PHE and TYR), monoterpenes (limonone, camphor, citral, thymol, menthol), and mixtures of hydrolytic enzymes (Glucanex) as elicitors for boosting fungal RV. Batch cultures with variable concentrations of PHE and TYR (50–500 mg L−1) stimulated RV production from 127.9 ± 4.6 to 221.8 ± 5.2 mg L−1 in basic cultures developed in PDB (pH 7) added with 10 g L−1 peptone at 30 °C. Maximum levels of RV and biomass were maintained during days 6–8 under these conditions, whereas a dramatic RV decrease was observed from days 10–12 without any loss of biomass. Among the tested volatiles, citral (50 mg L−1) enhanced RV production until 187.8 ± 2.2 mg L−1 in basic cultures, but better results were obtained with Glucanex (100 mg L−1; 198.3 ± 7.6 mg L−1 RV). Optimized batch cultures containing TYR (200 mg L−1), citral (50 mg L−1), thymol (50 mg L−1), and Glucanex (100 mg L−1) produced up to 237.6 ± 4.7 mg L−1 of RV. Our results suggest that low concentrations of volatiles and mixtures of isoenzymes with β-1, 3 glucanase activity increase the biosynthesis of fungal RV produced by A. aureus MaC7A in batch cultures.

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Methods of Random Mutagenesis of Aspergillus Strain for Increasing Kojic Acid Production

Current Pharmaceutical Biotechnology ◽

10.2174/1389201022666210615125004 ◽

2021 ◽

Vol 22 ◽

Author(s):

Herman Suryadi ◽

Marina Ika Irianti ◽

Tri Hastuti Septiarini

Keyword(s):

Acid Production ◽

Kojic Acid ◽

Ion Beam ◽

Random Mutagenesis ◽

Strain Improvement ◽

Temperature Plasma ◽

Acid Yield ◽

Chemical Mutagens ◽

Kojic Acid Production ◽

Physical And Chemical

: Kojic acid is an organic acid that is commonly used in the pharmaceutical and cosmetic industries. This acid compound is a secondary metabolite produced by various microorganisms, one of which is Aspergillus oryzae. Typically, improving the strain can enhance kojic acid production. A mutation is one of the tools to perform strain improvement because the change in kojic acid-producing genes effectively increases kojic acid yield. Random mutagenesis is a classic approach for inducing and producing mutants with random mutations. The mutagenesis can be generated by the individual physical and chemical mutagen, combined physical and chemical mutagens, or initiate by protoplast preparation. Aspergillus strains that are exposed to physical mutagens (e.g., UV) or chemical mutagens (e.g., N-methyl-N-nitro-N-nitrosoguanidine (NTG)) showed their abilities in increasing kojic acid production. Several new mutation methods, such as Ion Beam Implantation and Atmospheric and room temperature plasma (ARTP), also showed good responses in enhancing the production of biological products such as kojic acid. This review compared different random mutagenesis methods of Aspergillus strain with various mutagen types to provide better insight for researchers in choosing the most suitable method to increase kojic acid production.

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Adaptive Laboratory Evolution for Multistress Tolerance, including Fermentability at High Glucose Concentrations in Thermotolerant Candida tropicalis

Energies ◽

10.3390/en15020561 ◽

2022 ◽

Vol 15 (2) ◽

pp. 561

Author(s):

Koudkeo Phommachan ◽

Chansom Keo-oudone ◽

Mochamad Nurcholis ◽

Nookhao Vongvilaisak ◽

Mingkhuan Chanhming ◽

...

Keyword(s):

High Temperatures ◽

Candida Tropicalis ◽

High Glucose ◽

Glucose Transporter ◽

Major Facilitator Superfamily ◽

Cellulosic Biomass ◽

High Concentration ◽

Adaptive Laboratory Evolution ◽

Membrane Spanning ◽

Major Facilitator

Candida tropicalis, a xylose-fermenting yeast, has the potential for converting cellulosic biomass to ethanol. Thermotolerant C. tropicalis X-17, which was isolated in Laos, was subjected to repetitive long-term cultivation with a gradual increase in temperature (RLCGT) in the presence of a high concentration of glucose, which exposed cells to various stresses in addition to the high concentration of glucose and high temperatures. The resultant adapted strain demonstrated increased tolerance to ethanol, furfural and hydroxymethylfurfural at high temperatures and displayed improvement in fermentation ability at high glucose concentrations and xylose-fermenting ability. Transcriptome analysis revealed the up-regulation of a gene for a glucose transporter of the major facilitator superfamily and genes for stress response and cell wall proteins. Additionally, hydropathy analysis revealed that three genes for putative membrane proteins with multiple membrane-spanning segments were also up-regulated. From these findings, it can be inferred that the up-regulation of genes, including the gene for a glucose transporter, is responsible for the phenotype of the adaptive strain. This study revealed part of the mechanisms of fermentability at high glucose concentrations in C. tropicalis and the results of this study suggest that RLCGT is an effective procedure for improving multistress tolerance.

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Random Mutagenesis of Filamentous Fungi Strains for High-Yield Production of Secondary Metabolites: The Role of Polyamines

10.5772/intechopen.93702 ◽

2020 ◽

Author(s):

Alexander A. Zhgun

Keyword(s):

Filamentous Fungi ◽

Random Mutagenesis ◽

Strain Improvement ◽

Main Tool ◽

High Yield ◽

Individual Species ◽

Airborne Spores ◽

Yield Production ◽

Improvement Programs ◽

The Individual

A filamentous fungus (also called molds or moldy fungus) is a taxonomically diverse organism from phylum Zygomycota and Ascomycota with filamentous hyphae and has the ability to produce airborne spores or conidia. Currently, more than 70,000 molds are known, and some of them contain unique and unusual biochemical pathways. A number of products from such pathways, especially, the secondary metabolite (SM) pathways are used as important pharmaceuticals, including antibiotics, statins, and immunodepresants. Under different conditions, the individual species can produce more than 100 SM. The strain improvement programs lead to high yielding in target SM and significant reduction of spin-off products. The main tool for the strain improvement of filamentous fungi is random mutagenesis and screening. The majority of industrial overproducing SM strains were developed with the help of such technique over the past 50–70 years; the yield of the target SM increased by 100- to 1000-fold or more. Moreover, most of the strains have reached their technological limit of improvement. A new round of mutagenesis has not increased overproduction. Recently, it was shown that that the addition of exogenous polyamines may increase the production of such improved strains of filamentous fungi. The possible molecular mechanism of this phenomenon and its biotechnological applications are discussed.

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Green seaweeds (Ulva fasciata sp.) as nitrogen source for fungal cellulase production

World Journal of Microbiology and Biotechnology ◽

10.1007/s11274-019-2658-1 ◽

2019 ◽

Vol 35 (6) ◽

Cited By ~ 1

Author(s):

Joseph A. Bentil ◽

Anders Thygesen ◽

Lene Lange ◽

Moses Mensah ◽

Anne S. Meyer

Keyword(s):

Nitrogen Source ◽

Cellulase Production ◽

Ulva Fasciata ◽

Fungal Cellulase

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How Carbon Source and Degree of Oligosaccharide Polymerization Affect Production of Cellulase-Degrading Enzymes by Fusarium oxysporum f. sp. lycopersici

Frontiers in Microbiology ◽

10.3389/fmicb.2021.652655 ◽

2021 ◽

Vol 12 ◽

Author(s):

Nasim Najjarzadeh ◽

Leonidas Matsakas ◽

Ulrika Rova ◽

Paul Christakopoulos

Keyword(s):

Carbon Source ◽

Fusarium Oxysporum ◽

Filamentous Fungi ◽

Cellulase Production ◽

Degree Of Polymerization ◽

Cellulase Induction ◽

Degrading Enzymes

Cellulases are a group of enzymes responsible for the degradation of cellulose, which is one of the most abundant polymers on Earth. The three main groups of cellulases are endoglucosidases, exoglucosidases, and β-glucosidases; however, the mechanism of induction of these enzymes remains poorly characterized. Cellooligosaccharides are among the main inducers of these enzymes in filamentous fungi, yet it is not clear how their degree of polymerization may affect the strength of induction. In the present study, we investigated the effect of different carbohydrate-based inducers, such as lactose, sophorose, cellooligosaccharides, and xylooligosacharides, characterized by different concentrations and degree of polymerization, on cellulases production by the fungus Fusarium oxysporum f. sp. lycopersici, which is one of the most studied lignocellulose degrading fungi with the ability to consume both cellulose and hemicellulose. Moreover, the effect of carbon source on cellulase induction was assessed by growing the biomass on sucrose or glycerol. Results showed a correlation between induction efficiency and the cellooligosaccharides’ concentration and size, as well as the carbon source available. Specifically, cellotetraose was a better inducer when sucrose was the carbon source, while cellobiose yielded a better result on glycerol. These findings can help optimize industrial cellulase production.

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Cellulase Enzyme Production from Filamentous Fungi Trichoderma reesei and Aspergillus awamori in Submerged Fermentation with Rice Straw

Journal of Fungi ◽

10.3390/jof7100868 ◽

2021 ◽

Vol 7 (10) ◽

pp. 868

Author(s):

Laila Naher ◽

Siti Noor Fatin ◽

Md Abdul Halim Sheikh ◽

Lateef Adebola Azeez ◽

Shaiquzzaman Siddiquee ◽

...

Keyword(s):

Trichoderma Reesei ◽

Filamentous Fungi ◽

Rice Straw ◽

Submerged Fermentation ◽

Enzyme Production ◽

Dry Weight ◽

Cellulase Production ◽

Fungal Species ◽

Aspergillus Awamori ◽

Cellulase Enzyme

Fungi are a diverse group of microorganisms that play many roles in human livelihoods. However, the isolation of potential fungal species is the key factor to their utilization in different sectors, including the enzyme industry. Hence, in this study, we used two different fungal repositories—soil and weed leaves—to isolate filamentous fungi and evaluate their potential to produce the cellulase enzyme. The fungal strains were isolated using dichloran rose bengal agar (DRBA) and potato dextrose agar (PDA). For cellulase enzyme production, a rice straw submerged fermentation process was used. The enzyme production was carried out at the different incubation times of 3, 5, and 7 days of culture in submerged conditions with rice straw. Fungal identification studies by morphological and molecular methods showed that the soil colonies matched with Trichoderma reesei, and the weed leaf colonies matched with Aspergillus awamori. These species were coded as T. reesei UMK04 and A. awamori UMK02, respectively. This is the first report of A. awamori UMK02 isolation in Malaysian agriculture. The results of cellulase production using the two fungi incorporated with rice straw submerged fermentation showed that T. reesei produced a higher amount of cellulase at Day 5 (27.04 U/mg of dry weight) as compared with A. awamori (15.19 U/mg of dry weight), and the concentration was significantly different (p < 0.05). Our results imply that T. reesei can be utilized for cellulase production using rice straw.

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OPTIMIZATION OF CELLULASE ENZYME PRODUCTION FROM Aspergillus oryzae FOR INDUSTRIAL APPLICATIONS

World Journal of Biology and Biotechnology ◽

10.33865/wjb.002.02.0088 ◽

2017 ◽

Vol 2 (2) ◽

pp. 155 ◽

Cited By ~ 1

Author(s):

Hassan Sher ◽

Muhammad Faheem ◽

Abdul Ghani ◽

Rashid Mehmood ◽

Hamza Rehman ◽

...

Keyword(s):

Enzyme Production ◽

Time Course ◽

Value Added ◽

Cellulase Production ◽

Industrial Applications ◽

Inoculum Size ◽

Cellulosic Biomass ◽

High Yield ◽

Carboxymethyl Cellulase ◽

Cellulase Enzyme

Cellulases are the hydrolytic group of enzymes, responsible for release of sugars in the bioconversion of the cellulosic biomass into a variety of value added industrial products. Fungal isolated cellulases are well studied and playing a significant role in various industrial processes. Enzymatic depolymerisation of cellulosic material has been done by the various fungal isolated enzymes. In the present study, the cultivation conditions for cellulase production from Aspergillus species were optimized. Optimization of scarification conditions such as time course, inoculum size, carbon source and concentration, nitrogen source, various pH levels were performed for the production of extracellular carboxymethyl cellulase and endoglucanase enzyme. The result exhibited, 15 % inoculums size, corncobs 2 % concentration, Urea and medium pH 7 at 30oC supported high yield of carboxymethyl cellulase (38.80 U/ml/min) and exoglucanase enzyme (10.94 U/ml/min) through a submerged fermentation (SmF). In future biotechnological applications in cellulase enzyme production attain a vital role to obtain high degradable yield.

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