Production Of Lignocellulolytic Enzymes Research Articles

Developing endophytic Penicillium oxalicum as a source of lignocellulolytic enzymes for enhanced hydrolysis of biorefinery relevant pretreated rice straw.

Endophytic fungi, as plant symbionts, produce an elaborate array of enzymes for efficient disintegration of lignocellulosic biomass into constituent monomeric sugars, making them novel source of lignocellulolytic CAZymes with immense potential in future biorefineries. The present study reports lignocellulolytic enzymes production potential of an endophytic halotolerant Penicillium oxalicum strain isolated from Citrus limon, under submerged and solid-state fermentation (SmF & SSF, respectively), in the presence and absence of salt (1M NaCl). The comparative QTOF-LC/MS-based exoproteome analysis of the culture extracts unveiled differential expression of CAZymes, with the higher abundance of GH6 and GH7 family cellobiohydrolase in the presence of 1M salt. The strain improvement program, employing cyclic mutagenesis and diploidization, was utilized to develop hyper-cellulase producing mutant strains of P. oxalicum. The enzyme production of the developed strain (POx-M35) was further enhanced through statistical optimization of the culture conditions utilizing glucose mix disaccharides (GMDs) as an inducer. This optimization process resulted in the lignocellulolytic cocktail that contained high titers (U/mL) of endoglucanase (EG) (146.16), cellobiohydrolase (CBHI) (6.99), β-glucosidase (β-G) (26.21), xylanase (336.05) and FPase (2.02 U/mL), which were 5.47-, 5.54-, 8.55-, 4.96-, and 4.39-fold higher when compared to the enzyme titers obtained in wild HP1, respectively. Furthermore, the lignocellulolytic cocktails designed by blending secretome produced by mutant POx-M35 with xylanases (GH10 and GH11) derived from Malbranchea cinnamomea resulted in efficient hydrolysis of unwashed acid pretreated (UWAP) rice straw slurry and mild alkali deacetylated (MAD) rice straw. This study underscores the potential of bioprospecting novel fungus and developing an improved strain for optimized production and constitution of lignocellulolytic cocktails that can be an important determinant in advancing biomass conversion technologies.

Genomic analysis of a halophilic bacterium Nesterenkonia sp. CL21 with ability to produce a diverse group of lignocellulolytic enzymes.

Halophilic bacteria are extremophiles that thrive in saline environment. Their ability to withstand such harsh conditions makes them an ideal choice for industrial applications such as lignocellulosic biomass degradation. In this study, a halophilic bacterium with the ability to produce extracellular cellulases and hemicellulases, designated as Nesterenkonia sp. CL21, was isolated from mangrove sediment in Tanjung Piai National Park, Malaysia. Thus far, studies on lignocellulolytic enzymes concerning bacterial species under this genus are limited. To gain a comprehensive understanding of its lignocellulose-degrading potential, the whole genome was sequenced using the Illumina NovaSeq 6000 platform. The genome of strain CL21 was assembled into 25 contigs with 3,744,449 bp and a 69.74% GC content and was predicted to contain 3,348 coding genes. Based on taxonomy analysis, strain CL21 shares 73.8 to 82.0% average nucleotide identity with its neighbouring species, below the 95% threshold, indicating its possible status as a distinct species in Nesterenkonia genus. Through in-depth genomic mining, a total of 81 carbohydrate-active enzymes were encoded. Among these, 24 encoded genes were identified to encompass diverse cellulases (GH3), xylanases (GH10, GH11, GH43, GH51, GH127 and CE4), mannanases (GH38 and GH106) and pectinases (PL1, PL9, and PL11). The production of lignocellulolytic enzymes was tested in the presence of several substrates. This study revealed that strain CL21 can produce a diverse array of enzymes which are active at different time points. By combining experimental data with genomic information, the ability of strain CL21 to produce lignocellulolytic enzymes has been elucidated, with potential applications in biorefinery industry.

Influence of metals on lignocellulolytic enzyme production and metal removal by Phlebia brevispora in a wheat straw based fixed‐bed bioreactor

AbstractHeavy metals contamination poses a significant environmental threat, which requires the development of eco‐friendly bioremediation techniques. Present research work was conducted using wheat straw in submerged conditions for the growth of white rot fungus, Phlebia brevispora in the presence of varying concentrations of Cd, Pb, Ni, and Cr. The lignocellulolytic enzyme production ability of fungus was monitored. A wheat straw‐based fixed‐bed bioreactor was designed to treat metal‐containing water in continuous mode. The fungus showed a positive influence on laccase production, cellulase production, and lignin peroxidase activity. As observed through atomic absorption spectroscopy (AAS), Cr removal efficiency was 78%–80%, regardless of the initial metal concentration. The Cr was also found on the surface of fungal mycelium as per the results obtained from SEM–EDX. The continuous bioreactor achieved 98%–99% metal removal, making it a natural and cost‐effective solution for metal removal from wastewater using P. brevispora.

Small GTPase Rab7 is involved in stress adaptation to carbon starvation to ensure the induced cellulase biosynthesis in Trichoderma reesei.

The saprophytic filamentous fungus Trichoderma reesei represents one of the most prolific cellulase producers. The bulk production of lignocellulolytic enzymes by T. reesei not only relies on the efficient transcription of cellulase genes but also their efficient secretion after being translated. However, little attention has been paid to the functional roles of the involved secretory pathway in the high-level production of cellulases in T. reesei. Rab GTPases are key regulators in coordinating various vesicle trafficking associated with the eukaryotic secretory pathway. Specifically, Rab7 is a representative GTPase regulating the transition of the early endosome to the late endosome followed by its fusion to the vacuole as well as homotypic vacuole fusion. Although crosstalk between the endosomal/vacuolar pathway and the secretion pathway has been reported, the functional role of Rab7 in cellulase production in T. reesei remains unknown. A TrRab7 was identified and characterized in T. reesei. TrRab7 was shown to play important roles in T. reesei vegetative growth and vacuole morphology. Whereas knock-down of Trrab7 significantly compromised the induced production of T. reesei cellulases, overexpression of the key transcriptional activator, Xyr1, restored the production of cellulases in the Trrab7 knock-down strain (Ptcu-rab7KD) on glucose, indicating that the observed defective cellulase biosynthesis results from the compromised cellulase gene transcription. Down-regulation of Trrab7 was also found to make T. reesei more sensitive to various stresses including carbon starvation. Interestingly, overexpression of Snf1, a serine/threonine protein kinase known as an energetic sensor, partially restored the cellulase production of Ptcu-rab7KD on Avicel, implicating that TrRab7 is involved in an energetic adaptation to carbon starvation which contributes to the successful cellulase gene expression when T. reesei is transferred from glucose to cellulose. TrRab7 was shown to play important roles in T. reesei development and a stress response to carbon starvation resulting from nutrient shift. This adaptation may allow T. reesei to successfully initiate the inducing process leading to efficient cellulase production. The present study provides useful insights into the functional involvement of the endosomal/vacuolar pathway in T. reesei development and hydrolytic enzyme production.

Successive process for efficient biovalorization of Brewers’ spent grain to lignocellulolytic enzymes and lactic acid production through simultaneous saccharification and fermentation

This study aimed to increase the value of brewers’ spent grain (BSG) by using it as feedstock to produce lignocellulolytic enzymes and lactic acid (LA). Twenty-two fungal strains were screened for lignocellulolytic enzyme production from BSG. Among them, Trichoderma sp. showed the highest cellulase activity (35.84 ± 0.27 U/g-BSG) and considerably high activities of xylanase (599.61 ± 23.09 U/g-BSG) and β-glucosidase (16.97 ± 0.77 U/g-BSG) under successive solid-state and submerged fermentation. The processes were successfully scaled up in a bioreactor. The enzyme cocktail was recovered and characterized. The maximum cellulase and xylanase activities were found at pH 5.0 and 50 °C, and the activities were highly stable at pH 4–8 and 30–50 °C. The enzyme cocktail was applied in simultaneous saccharification and fermentation of acid-pretreated BSG for LA production. The maximum LA obtained was 59.3 ± 1.0 g/L. This study has shown the efficient biovalorization of BSG, and this approach may also be applicable to other agro-industrial wastes.

Effect of co-cultivation of white and brown rot species on basidiome production, lignocelluloytic enzyme activity and dye decolourisation

The aim of this work was studying the impact of co-cultivating two mushroom species: a white (Pleurotus albidus CLA 45) and a brown rot one (Laetiporus sulphureus BAFC 205) in substrates based on poplar or pine sawdust, on their lignocellulolytic enzyme production, yield values and basidiomes properties. Laetiporus sulphureus only developed basidiome primordia, but P. albidus monoculture and co-culture in pine sawdust achieved biological efficiencies of up to 50–55 %. Co-cultivation on diverse substrates rendered varied enzyme titers. Laccase and Manganese peroxidase titers were highest in pine co-culture and P. albidus poplar monoculture, respectively. Enzymatic extracts obtained from spent poplar substrate of dual cultures displayed potential for treating non-sterile textile-coloured effluents, achieving 35 % decolourisation after 120 h. The knowledge available on the effects of co-culture of white and brown rot fungi is still limited. This study represents an initial exploration of the interaction between them within intensive cultivation conditions.

A closed-loop strategy for on-site production of saccharolytic enzymes for lignocellulose biorefinery using internal lignocellulosic hydrolysates

The cost-effective production of lignocellulose-saccharolytic enzymes is crucial for the bioconversion of plant biomass into fuels and chemicals. The integrated mode for producing these enzymes using lignocellulosic feedstocks as natural inducers has a cost advantage; however, enzyme production level is limited because of the solid-state and complex composition of the raw materials. This study aimed to establish an innovative process for the integrated production of lignocellulolytic enzymes using lignocellulosic hydrolysate as the carbon source. The cellulase-producing workhorse Trichoderma reesei was reprogrammed by combinatorial engineering of transcription factors and the introduction of heterologous β-glucosidases. This enabled inducer-independent production of a complete lignocellulose-saccharolytic enzyme cocktail using lignocellulose-derived monosaccharides as sole carbon source. Via continuous feeding of the enzymatic hydrolysate of pretreated corn stover, crude enzymes with filter paper enzyme activity of 60.4 U/ml and β-glucosidase activity of 503 U/ml were produced. Crude fermentation broth can be used directly to prepare hydrolysates, enabling nex-round enzyme production by recycling 3.1% of the hydrolysate. These results demonstrate a closed-loop strategy for integrated enzyme production using internally produced lignocellulosic sugars from biorefinery plants.

Isolation, screening wood rot fungi from the tropical forest of Thailand and their lignocellulolytic enzyme production under solid-state fermentation using agricultural waste as substrate

Isolation, screening wood rot fungi from the tropical forest of Thailand and their lignocellulolytic enzyme production under solid-state fermentation using agricultural waste as substrate

Structure-guided engineering of transcriptional activator XYR1 for inducer-free production of lignocellulolytic enzymes in Trichoderma reesei

The filamentous fungus Trichoderma reesei is widely used for the production of lignocellulolytic enzymes in industry. XYR1 is the major transcriptional activator of cellulases and hemicellulases in T. reesei. However, rational engineering of XYR1 for improved lignocellulolytic enzymes production has been limited by the lack of structure information. Here, alanine 873 was identified as a new potential target for the engineering of XYR1 based on its structure predicted by AlphaFold2. The mutation of this residue to tyrosine enabled significantly enhanced production of xylanolytic enzymes in the medium with cellulose as the carbon source. Moreover, xylanase and cellulase production increased by 56.7- and 3.3-fold, respectively, when glucose was used as the sole carbon source. Under both conditions, the improvements of lignocellulolytic enzyme production were higher than those in the previously reported V821F mutant. With the enriched hemicellulases and cellulases, the crude enzymes secreted by the A873Y mutant strain produced 51 % more glucose and 52 % more xylose from pretreated corn stover than those of the parent strain. The results provide a novel strategy for engineering the lignocellulolytic enzyme-producing capacity of T. reesei, and would be helpful for understanding the molecular mechanisms of XYR1 regulation.

CRISPR/Cas9 mediated gene editing of transcription factor ACE1 for enhanced cellulase production in thermophilic fungus Rasamsonia emersonii

BackgroundThe filamentous fungus Rasamsonia emersonii has immense potential to produce biorefinery relevant thermostable cellulase and hemicellulase enzymes using lignocellulosic biomass. Previously in our lab, a hyper-cellulase producing strain of R. emersonii was developed through classical breeding and system biology approaches. ACE1, a pivotal transcription factor in fungi, plays a crucial role in negatively regulating the expression of cellulase genes. In order to identify the role of ACE1 in cellulase production and to further improve the lignocellulolytic enzyme production in R. emersonii, CRISPR/Cas9 mediated disruption of ACE1 gene was employed.ResultsA gene-edited ∆ACE1 strain (GN11) was created, that showed 21.97, 20.70 and 24.63, 9.42, 18.12%, improved endoglucanase, cellobiohydrolase (CBHI), β-glucosidase, FPase, and xylanase, activities, respectively, as compared to parental strain M36. The transcriptional profiling showed that the expression of global regulator (XlnR) and different CAZymes genes including endoglucanases, cellobiohydrolase, β-xylosidase, xylanase, β-glucosidase and lytic polysaccharide mono-oxygenases (LPMOs) were significantly enhanced, suggesting critical roles of ACE1 in negatively regulating the expression of various key genes associated with cellulase production in R. emersonii. Whereas, the disruption of ACE1 significantly down-regulated the expression of CreA repressor gene as also evidenced by 2-deoxyglucose (2-DG) resistance phenotype exhibited by edited strain GN11 as well as appreciably higher constitutive production of cellulases in the presence of glucose and mixture of glucose and disaccharide (MGDs) both in batch and flask fed batch mode of culturing. Furthermore, ∆ACE1 strains were evaluated for the hydrolysis of biorefinery relevant steam/acid pretreated unwashed rice straw slurry (Praj Industries Ltd; 15% substrate loading rate) and were found to be significantly superior when compared to the benchmark enzymes produced by parent strain M36 and Cellic Ctec3.ConclusionsCurrent work uncovers the crucial role of ACE1 in regulating the expression of the various cellulase genes and carbon catabolite repression mechanism in R. emersonii. This study represents the first successful report of utilizing CRISPR/Cas9 genome editing technology to disrupt the ACE1 gene in the thermophlic fungus R. emersonii. The improved methodologies presented in this work might be applied to other commercially important fungal strains for which genetic manipulation tools are limited.

Lignocellulose Degrading Weizmannia coagulans Capable of Enantiomeric L-Lactic Acid Production via Consolidated Bioprocessing

Second-generation lactic acid production requires the development of sustainable and economically feasible processes and renewable lignocellulose biomass as a starting raw material. Weizmannia coagulans MA42 was isolated from a soil sample in Chiang Mai province, Thailand and showed the highest production of L-lactic acid and lignocellulolytic enzymes (cellulase, β-mannanase, xylanase, β-glucosidase, β-mannosidase, and β-xylosidase) compared to other isolates. Weizmannia coagulans MA42 was able to grow, secrete lignocellulolytic enzymes, and directly produce L-lactic acid in the medium containing various lignocellulosic feedstocks as the sole carbon source. Moreover, L-lactic acid production efficiency was improved after the substrates were pretreated with diluted sulfuric acid and diluted sodium hydroxide. The highest L-lactic acid production efficiency of 553.4 ± 2.9, 325.4 ± 4.1, 326.6 ± 4.4, 528.0 ± 7.2, and 547.0 ± 2.2 mg/g total available carbohydrate was obtained from respective pretreated substrates including sugarcane bagasse, sugarcane trash, corn stover, rice straw, and water hyacinth. It is suggested that structural complexity of the lignocellulosic materials and properties of lignocellulolytic enzymes are the key factors of consolidated bioprocessing (CBP) of lignocellulosic feedstocks to lactic acid. In addition, the results of this study indicated that W. coagulans MA42 is a potent bacterial candidate for CBP of a variety of lignocellulosic feedstocks to L-lactic acid production; however, further bioprocess development and genetic engineering technique would provide higher lactic acid production efficiency, and this would lead to sustainable lactic acid production from lignocellulosic feedstocks.

Single Mutation in Transcriptional Activator Xyr1 Enhances Cellulase and Xylanase Production in Trichoderma reesei on Glucose.

To achieve cost-effective production of lignocellulolytic enzymes for biorefinery processes, engineering transcription factors represents a powerful strategy to boost cellulase and xylanase in Trichoderma reesei. In this study, a novel mutation (R434L) in xylanase regulator 1 (Xyr1) was identified based on the yeast one-hybrid screening system. The point mutation was located in the middle homology region of Xyr1 with unclear functions, indicating a significant role for this domain in tuning Xyr1 transactivation. When constitutively expressed in T. reesei Δxyr1 (OEXR434L), Xyr1R434L led to highly improved production of both cellulases and xylanases on glucose compared with a strain similarly expressing Xyr1 (OEX). The respective 0.8- and 0.7-fold increases in extracellular pNPCase and xylanolytic activity were further verified to result from the greatly elevated transcription of major cellulase and xylanase genes in OEXR434L. Moreover, the saccharification efficiency of corn stover with OEXR434L enzyme cocktails was enhanced by 21% compared with that of OEX.

Lignocellulolytic enzyme cocktail produced by plant endophytic Chaetomium globosum exhibits a capacity for high-efficient saccharification of raw rice straw

The aim of this work was to optimize the production of lignocellulolytic enzyme cocktail by a plant endophyte Chaetomium globosum DX-THS3 under solid-state fermentation, and to evaluate the saccharification efficiency of this enzyme cocktail on raw rice straw. The process variables, including substrate type, substrate size, initial moisture content of substrate, culture temperature and fermentative time, were optimized using One-factor at a time approach and Response surface methodology. The results showed that the maximum CMCase, FPase and xylanase production reached 237.04 U/g, 54.8 U/g, and 987.33 U/g, respectively. Meanwhile, the lignocellulolytic enzyme cocktail exhibited good thermal stability between 30 °C and 60 °C, and excellent tolerance to acidic pH values of 3–6. Subsequently, the saccharification of raw rice straw by the enzyme cocktail was performed. After 24 h hydrolysis of raw rice straw, high reducing sugars yield (294.36 mg/g) and hydrolytic rate (47.58 %) were achieved. Furthermore, scanning electron micrograph also indicated that the enzyme cocktail had high hydrolysis performance for raw rice straw. Overall, our result provides a novel class of lignocellulolytic enzyme cocktail with a capacity for high-efficient and eco-friendly saccharification of raw rice straw, and strongly suggests plant endophytic fungi as excellent candidates for the production of lignocellulolytic enzymes.

Efecto del tamaño de partícula, aireación y altura del sustrato sobre la producción de enzimas lignocelulolíticas producidas por Trametes polyzona HHM001 crecido sobre residuos de hojas de maíz

The objective of the work was to evaluate the production of lignocllulolytic enzymes produced by Trametes polyzona HHM001 during its growth on corn leaf residues. Two particle sizes (PS) (PS8 and PS12), two levels of aeration (1 vvm and 0 vvm), as well as the height of the substrate in the production of lignocellulolytic enzymes were tested. The enzymatic activities of Laccase (Lcc), Lignin peroxidase (LiP), and Manganese peroxidase (MnP) were favored under aerated conditions (1 vvm). The enzymatic activity of Lcc was the most favored, with 80 Activity Units (AU)/mL, compared to the culture without aeration, in which 40 AU/mL was obtained. The production of xylanases (Xyl) and cellulases (Cel) were not influenced by aeration under the tested conditions. The results indicated that the particle size has more effect on enzyme production than the presence or absence of air, with a particle size of 8 where the best levels of enzymatic activity were observed. It was observed that the height of the substrate in the fermentation column strongly affects the activity of ligninolytic enzymes and not of the hydrolytic; at 5 cm, the highest ligninolytic activity was detected, where aeration favors oxidative conditions.

Conversion of lignocellulosic biomass: Production of bioethanol and bioelectricity using wheat straw hydrolysate in electrochemical bioreactor

The present study evaluated efficiency of wheat straw (WS) hydrolysate obtained through fungal pre-treatment to produce ethanol and electricity in an electrochemical bioreactor. Three white rot fungi Phanerochaete chrysosporium, Phlebia floridensis and Phlebia brevispora were used to degrade WS for hydrolysate preparation, Lignocellulolytic enzyme production was also monitored during the pretreatment. Yeast Pichia fermentans was allowed to ferment all three hydrolysates up to 12 days. The yeast showed maximum electrochemical response as open circuit voltage (0.672 V), current density 542.42 mA m−2, and power density of 65.09 mW m−2 on 12th day in the hydrolysate prepared using Phlebia floridensis. Maximum ethanol production of 9.2% (w/v) was achieved on 7th day with a fermentation efficiency of about 62.1%. Further, the coulombic efficiency improved from 0.06 to 1.46% during 12 days of the experiment. Thus, the results indicated towards the possible conversion of lignocellulosic biomass into bioethanol along with bioelectricity generation.

Characterization of fungi isolated from cattle dung with potential lignocellulolytic activities

Agricultural wastes not only serve as sources of environmental pollutions, but also serve as magnificent source of plant nutrients and natural repositories of bio-tools especially fungi of industrial interest for lignocellulolytic enzymes production. However, residues management particularly in rice–wheat system is a tedious phenomenon due to narrow gap between harvesting and sowing of these two crops. Hence, microbial intervention is one of the key options that may use successfully for their management. Thus, cellulolytic fungi were isolated from the cattle dung collected from various locations for cellulose manufacturing. The cellulolytic property of fungal isolates was confirmed by plate screening assay based on yellow zone formation surrounding of fungal culture on Congo red agar medium plate owing to production of cellulase enzyme. The colony diameter, clear zone diameter and cellulolytic index showing in descending order of CRD3, CRD5, CRD6 and CRD10 isolates, but –glucosidase activity was greater in CRD3 and CRD5 while, the lowest was in CRD9. However, isolates CRD7, CRD8 and CRD10 exhibited greater biochemical and enzymatic activity than rest of the promising fungal isolates. Similarly, isolates CRD1 and CRD2 utilized most of carbon sources as carbon substrate, but CRD7, CRD8 and CRD10 could not utilized mannitol and cellulose as carbon substrate. Overall, isolate CRD10 showed as a promising isolate in respect of biochemical, carbon utilization pattern and enzymatic activity than rest of the promising isolates.

Microbial Lignocellulolytic Enzymes for the Effective Valorization of Lignocellulosic Biomass: A Review

The urgent demand for alternative energy sources has been sparked by the tremendous burden on fossil fuels and the resulting acute energy crisis and climate change issues. Lignocellulosic biomass is a copious renewable and alternative bioresource for the generation of energy fuels and biochemicals in biorefineries. Different pretreatment strategies have been established to overcome biomass recalcitrance and face technological challenges, such as high energy consumption and operational costs and environmental hazards, among many. Biological pretreatment using microbial enzymes is an environmentally benign and low-cost method that holds promising features in the effective pretreatment of lignocellulosic biomass. Due to their versatility and eco-friendliness, cellulases, hemicellulases, and ligninolytic enzymes have been recognized as “green biocatalysts” with a myriad of industrial applications. The current review provides a detailed description of different types of lignocellulolytic enzymes, their mode of action, and their prospective applications in the valorization of lignocellulosic biomass. Solid state fermentation holds great promise in the microbial production of lignocellulolytic enzymes owing to its energy efficient, environment friendly, and higher product yielding features utilizing the lignocellulosic feedstocks. The recent trends in the application of enzyme immobilization strategies for improved enzymatic catalysis have been discussed. The major bottlenecks in the bioprocessing of lignocellulosic biomass using microbial enzymes and future prospects have also been summarized.

Tailoring the expression of Xyr1 leads to efficient production of lignocellulolytic enzymes in Trichoderma reesei for improved saccharification of corncob residues

BackgroundThe filamentous fungus Trichoderma reesei is extensively used for the industrial-scale cellulase production. It has been well known that the transcription factor Xyr1 plays an important role in the regulatory network controlling cellulase gene expression. However, the role of Xyr1 in the regulation of cellulase expression has not been comprehensively elucidated, which hinders further improvement of lignocellulolytic enzyme production.ResultsHere, the expression dosage of xyr1 was tailored in T. reesei by differentially overexpressing the xyr1 gene under the control of three strong promoters (Pegl2, Pcbh1, and Pcdna1), and the transcript abundance of xyr1 was elevated 5.8-, 12.6-, and 47.2-fold, respectively. We found expression of cellulase genes was significantly increased in the Pegl2-driven xyr1 overexpression strain QE2X, whereas relatively low in the Pcbh1- and Pcdna1-driven overexpression strains. We also found that the Pegl2-driven overexpression of xyr1 caused a more significant opening of chromatin in the core promoter region of the prominent cellulase genes. Furthermore, the cellulase activity showed a 3.2-fold increase in the strain QE2X, while insignificant improvement in the Pcbh1- and Pcdna1-driven strains. Finally, the saccharification efficiency toward acid-pretreated corncob residues containing high-content lignin by the crude enzyme from QE2X was increased by 57.2% compared to that from the parental strain. Moreover, LC–MS/MS and RT-qPCR analysis revealed that expression of accessory proteins (Cip1, Cip2, Swo1, and LPMOs) was greatly improved in QE2X, which partly explained the promoting effect of the Pegl2-driven overexpression on enzymatic hydrolysis of lignocellulose biomass.ConclusionsOur results underpin that the precise tailoring expression of xyr1 is essential for highly efficient cellulase synthesis, which provide new insights into the role of Xyr1 in regulating cellulase expression in T. reesei. Moreover, these results also provides a prospective strategy for strain improvement to enhance the lignocellulolytic enzyme production for use in biorefinery applications.

Integrating 1G with 2G Bioethanol Production by Using Distillers’ Dried Grains with Solubles (DDGS) as the Feedstock for Lignocellulolytic Enzyme Production

First-generation (1G) bioethanol is one of the most used liquid biofuels in the transport industry. It is generated by using sugar- or starch-based feedstocks, while second-generation (2G) bioethanol is generated by using lignocellulosic feedstocks. Distillers’ dried grains with solubles (DDGS) is a byproduct of first-generation bioethanol production with a current annual production of 22.6 million tons in the USA. DDGS is rich in fiber and valuable nutrients contents, which can be used to produce lignocellulolytic enzymes such as cellulases and hemicellulases for 2G bioethanol production. However, DDGS needs a pretreatment method such as dilute acid, ammonia soaking, or steam hydrolysis to release monosaccharides and short-length oligosaccharides as fermentable sugars for use in microbial media. These fermentable sugars can then induce microbial growth and enzyme production compared to only glucose or xylose in the media. In addition, selection of one or more suitable microbial strains, which work best with the DDGS for enzyme production, is also needed. Media optimization and fermentation process optimization strategies can then be applied to find the optimum conditions for the production of cellulases and hemicellulases needed for 2G bioethanol production. Therefore, in this review, a summary of all such techniques is compiled with a special focus on recent findings obtained in previous pieces of research conducted by the authors and by others in the literature. Furthermore, a comparison of such techniques applied to other feedstocks and process improvement strategies is also provided. Overall, dilute acid pretreatment is proven to be better than other pretreatment methods, and fermentation optimization strategies can enhance enzyme production by considerable folds with a suitable feedstock such as DDGS. Future studies can be further enhanced by the technoeconomic viability of DDGS as the on-site enzyme feedstock for the manufacture of second-generation bioethanol (2G) in first-generation (1G) ethanol plants, thus bridging the two processes for the efficient production of bioethanol using corn or other starch-based lignocellulosic plants.

Genome sequence analysis of halophilic Luteibacter sp. CQ10 to prospect its dual roles in antioxidants production and lignocellulose degradation

Luteibacter is a bacterial genus affiliated to Rhodanobacteraceae family with five species reported up to date. Currently, limited information regarding this genus was revealed, especially on its genomic perspective and its potential application. Here, we present the whole genome sequence of Luteibacter sp. CQ10, which the strain was isolated from mangrove bark collected at Tanjung Piai National Park, Johor, Malaysia. The genome of Luteibacter sp. CQ10 contains 30 contigs, with a genome size of 4.87 Mbp, a GC content of 66.44 % and a total of 4197 predicted genes. The preliminary analysis elucidated that a total of 10 and 22 annotated genes were predicted to participate in biosynthesis of antioxidants and production of lignocellulolytic enzymes, respectively. These biomolecules are beneficial to the food and biorefinery industries.