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Fucoidans are fucose rich sulfated polysaccharides that are found in the cell wall of brown seaweeds and have been shown to have several beneficial bioactivities. In the present study, we report a new enzymatic extraction technique for the production of pure and intact fucoidans from the two brown seaweeds Saccharina latissima and Alaria esculenta. This new extraction protocol uses the commercial cellulase blend Cellic® CTec2 in combination with endo - and exo -acting thermophilic alginate lyases. The fucoidans obtained by this extraction technique are compared to traditionally extracted fucoidans in terms of chemical compositions and molecular weights and are shown to contain significantly higher amounts of fucose and sulfate, the main components of fucoidans, while cellulose, laminarin, and alginate contamination is low. Thus, by using this combination of enzymes, the extracted fucoidans do not undergo depolymerization during extraction and additional purification steps are not needed. The high purity fucoidans isolated by this new enzymatic extraction technique can be used to provide insight into the different fucoidan structures and biological activities. • New enzymatic extraction technique for the production of pure and intact fucoidans • Efficient fucoidan extraction using alginate lyases and cellulases • Pure fucoidans with very low content of cellulose, laminarin and alginate • Enzymically extracted fucoidans with high molecular weights • No need for purification steps after fucoidan extraction [ABSTRACT FROM AUTHOR]

In this study, Lentinula edodes and Pleurotus ostreatus were selected as representatives to explore the effects of corn straw and xylosma sawdust on the production and activity of polysaccharides by edible fungi during solid-state fermentation. The results clearly indicated that the addition of lignocellulose could promote the polysaccharide content compared to the control group. Meanwhile, the hydroxyl radical scavenging activity of polysaccharides reached the maximum when the glucose concentration was 1.5 %, and among them, the xylosma sawdust groups for two fungi (L. edodes -32.37 %, P. ostreatus -25.86 %) both performed better than corn straw groups (L. edodes -24.96 %, P. ostreatus -20.80 %). In addition, structural characterization and degradation mode analysis were carried out. The results showed that the structure of the xylosma had a stronger destruction than corn straw. The activities of lignocellulolytic enzymes such as carboxymethyl cellulase, filter paper enzyme, β-glucosidase and xylanase increased rapidly in the early stage of fermentation, which could degrade the cellulose and hemicellulose as raw materials for the synthesis of active polysaccharides. [ABSTRACT FROM AUTHOR]

Two novel Bacillus glycoside hydrolase family 53 (GH53) endo-1,4-β-galactanases (Bs936 and Bs4828) were identified. The recombinant Bs936 and Bs4828 displayed maximal activities at pH 5.5/55 °C and pH 6.5/65 °C, respectively. Stability analyses revealed that the enzymes were stable between pH 4.5–10, retaining over 80% activities after 12 h incubation at 25 °C. Moreover, Bs936 maintained about 75% activity after being treated at 45 °C for 2 h, while Bs4828 kept full activity after 4 h of incubation at 50 °C. Importantly, Bs936 and Bs4828 exhibited good activity towards β-1,4-galactan, showing specific activities of 1859.46- and 3110.79 U/mg towards potato galactan, respectively. The corresponding K m and k cat /K m were 4.93 mg/mL and 296.35 mL/mg/s for Bs936, and 6.60 mg/mL and 707.12 mL/mg/s for Bs4828. Extensive synergy assays revealed that each endo-1,4-β-galactanase acted synergistically with the other β-galactosidase (Bs937 or Bs4829), endo-1,5-α-L-arabinosidase (BsAbn2) and an endoxylanase/endoglucanase+cellulase mixture (BaXynA + LsCel) towards the hydrolysis of galactan, debranched arabinan/arabinan, and pretreated corn stover, respectively. The corresponding degrees of synergy were 1.28, 1.17/1.09, and 1.15 for Bs486 and 1.55, 1.04/1.08, and 1.11 for Bs4828, respectively. This study expands the repertoire of GH53 enzymes, and demonstrates the role of two highly active endo-1,4-β-galactanases in polysaccharides hydrolysis. [ABSTRACT FROM AUTHOR]

Low pretreatment temperatures are beneficial for enhancing the activity of both soluble lignin and lignin in pretreated solids. To achieve high-activity lignin and a higher glucose yield at a lower lignin removal rate and reduced enzymatic loading, the synergistic effects of octyl glucoside and tea saponin during low-temperature ammonia‑oxygen pretreatment and enzymatic hydrolysis were explored. Utilizing a moderated lignin extraction at 38 %, with pretreatment at 110 °C, and enriched with 1 % octyl glucoside and 0.06 g/g tea saponin, the biochemical conversion was galvanized. This manifested in an enzymatic hydrolysis efficiency of 96 % and a glucose yield in excess of 85 % at an enzyme loading of 5 FPU/g of pretreated-solids. Conversely, the lignin removal rate without surfactants was 26 %, resulting in 70 % hydrolysis efficiency even at a higher enzyme loading of 15 FPU/g of pretreated-solids. In order to minimize the pretreatment temperature and enzyme load as much as possible without affecting the efficiency of enzymatic hydrolysis and glucose yield, the goal of this research is to investigate the roles of octyl glucoside in the pretreatment process and tea saponin in the enzymatic hydrolysis process. This study provides a new approach for refining lignocellulosic biomass. • Using surfactants to sustain high enzyme activity with low intensive pretreatment • Enzyme hydrolysis efficiency raised by adding octyl glucoside and tea saponin • The glucose yield increased to over 85 % by adding surfactants. • Low enzyme loading (5 FPU/g–PSs) was used and 67 % of cellulase can be saved. • This study provided a foundation for the high-value utilization of lignin. [ABSTRACT FROM AUTHOR]

This study focused on the morphology regulation of nanocelluloses (NCs) aiming at preparing NCs with the controllable properties. NCs with rod-shaped, spherical, and rod-shaped and spherical composite structures were prepared from waste paper by a green enzymatic hydrolysis. The critical enzyme concentration for the transition of NCs from one morphology to another was explored. Several analytical methods, including TGA, XRD, SEM, TEM, DSC and WCA, were employed to characterize the prepared NCs. The results showed that with the cellulase enzyme concentration lower than 6 μ/ml or higher than 30 μ/ml, the prepared NCs had a single rod-shaped or spherical structure. When the cellulase concentration was between 6 and 30 μ/ml, the prepared NCs had a composite morphology of rods and spheres. Besides, NCs with a composite morphology dominated by rod-like or spherical structures had a higher thermal stability. Moreover, NCs prepared with cellulase enzyme concentration of 6 and 35 μ/ml had the highest and lowest crystallinity of 81.7 % and 59.3 %, respectively. Hence, this work achieved the morphology and property regulation of NCs by a green method which expanded the application scope of NCs and realized the high value utilization of waste paper. [Display omitted] • Prepared nanocellulose by a green hydrolysis method • Achieved the morphology and property regulation of nanocellulose • Explored critical enzyme concentration of nanocellulose morphology transition • Achieved the high-value utilization of waste paper [ABSTRACT FROM AUTHOR]

The cost of enzymolysis is a major bottleneck for the industrialisation of lignocellulosic enzymatic hydrolysis technology, and recycling cellulase can reduce this cost. Herein, a sulfobetaine prepolymer (CPS) with terminal chlorine was grafted onto enzymatic hydrolysis residual lignin (EHL) from corncob to construct thermosensitive lignin-based "molecular glues" (lignin-based sulfobetaine polymers, L-CPS) that were used to recover and recycle cellulase. L-CPS 2 (1.0 g/L) was added to the corncob residue (CCR) enzymolysis system (50 °C, pH 4.5). After hydrolysis, L-CPS 2 co-precipitated with cellulase through hydrophobic binding when cooling to 25 °C. This co-precipitation decreased the amount of cellulase by 40 %. In summary, a thermally responsive lignin-based molecular glue was constructed for green recycling of cellulase, providing a new approach to decreasing the cost of lignocellulosic enzymolysis and high value utilisation of industrial lignin. [Display omitted] [ABSTRACT FROM AUTHOR]

Castor stalk from hemp plants is an attractive lignocellulosic feedstock for biomass refining valorization due to its similar chemical composition to hardwoods. In this study, the castor stalk fibers were pretreated with efficient dual-functional ethanolamine to achieve delignification and swelling of the cellulosic fibers, followed by cellulase enzymatic digestion for biomass conversion. Experimental results showed that ethanolamine pretreatment at 160 °C for 1 h effectively removed 69.20 % of lignin and 43.18 % of hemicellulose. In addition to efficient delignification and removal of hemicellulose, the study also revealed that supramolecular structure of cellulose was another major factor affecting enzymatic hydrolysis performance. The lowered crystallinity (60–70 %) and swelled crystal sizes (2.95–3.04 nm) promoted enzymatic hydrolysis efficiency during the heterogeneous reaction process. Under optimal conditions (160 °C, 1 h; enzyme loading: 15 FPU/g substrate), promoted yields of 100 % glucose and over 90 % xylose were achieved, which were significantly higher than those obtained from untreated castor stalk. These findings highlighted the effectiveness of the dual-functional ethanolamine pretreatment strategy for efficient bioconversion of lignocellulosic feedstocks. Overall, this study provides valuable insights into the development of new strategies for the efficient utilization of biomass resources, which is essential for the sustainable development of our society. [Display omitted] [ABSTRACT FROM AUTHOR]

In this study, pectin extracted from pomelo peel was investigated using three different combination methods of pulsed electric field (PEF) and cellulase. Three action sequences were performed, including PEF treatment followed by enzymatic hydrolysis, enzymatic hydrolysis followed by PEF treatment, and enzymatic hydrolysis simultaneously treated by PEF. The three corresponding pectins were namely PEP, EPP and SP. The physiochemical, molecular structural and functional properties of the three pectins were determined. The results showed that PEP had excellent physiochemical properties, with the highest yield (12.08 %), total sugar (80.17 %) and total phenol content (38.20 %). The monosaccharide composition and FT-IR analysis indicated that the three pectins were similar. The molecular weights of PEP, EPP and SP were 51.13, 88.51 and 40.00 kDa, respectively. PEP showed the best gel properties, emulsification stability and antioxidant capacity among the three products, due to its high galacturonic acid and total phenol content, appropriate protein and low molecular weight. The mechanism of PEF-assisted cellulase hydrolysis of pomelo peel was also revealed by SEM analysis. These results suggested that PEF pretreatment was the best method, which not only improved the efficiency of enzymatic extraction, but also reduced resource waste and increased financial benefits. [ABSTRACT FROM AUTHOR]

Cellulases have been widely used in many fields such as animal feed, textile, food, lignocellulose bioconversion, etc. Efficient and low-cost production of cellulases is very important for its industrial application, especially in bioconversion of lignocellulosic biomass. Filamentous fungi are currently widely used in industrial cellulase production due to their ability to secrete large amounts of active free cellulases extracellularly. This review comprehensively summarized the research progress on cellulases from filamentous fungi in recent years, including filamentous fungi used for cellulase production and its modification strategies, enzyme compositions, characterization methods and application of fungal cellulase systems, and the production of fungal cellulase includes production processes, factors affecting cellulase production such as inducers, fermentation medium, process parameters and their control strategies. Also, the future perspectives and research topics in fungal cellulase production are presented in the end of the review. The review helps to deepen the understanding of the current status of fungal cellulases, thereby promoting the production technology progress and industrial application of filamentous fungal cellulase. [Display omitted] [ABSTRACT FROM AUTHOR]

This study investigates the synthesis of (hemi)cellulolytic enzymes, including endoglucanase (CMCase), xylanase, and β-glucosidase, employing Trichoderma reesei RUT-C30 and deoiled oil palm mesocarp fiber (OPMF) through solid-state fermentation (SSF). The objective was to determine the optimal process conditions for achieving high enzyme activities through a one-factor-at-a-time approach. The study primarily focused on the impact of the solid-to-liquid ratio, incubation period, initial pH, and temperature on enzyme activity. The effects of OPMF pretreatment, particularly deoiling and fortification, were explored. This approach significantly improved enzyme activity levels compared to the initial conditions, with CMCase increasing by 111.6 %, xylanase by 665.2 %, and β-Glucosidase by 1678.1 %. Xylanase and β-glucosidase activities, peaking at 1346.75 and 9.89 IU per gram dry substrate (GDS), respectively, under optimized conditions (1:4 ratio, pH 7.5, 20 °C, 9-day incubation). With lower moisture levels, CMCase reached its maximum activity of 227.84 IU/GDS. The study highlights how important it is for agro-industrial byproducts to support environmentally sustainable practices in the palm oil industry. It also emphasizes how differently each enzyme reacts to changes in process parameters. [ABSTRACT FROM AUTHOR]

In this work, five different NaOH-catalyzed ethylene glycol (EG) pretreatments together with water-soluble sulfonated lignin (SL) were used for enhancing sugarcane bagasse (SCB) enzymatic digestion. The results showed that the coupling of NaOH and EG into a one-pot pretreatment (10%NaOH/EG) was more beneficial to improve SCB enzymatic hydrolysis than that of single 10%NaOH or EG pretreatment, or the two-step pretreatment of NaOH and EG in different sequence (10%NaOH+EG and EG + 10%NaOH, respectively). The highest glucose yield of this work was 91.2 %, mainly released from the SCB that pretreated with 10%NaOH/EG at 130 °C for 60 min and 72 h enzymatic hydrolysis. The adding of SL into the enzymatic hydrolysis step could significantly lower the cellulase dosage and hydrolysis time from 20 FPU/g and 72 h to 10 FPU/g and 24 h, respectively, meanwhile keeping a high glucose yield of 90.4 %. The characterization of various pretreated or un-pretreated SCB confirmed that the improvement of hydrolysis efficiency of SCB after 10%NaOH/EG pretreatment was closely related to the removal of various components barriers in SCB and the fragmentation of pretreated solid. It can be concluded that the developed NaOH-catalyzed ethylene glycol pretreatment was an efficiency way to enhance the sugar release from SCB. [Display omitted] • Five different NaOH-catalyzed glycol pretreatments were developed in this work. • 10%NaOH/EG pretreatment displayed highest efficiency for enhancing SCB hydrolysis. • Sulfonated lignin could remarkably reduce the cellulase dosage and hydrolysis time. • A certain amount of glycol lignin (∼50.1 %) also can be obtained in pretreated liquid. [ABSTRACT FROM AUTHOR]

Drug conjugation with enzymes is one of the innovative antibacterial nanocarriers used as a delivery system for cancer therapy. Zeolitic imidazolate framework-8 (ZIF-8) was synthesized, dual encapsulated with cellulase (CL) enzyme, and resveratrol (Resv) drug formed ZIF-8@CL&Resv. Cellulase and resveratrol hydrophobic nature have bound them together and imparted a negative charge on the ZIF-8, resulting in the decrease of zeta potential from 22.7 mV (ZIF-8) to 3.82 mV (ZIF-8@CL&Resv). The cellulase like a scaffold regulated the pH-responsive release of resveratrol enhancing its bioavailability. Molecular docking studies provided evidence of the major interaction between the biofilm-related proteins with cellulase and resveratrol. The encapsulated cellulase showed high enzymatic activity and possibly exhibited antibacterial effects by dissolving the biofilm and exposing bacteria to resveratrol action. Resveratrol released sustainably exhibited significant antioxidant and antibacterial activity against selected bacterial species. ZIF-8@CL&Resv exhibited high biocompatibility and had a potent cytotoxic effect against triple-negative breast cancer cells MDA MB 231 with an IC 50 -value of 17.18 μg/mL compared to ZIF-8 control with 90.47 μg/mL. ZIF-8@CL&Resv treatment led to 61.81 % cell death, apoptosis induction, increased ROS generation, and decreased mitochondrial membrane potential. Overall, data demonstrated that ZIF-8@CL&Resv is a novel drug release system and a potential catalytic nanoparticle for antimicrobial and anticancer applications., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Cellulases are divided into endoglucanase, exoglucanase, and β-glucosidase based on their catalytic activity. Eight cellulases were recombinantly expressed in Escherichia coli BL21 to investigate their effects on the enzymatic hydrolysis of wheat straw. Among them, cellulase 2006 exhibited the highest endoglucanase activity (432.25 U/mg), while Bf1 displayed superior exoglucanase and β-glucosidase activities (577.46 and 1991.63 U/mg respectively). Bioinformatic and enzymatic analyses revealed that both cellulases displayed notable thermal and pH stability. The enzyme kinetics parameters revealed that Km values for cellulases 2006 and Bf1 were 15.98 and 14.19 mg/mL, respectively, with Vmax values of 20.78 and 16.38 μmol/min/mg. In a prokaryotic co-expression system, the mixed cellulase Bf2006 exhibited endoglucanase, exoglucanase, and β-glucosidase activities (130.78, 1406.36, and 1119.25 U/mg, respectively). The enzymatic hydrolysis assay revealed that these three cellulases acted on the cellulose macromolecules in wheat straw, increasing reducing sugar content and decreasing crystallinity. Endoglucanase 2006 acted on various organic compounds rich in phenols and aromatic heterocycles, while Bf1 primarily acted on compounds containing glucose units. Bf2006 significantly affected the content of lignin, neutral detergent fiber, acid detergent fiber, and the microstructure of wheat straw, with degradation products primarily consisting of disaccharides, oligosaccharides, polysaccharides, glycosides, and other carbohydrates. This study provides theoretical guidance for the production and application of mixed cellulase Bf2006, serving as a reference for the industrial use of different cellulase types., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Microbial cellulases have become significant biocatalysts because of their complex composition and extensive industrial applications. This study aimed to isolate an efficient cellulase-producing strain, followed by molecular identification, enzyme purification, and characterization. Among 110 isolates, Bacillus amyloliquefaciens OKB3 was selected for its significant cellulase production, with optimal activity at pH 5.0 and 34 °C. The purification using ammonium sulfate and Sephadex G-100 chromatography resulted in specific activity of 2720.76 U/mg, 2.91 fold purification, and 29.44 % yield. The purified cellulase named CelB was a dimeric macromolecule of 123 kDa consisting of 67 and 54 kDa subunits. CelB was most active at 60 °C and pH 6, and it was stable at pH 5.5 to 6.0 and 0 °C to 4 °C. CelB was unaffected by metal cofactors and inhibited in the presence of divalent cations Cu 2+ , Hg 2+ , Cd 2+ , and Ag 2+ . The CelB has higher specificity of CMC compared to other substrates. The K m , V max, and K cat values were 0.037 mM, 188.67 μmole/min, and 7430 S -1 respectively. The unique attributes of CelB make it a very promising candidate for various biotechnological applications., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Hyperbranched polysaccharide from Pleurotus tuber-regium (PTR-HBPS) is a β-glucan with high degree of branching (DB, 0.69) and a molecular weight (Mw) of 31.2 × 105 g/mol with mixed β-1, 4/β-1, 4, 6/β-1, 6 glucosidic linkages. PTR-HBPS was depolymerized by cellulase and β-glucosidase under optimized conditions to form PC (PTR-HBPS depolymerized by cellulase) and PG (PTR-HBPS depolymerized by β-glucosidase) fractions with a minimum Mw of 2.74 × 105 and 3.98 × 105 g/mol, respectively. PC fractions had no significant changes for its primary structure in terms of glycosidic linkages, DB, and triple helical structure, while the DB of PG fractions was reduced to 0.63 with the loss of triple helical structure. Nanoparticles fabricated by PC fractions with zein showed better stability under different pH conditions. Enzymatic depolymerized low Mw β-glucan derived from PTR-HBPS with similar structural characteristics as the native one has potential as nanocarriers for food bioactive substances. • Cellulase could depolymerize Mw of PTR-HBPS up to one-eleventh of its original size. • Cellulase cleaved β-1,4 glucosidic bonds in backbone but did not change configuration. • β-Glucosidase could degrade Mw of PTR-HBPS up to one-eighth of its original size. • β-Glucosidase cuts more side branches from non-reducing end caused a decrease of DB. • PC3/zein nanoparticles have better pH stability. [ABSTRACT FROM AUTHOR]

In this research, a magnetic reusable nickel nanoparticle (NiNPs) supporting materials were prepared for cellulase enzyme immobilization. The immobilized cellulase showed high activity recovery, large & fast immobilization capacity and improved pH & temperature tolerance. The excellent stability and reusability enabled the immobilized cellulase to retain 84% of its initial activity after ten cycles. At 2 mg/mL enzyme concentration, highest 93% immobilization efficiency was achieved within two hours of immobilization. When the treatment temperature reached 40 °C and pH 5, the immobilized cellulase exhibited highest residual activity. The immobilized cellulase could be separated from the solution by a magnetic force. This study introduced a novel supporting material for cellulase immobilization, and the immobilized cellulase poses a great potential in the hydrolysis of lignocellulosic biomass which can used as an easily applicable and sustainable pre-treatment step for advanced biofuel production. [Display omitted] [ABSTRACT FROM AUTHOR]

Lignin-cellulose nanofibrils (LCNF) are of attracting an increasing interest due to the benefits of maintaining the lignin in the nanomaterial composition. The production of LCNF requires considerable energy consumption, which has been suppressed employing pretreatment of biomass, in which it highlights those that employ enzymes that have the advantage of being more environmentally friendly. Some negative aspects of the presence of lignin in the fiber to obtain cellulose nanofibrils is that it can hinder the delamination of the cell wall and act as a physical barrier to the action of cellulase enzymes. This study aimed to evaluate the impact of a combined enzymatic pretreatment of laccase and endoglucanase for high content lignin LCNF production. The morphological and chemical properties, visual aspect and stability, crystallinity, mechanical properties, rheology, barrier properties and quality index were used to characterize the LCNF. The laccase loading used was efficient in modifying the lignin to facilitate the action of the endoglucanase on cellulose without causing the removal of this macromolecule. This pretreatment improved the quality of LCNF (61 ± 3 to 71 ± 2 points) with an energy saving of 42% and, therefore, this pretreatment could be suitable for industrial production for a variety of applications. [Display omitted] [ABSTRACT FROM AUTHOR]

Immobilized enzymes are one of the most common tools used in enzyme engineering, as they can substantially reduce the cost of enzyme isolation and use. However, efficient catalysis of solid substrates using immobilized enzymes is challenging, hydrolysis of insoluble cellulose by immobilized cellulases is a typical example of this problem. In this study, inspired by bees and honeycombs, we prepared gelatin-modified cellulase (BEE) and gelatin hydrogels (HONEYCOMB) to achieve reversible recycling versus release of cellulase through temperature-responsive changes in the triple-stranded helix-like interactions between BEE and HONEYCOMB. At elevated temperatures, BEE was released from HONEYCOMB and participated in hydrolytic saccharification. After 24 h, the glucose yields of both the free enzyme and BEE reached the same level. When the temperature was decreased, BEE recombined with HONEYCOMB to facilitate the effective separation and recycling of BEE from the system. The enzymatic system retained >70 % activity after four reuse cycles. In addition, this system showed good biocompatibility and environmental safety. This method increases the mass transfer capacity and enables easy recovery of immobilized cellulase, thereby serving as a valuable strategy for the immobilization of other enzymes. [Display omitted] [ABSTRACT FROM AUTHOR]

Gums are high-molecular-weight compounds with hydrophobic or hydrophilic characteristics, which are mainly comprised of complex carbohydrates called polysaccharides, often associated with proteins and minerals. Various innovative modification techniques are utilized, including ultrasound-assisted and microwave-assisted techniques, enzymatic alterations, electrospinning, irradiation, and amalgamation process. These methods advance the process, reducing processing times and energy consumption while maintaining the quality of the modified gums. Enzymes like xanthan lyases, xanthanase, and cellulase can selectively modify exudate gums, altering their structure to enhance their properties. This precise enzymatic approach allows for the use of exudate gums for specific applications. Exudate gums have been employed in nanotechnology applications through techniques like electrospinning. This enables the production of nanoparticles and nanofibers with improved properties, making them suitable for the drug delivery system, tissue engineering, active and intelligient food packaging. The resulting modified exudate gums exhibit improved rheological, emulsifying, gelling, and other functional properties, which expand their potential applications. This paper discusses novel applications of these modified gums in the pharmaceutical, food, and industrial sectors. The ever-evolving field presents diverse opportunities for sustainable innovation across these sectors. [Display omitted] • Exudate gums exhibit both hydrophobic and hydrophilic characteristics, emphasizing their versatile nature. • The study highlights a range of cutting-edge modification techniques aimed at enhancing the functional properties. • Precise enzymatic approach tailors the structure of the gums to enhance their properties for specific applications. • The modifications impart improved rheological, emulsifying, and gelling properties to the exudate gums. [ABSTRACT FROM AUTHOR]

Anthropogenic activities have led to a drastic shift from natural fuels to alternative renewable energy reserves that demand heat-stable cellulases. Cellobiohydrolase is an indispensable member of cellulases that play a critical role in the degradation of cellulosic biomass. This article details the process of cloning the cellobiohydrolase gene from the thermophilic bacterium Caldicellulosiruptor bescii and expressing it in Escherichia coli (BL21) CondonPlus DE3-(RIPL) using the pET-21a(+) expression vector. Multi-alignments and structural modeling studies reveal that recombinant CbCBH contained a conserved cellulose binding domain III. The enzyme's catalytic site included Asp-372 and Glu-620, which are either involved in substrate or metal binding. The purified CbCBH, with a molecular weight of 91.8 kDa, displayed peak activity against p NPC (167.93 U/mg) at 65°C and pH 6.0. Moreover, it demonstrated remarkable stability across a broad temperature range (60–80°C) for 8 h. Additionally, the Plackett-Burman experimental model was employed to assess the saccharification of pretreated sugarcane bagasse with CbCBH, aiming to evaluate the cultivation conditions. The optimized parameters, including a pH of 6.0, a temperature of 55°C, a 24-hour incubation period, a substrate concentration of 1.5% (w / v), and enzyme activity of 120 U, resulted in an observed saccharification efficiency of 28.45%. This discovery indicates that the recombinant CbCBH holds promising potential for biofuel sector. • The increasing energy demand has intensified the need of renewable energy sources. • A novel cellulose-binding domain containing cellobiohydrolase (CbCBH) cloned from Caldicellulosiruptor bescii. • CbCBH exhibited great thermostability making it a promising candidate for biofuel. • Plackett-Burman model evaluated the saccharification conditions of sugarcane bagasse. • Optimized conditions yielded a remarkable 28.45% saccharification efficiency. [ABSTRACT FROM AUTHOR]

In recent years, the process of producing bioethanol from lignocellulosic biomass through biorefining has become increasingly important. However, to obtain a high yield of ethanol, the complex structures in the feedstock must be broken down into simple sugars. A cost-effective and innovative method for achieving this is ionic liquid pre-treatment, which is widely used to efficiently hydrolyze the lignocellulosic material. The study aims to produce a significant profusion of bioethanol via catalytic hydrolysis of ionic liquid-treated lignocellulose biomass. The current study reports the purification of Streptomyces sp. MS2A cellulase via ultrafiltration and gel permeation chromatography. The kinetic parameters and the biochemical nature of the purified cellulase were analyzed for the effective breakdown of the EMIM[OAC] treated lignocellulose chain. The two-step cellulase purification resulted in 6.28 and 12.44 purification folds. The purified cellulase shows a Km value of 0.82 ± 0.21 mM, and a Vmax value of 85.59 ± 8.87 μmol min−1 mg−1 with the catalytic efficiency of 1.027 S−1. The thermodynamic parameters like ΔH, ΔS, and ΔG of the system were studied along with the thermal deactivation kinetics of cellulase. The optimal temperature and pH of the purified cellulase enzyme for hydrolysis was found to be 40 °C and 7. The rice husk and wheat husk used in this study were pretreated with the EMIM [OAC] ionic liquid and the change in the structure of lignocellulosic biomass was observed via HRSEM. The ionic liquid treated biomass showed the highest catalytic hydrolysis yield of 106.66 ± 0.19 mol/ml on the third day. The obtained glucose was fermented with Saccharomyces cerevisiae to yield 23.43 g of ethanol/l of glucose from the rice husk (RH) and 24.28 g of ethanol/l of glucose from the wheat husk (WH). • Streptomyces sp.MS2A cellulase was 12.4 fold purified by ultrafiltration and gel permeation chromatography. • EMIM[OAC] pretreatment reduced the crystallinity index of lignocellulose biomass. • 24.28 g/L and 23.43 g/L of ethanol was produced from the fermentation of ionic liquid pretreated wheat husk and ricehusk. [ABSTRACT FROM AUTHOR]

The recently developed technologies for immobilization of cellulase may address the challenges in costly hydrolysis of cellulose for cellulosic butanol production. In this study, a "hybrid" hydrolysis was developed based on chemical hydrolysis of cellulose to its oligomers followed by enzymatic post-hydrolysis of the resulting "soluble oligomers" by cellulase immobilized on chitosan-coated Fe 3 O 4 nanoparticles. This hybrid hydrolysis stage was utilized in the process of biobutanol production from a waste textile, jeans waste, leading to selective formation of glucose and high yield of butanol production by Clostridium acetobutylicum. After validating the immobilization process, the optimum immobilization parameters including enzyme concentration and time were achieved on 8 h and 15.0 mg/mL, respectively. The reusability of immobilized enzyme showed that immobilized cellulase could retain 51.5% of its initial activity after three times reuses. Dilute acid hydrolysis of regenerated cellulose at 120–180 °C for 60 min 0.5–1.0% phosphoric acid led to less than 10 g/L glucose production, and enzymatic post-hydrolysis of the oligomers resulted in up to 51.5 g/L glucose. Fermentation of the hydrolysate was accompanied by 5.3 g/L acetone-butanol-ethanol (ABE) production. The simultaneous co-saccharification and fermentation (SCSF) of soluble and insoluble oligomers of cellulose led to 17.4 g/L ABE production. [Display omitted] • Post-hydrolysis of chemically derived cellulose oligomers by immobilized cellulase. • Cellulase was immobilized on chitosan-grafted magnetic nanoparticles. • The hybrid hydrolysis led to more than 50 g/L glucose fermented to 5.3 g/L ABE. • The immobilized enzyme was reused up to three cycles with near half activity. • Simultaneous co-saccharification and fermentation led to 17.4 g/L ABE production. [ABSTRACT FROM AUTHOR]

Cellulose nanostructures obtained from lignocellulosic biomass by the enzymatic route can offer advantages in terms of material properties and processing sustainability. However, most of the enzymatic cocktails commonly used in the saccharification of biomass are designed to promote the complete depolymerization of the cellulose structure into soluble sugars. Here, investigation was made of the way that the action of different commercially available cellulase enzyme cocktails can affect the production of nanocellulose. For this, enzymatic cocktails designed for complete or partial saccharification were compared, using eucalyptus cellulose pulp as a model feedstock. The results showed that all the enzymatic cocktails were effective in the formation of nanocellulose structures, with the complete saccharification enzymes being more efficient in promoting the coproduction of glucose (36.5 g/L, 87% cellulose conversion). The presence of auxiliary enzymes, especially xylanases, acted cooperatively to favor the production of nanostructures with higher crystallinity (up to 79%), higher surface charge (zeta potential up to −30.9 mV), and more uniform dimensions within the size range of cellulose nanocrystals (80 to 350 nm). Interestingly, for the enzymatic cocktails designed for partial saccharification, the xylanase activity was more important than the endoglucanase activity in the production of nanocellulose with improved properties. The findings showed that the composition of the enzymatic cocktails already used for complete biomass saccharification can be suitable for obtaining nanocellulose, together with the release of a glucose stream, in a format compatible with the biorefinery concept. • Cellulose nanostructures using enzymes can offer advantages in their properties. • The action of different enzyme cocktails affects nanocellulose production. • Xylanase was more important than endoglucanase in terms of nanocellulose properties. • Enzyme cocktails for complete saccharification are suitable to obtain nanocellulose. [ABSTRACT FROM AUTHOR]

An integrated treatment coupling alkali, steam explosion and ammonia/chlorine-free bleaching with sequential mild acid pretreatment were performed to isolate and characterize cellulose from banana agrowastes followed by optimized enzymatic hydrolysis to glucose. The cellulose yield, compositional, microstructural, and morphological analysis initially obtained from three post-harvest banana agrowastes (peel, pseudostem, and peduncle) were surveyed. Isolation parameters for banana peduncle agrowastes, the most efficient precursor, were reconfigured for acid hydrolysis by applying an orthogonal L 9 array of Taguchi design. Effects of solution-to-pulp ratio, acid concentration, temperature, and reaction time on physicochemical parameters were assessed resulting in ~81% cellulose recovery. Subsequently, cellulase driven enzymatic conversion to glucose was modelled using response surface methodology (RSM), where the mutual influences of incubation time, enzyme concentration, substrate concentration, and surfactant concentration were investigated. Artificial Neural Network (ANN) modelling further improved upon RSM optimizations ensuing ~ 97% optimized glucose yield, verified experimentally. • Cellulose extraction coupled with glucose production in an integrated scheme. • Ammonia/chlorine free, low-acid hydrolytic isolation of cellulose is environmentally benign. • Taguchi design optimized low-acid hydrolysis yields ~81% cellulose. • Low DP celluloses enzymatically converted to high yield glucose (~97%). • RSM and ANN modelling used for optimizing enhanced glucose production. [ABSTRACT FROM AUTHOR]

Endoglucanases are carbohydrate-degrading enzymes widely used for bioethanol production as part of the enzymatic cocktail. However, family 5 subfamily 5 (GH5_5) endoglucanases are still poorly explored in depth. The Trichoderma reesei representative is the most studied enzyme, presenting catalytic activity in acidic media and mild temperature conditions. Though biochemically similar, its modular structure and synergy with other components vary greatly compared to other GH5_5 members and there is still a lack of specific studies regarding their interaction with other cellulases and application on novel and better mixtures. In this regard, the threedimensional structure elucidation is a highly valuable tool to both uncover basic catalytic mechanisms and implement engineering techniques, proved by the high success rate GH5_5 endoglucanases show. GH5_5 enzymes must be carefully evaluated to fully uncover their potential in biomass-degrading cocktails: the optimal industrial conditions, synergy with other cellulases, structural studies, and enzyme engineering approaches. We aimed to provide the current understanding of these main topics, collecting all available information about characterized GH5_5 endoglucanases function, structure, and bench experiments, in order to suggest future directions to a better application of these enzymes in the industry. [ABSTRACT FROM AUTHOR]

Exoglucanase (CBH) is the rate limiting enzyme in the process of cellulose degradation. The carbohydrate binding module (CBM) can improve the accessibility of cellulase to substrate, thereby promoting the enzymatic hydrolysis of cellulase. In this study, the influence of CBM on the properties of GH6 exoglucanase from Chaetomium thermophilum (CtCBH) is systematically explored from three perspectives: the fusion of exogenous CBM, the exogenous CBM replacement of its own CBM, and the deletion of its own CBM. The parental and reconstructed CtCBH presented the same optimum pH (6.0) and temperature (60 °C) for maximum activity. Fusion of exogenous CBM increased the binding capacity of CtCBH to Avicel by 8% and 9%, respectively, but it had no significant effect on its catalytic activity. The exogenous CBM replacement of its own CBM resulted in a 12% reduction in the binding ability of CtCBH to Avicel, and a 26% reduction in the catalytic activity of Avicel. The deletion of its own CBM significantly reduced the binding ability of CtCBH to Avicel by approximately 53%, but its catalytic activity was not obviously reduced. These observations suggest that binding ability of CBM is not necessary for the catalysis of CtCBH. • CBM is not necessary for the catalysis of CtCBH. • The deletion of CBM significantly reduced the binding ability of CtCBH. • The deletion of CBM did not significantly reduce its catalytic activity. • The fusion of exogenous CBM can improve the binding ability of CtCBH. • The replacement of CBM affected the catalytic activity and binding ability of CtCBH. [ABSTRACT FROM AUTHOR]

Cellulases play a promising role in the bioconversion of renewable lignocellulosic biomass into fermentable sugars which are subsequently fermented to biofuels and other value-added chemicals. Besides biofuel industries, they are also in huge demand in textile, detergent, and paper and pulp industries. Low titres of cellulase production and processing are the main issues that contribute to high enzyme cost. The success of ethanol-based biorefinery depends on high production titres and the catalytic efficiency of cellulases functional at elevated temperatures with acid/alkali tolerance and the low cost. In view of their wider application in various industrial processes, stable cellulases that are active at elevated temperatures in the acidic-alkaline pH ranges, and organic solvents and salt tolerance would be useful. This review provides a recent update on the advances made in thermostable cellulases. Developments in their sources, characteristics and mechanisms are updated. Various methods such as rational design, directed evolution, synthetic & system biology and immobilization techniques adopted in evolving cellulases with ameliorated thermostability and characteristics are also discussed. The wide range of applications of thermostable cellulases in various industrial sectors is described. • This review focuses on recent attempts made in evolving cellulases with ameliorated thermostability characteristics. • Several thermostable microbial cellulases, their sources and characteristics are discussed. • Various mechanisms and adaptations that contribute to thermostability of cellulases are also described. • The industrial applications of thermostable cellulases are comprehensively covered. [ABSTRACT FROM AUTHOR]

Nanostructured materials along with an added value of polymers-based support carriers have gained high interest and considered ideal for enzyme immobilization. The recently emerged nanoscience interface in the form of nanostructured materials combined with immobilized-enzyme-based bio-catalysis has now become research and development frontiers in advance and applied bio-catalysis engineering. With the involvement of nanoscience, various polymers have been thoroughly developed and exploited to nanostructured engineer constructs as ideal support carriers/matrices. Such nanotechnologically engineered support carriers/matrix possesses unique structural, physicochemical, and functional attributes which equilibrate principal factors and strengthen the biocatalysts efficacy for multipurpose applications. In addition, nano-supported catalysts are potential alternatives that can outstrip several limitations of conventional biocatalysts, such as reduced catalytic efficacy and turnover, low mass transfer efficiency, instability during the reaction, and most importantly, partial, or complete inhibition/deactivation. In this context, engineering robust and highly efficient biocatalysts is an industrially relevant prerequisite. This review comprehensively covered various biopolymers and nanostructured materials, including silica, hybrid nanoflower, nanotubes or nanofibers, nanomembranes, graphene oxide nanoparticles, metal-oxide frameworks, and magnetic nanoparticles as robust matrices for cellulase immobilization. The work is further enriched by spotlighting applied and industrially relevant considerations of nano-immobilized cellulases. For instance, owing to the cellulose-deconstruction features of nano-immobilized cellulases, the applications like lignocellulosic biomass conversion into industrially useful products or biofuels, improved paper sheet density and pulp beat in paper and pulp industry, fruit juice clarification in food industry are evident examples of cellulases, thereof are discussed in this work. [Display omitted] • Herein, nano-biocatalytic systems are comprehensively discussed with potential applications. • Immobilized-cellulase based bio-catalysis has become research and development frontiers. • Various applied perspectives are given with suitable examples. • Lignocellulosic biomass conversion into industrially useful products or biofuels • Fruit juice clarification in food industry is evident examples and discussed herein. [ABSTRACT FROM AUTHOR]

Nanocellulose isolation from lignocellulose is a tedious and expensive process with high energy and harsh chemical requirements, primarily due to the recalcitrance of the substrate, which otherwise would have been cost-effective due to its abundance. Replacing the chemical steps with biocatalytic processes offers opportunities to solve this bottleneck to a certain extent due to the enzymes substrate specificity and mild reaction chemistry. In this work, we demonstrate the isolation of sulphate-free nanocellulose from organosolv pretreated birch biomass using different glycosyl-hydrolases, along with accessory oxidative enzymes including a lytic polysaccharide monooxygenase (LPMO). The suggested process produced colloidal nanocellulose suspensions (ζ-potential −19.4 mV) with particles of 7–20 nm diameter, high carboxylate content and improved thermostability (T o = 301 °C, T max = 337 °C). Nanocelluloses were subjected to post-modification using LPMOs of different regioselectivity. The sample from chemical route was the least favorable for LPMO to enhance the carboxylate content, while that from the C1-specific LPMO treatment showed the highest increase in carboxylate content. • Nanocellulose isolated by chemical and enzymatic route from birch was compared. • Enzymes with endoglucanase, xylanase, laccase and LPMO activity were employed. • Enzymatically produced nanocellulose had high COOH content and thermostability. • C1-LPMO post-treatment yielded high aspect ratio fibers with higher COOH content. [ABSTRACT FROM AUTHOR]

Cellulose is a very abundant polymer that is found in nature. Cellulose has been used as a raw material for production of biofuels for many years. However, there are multiple processing steps that are required so that cellulose can be used as a raw material for biofuel production. One of the most important steps is the breakdown of cellulose into intermediate sugars which can then be a viable substrate for biofuel production. Cellulases are enzymes which play a role in the catalysis of the breakdown of cellulose into glucose. Nanomaterials and micromaterials have been gaining a lot of attention over the past few years for its potential in immobilizing enzymes for industrial procedures. Immobilization of enzymes on these nanomaterials has been observed to be of great value due to the improvement in thermal stability, pH stability, regenerative capacity, increase in activity and the reusability of enzymes. Similarly, there have been multiple reports of cellulase enzymes being immobilized on various nanoparticles. The immobilization of these cellulase enzymes have resulted in very efficient processing and provide a great and economic solution for the processing of cellulose for biofuel production. Hence in this paper, we review and discuss the various advantages and disadvantages of enzymes on various available nanomaterials. • Immobilization of cellulase onto MNPs makes the recovery process easier. • Cellulase immobilized on composite materials has high loading capacity and great reusability. • CNTs are suitable for enzyme immobilization especially for the purpose of biofuel production. • Immobilization on MOF was observed to be an excellent strategy for the improvement in enzyme activity. [ABSTRACT FROM AUTHOR]

A study was carried out to investigate the characterization of a novel Aspergillus sulphureus JCM01963 xylanase (AS-xyn10A) with a carbohydrate binding module (CBM) and its application in degrading alkali pretreated corncob, rapeseed meal and corn stover alone and in combination with a commercial cellulase. In this study, the 3D structure of AS-xyn10A, which contained a CBM at C-terminal. AS-xyn10A and its CBM-truncated variant (AS-xyn10A-dC) was codon-optimized and over-expressed in Komagaella phaffii X-33 (syn. Pichia pastoris) and characterized with optimal condition at 70 °C and pH 5.0, respectively. AS-xyn10A displayed high activity to xylan extracted from corn stover, corncob, and rapeseed meal. The concentration of hydrolyzed xylo-oligosaccharides (XOSs) reached 1592.26 μg/mL, 1149.92 μg/mL, and 621.86 μg/mL, respectively. Xylobiose was the main product (~70%) in the hydrolysis mixture. AS-xyn10A significantly synergized with cellulase to improve the hydrolysis efficiency of corn stover, corncob, and rapeseed meal to glucose. The degree of synergy (DS) was 1.32, 1.31, and 1.30, respectively. Simultaneously, XOSs hydrolyzed with AS-xyn10A and cellulase was improved by 46.48%, 66.13% and 141.45%, respectively. In addition, CBM variant decreased the yields of xylo-oligosaccharide and glucose in rapeseed meal degradation. This study provided a novel GH10 endo -xylanase, which has potential applications in hydrolysis of biomass. • The optimal condition for full-length and truncated AS-xyn10A is 70 °C and pH 5.0. • The removal of CBM had great influence on the synergistic action. • Xylobiose was the main product of AS-xyn10A degradation of alkali-pretreated corncob, rapeseed meal and corn stover. • AS-xyn10A showed synergistic effect with cellulase on biomasses hydrolyzing into glucose. [ABSTRACT FROM AUTHOR]

The investigation aims to determine the effect of enzymatic and alkali treatments on Sambucus ebulus L. stem fiber. For this purpose, Sambucus ebulus L. stem fibers were treated with alkali, cellulase, and pectinase enzymes. An image processing technique was developed and implemented to calculate the average thicknesses of Sambucus ebulus L. fibers. The thickness of alkali, cellulase and pectinase enzyme treated fibers was determined as 478.62 μm, 808.28 μm and 478.20 μm, respectively. Scanning electron microscopy analysis illustrated that enzymatic and alkali treatments lead to the breakage of fiber structure. Furthermore, enzymatic and alkali treatments induce variations in elemental ingredients. All treatments increased the crystallinity index of Sambucus ebulus L. fiber from 72 % (raw fiber) to 83 % (alkali treated), 75.2 % (cellulase enzyme treated) and 86.3 % (pectinase enzyme treated) due to the hydrolysis of hemicellulose. Fourier transform infrared analysis indicated that there are no significant differences in functional groups. Thermogravimetric analysis shows that enzymatic and alkali treatments improve final degradation temperature of the fiber. Mechanical behaviors of cellulase enzyme-treated fiber decrease compared to raw fiber, while pectinase enzyme and alkali treatment cause to improve mechanical properties. Tensile strength of samples was determined as 76.4 MPa (cellulase enzyme treated fiber), 210 MPa (pectinase enzyme treated fiber) and 240 MPa (alkali treated fiber). Young's modules of cellulase enzyme, pectinase enzyme and alkali treated fibers were predicted as 5.5 GPa, 13.1 GPa and 16.6 GPa. Elongation at break of samples was calculated as 5.5 % (cellulase enzyme treated fiber), 6.5 % (pectinase enzyme treated fiber) and 6 % (alkali treated fiber). The results suggest that enzymatic and alkali treatments can modify the functional and structural attributes of Sambucus ebulus L. fiber. [Display omitted] [ABSTRACT FROM AUTHOR]

Glycosylation, a general post-translational modification for fungal cellulase, has been shown to affect cellulase binding to its substrate. However, the exact impact of glycosylation on cellulase-lignin interaction remain unclear. Here, we demonstrated that the lignin isolated from tetrahydrofuran-pretreated corn stover exhibits strong adsorption capability to cellulase due to its negatively charged and porous structure. For the cellulases with varying glycosylation levels, the less-glycosylated protein showed high adsorption capability to lignin, and that trend was observed for the main cellulase components secreted by Penicillium oxilicum , including endoglucanase PoCel5B, cellobiohydrolase PoCel7A-2, and β-glucosidase PoBgl1. Additionally, N-glycan sites and motifs were examined using mass spectrometry, and protein structures with N-glycans were constructed, where PoBgl1 and PoCel7A-2 contained 13 and 1 glycosylated sites respectively. The results of molecular dynamics simulations indicated that the N-glycans impacted on the solvent-accessible surface area and secondary structure of protein, and the binding conformation of lignin fragment on cellulase, resulting in a decrease in binding energy (14 kcal/mol for PoBgl1 and 13 kcal/mol for PoCel7A-2), particularly for van der Waals and electrostatic interaction. Those findings suggested that glycosylation negatively impacted the lignin-cellulase interaction, providing a theoretical basis for the rational engineering of enzymes to reduce lignin-enzyme interaction. [ABSTRACT FROM AUTHOR]

Enhancing enzyme activity and stability in biomass degradation can improve substrate saccharification and, increases biorefinery efficiency. For the first time, we identified 20 lytic polysaccharide monooxygenases (LPMOs) AA9 genes in the genome of Thermothelomyces fergusii. Our results showed that TfAA9 was categorized into LPMOs1, LPMOs2, and LPMOs3 subgroups based on protein diversity. Protein- 3D structure analysis showed strong interactions between Myceliophthora thermophila AA9 proteins and 17 TfAA9 proteins. Gene ontology analysis indicated a high enrichment of cellulase activity in TfAA9 genes. KEGG pathways analysis revealed the role of TfAA9 proteins in the endohydrolysis of 1,4-beta-D-glucosidic linkages in cellulose. Numerous TfAA9s gene transcripts were up-regulated on avicel, cellobiose, and glucose, with a higher proportion on avicel. Protein concentration, endoglucanase, and cellulase activity were also boosted on avicel. However, limited fungal biomass was observed on avicel, despite the abundance of AA9 LPMOs in the T. fergusii genome. These findings expand our understanding of fungal AA9 genes and their role in lignocellulolytic degradation. The disparity between biomass and enzymatic activity suggests screening TfAA9 genes for highly active enzymes and redundant genes via heterologous expression. In short, functional characterization of these genes could contribute to improving the saccharification process of industrial raw materials. • Genomic analysis identified 20 LPMOs AA9 genes in Thermothelomyces fergusii. • Expression of LPMOs AA9 gene shows variation against carbon source media. • Protein 3D analysis showed strong interactions between MtAA9 and TfAA9 proteins. • GO analysis indicated a high enrichment of cellulase activity in TfAA9 genes. • Protein concentration, endoglucanase, and cellulase activity were boosted on avicel. [ABSTRACT FROM AUTHOR]

The natural interactions between various bacteria, fungi, and other cellulolytic microorganisms destroy lignocellulosic polymers. The efficacy of this process is determined by the combined action of three main enzymes: endoglucanases, exo -glucanases, and β-glucosidase. The enzyme attacks the polymeric structure's β-1,4-linkages during the cellulose breakdown reaction. This mechanism is crucial for the environment as it recycles cellulose in the biosphere. However, there are problems with enzymatic cellulose breakdown, including complex cellulase structure, insufficient degradation efficacy, high production costs, and post-translational alterations, many of which are closely related to certain unidentified cellulase properties. These issues impede the practical use of cellulases. A developing area of research is the application of this similar paradigm for industrial objectives. Cellulase enzyme exhibits greater promise in many critical industries, including biofuel manufacture, textile smoothing and finishing, paper and pulp manufacturing, and farming. However, the study on cellulolytic enzymes must move forward in various directions, including increasing the activity of cellulase as well as designing peptides to give biocatalysts their desired attributes. This manuscript includes an overview of current research on different sources of cellulases, their production, and biochemical characterization. [ABSTRACT FROM AUTHOR]

Microbial diversity from indigenous cultures has the potential to accelerate lignocellulose degradation through enzymes and make composting economically feasible. Therefore, this study is designed to boost cellulase output from a bacterial strain obtained from soil using a one-variable-at-a-time approach and response surface methodology. The bacteria recognized as Bacillus tequilensis (ON754229) produced the maximum cellulase at a temperature of 37 °C, pH -7.0, and incubation time of 72 h. A major contribution was anticipated by glucose (17 %) and ammonium sulfate (11 %) with cellulase activity of 0.56 U/mL in the optimized medium. The enzyme possessed activity of CMCase, FPase, and amylase of 0.589 μmol/min, 1.22 μmol/min, and 0.92 μmol/min respectively. SDS-PAGE showed a 65 kDa molecular weight of the enzyme capable of degrading cellulose, as confirmed by zymogram analysis. The enzyme showed relatively moderate thermo-stability towards neutral pH conditions possessing optimum conditions at pH 6.5 and temperature of 50 °C. The Km and Vmax values were 11.44 mM and 0.643 μmol/min respectively. The presence of MgSO 4 , ZnSO 4 , and Triton X- 100 increased the enzymatic reaction however AgNO 3 , EDTA, and HgCl 2 altered the activation process. These results showed cellulase from B. tequilensis SB125 would be suitable for conventional industrial processes that convert biomass into biofuels. [Display omitted] • A potential cellulase-producing bacteria, Bacillus tequilensis , was isolated. • Application of RSM technology significantly improved cellulase production by the bacteria. • The cellulase enzyme was confirmed for its broader substrate specificity. • Enzyme characterization validated its industrial candidacy. [ABSTRACT FROM AUTHOR]

Filamentous fungi are the main industrial source of cellulases which are important in the process of converting cellulose to fermentable sugars. In this study, transcriptome analysis was conducted on Aspergillus terreus NEAU-7 cultivated using corn stover and glucose as carbon sources. Four putative endoglucanases (EG5A, EG7A, EG12A, and EG12C) from A. terreus NEAU-7 were efficiently expressed in Pichia pastoris. Among them, EG7A exhibited the highest enzyme activity (75.17 U/mg) with an optimal temperature of 40 °C and pH 5.0. EG5A and EG12A displayed specific activities of 19.92 U/mg and 14.62 U/mg, respectively, at 50 °C. EG12C showed acidophilic characteristics with an optimal pH of 3.0 and a specific activity of 12.21 U/mg at 40 °C. With CMC-Na as the substrate, the Km value of EG5A, EG7A, EG12A or, EG12C was, 11.08 ± 0.87 mg/mL, 6.82 ± 0.74 mg/mL, 7.26 ± 0.64 mg/mL, and 9.88 ± 0.86 mg/mL, with V max values of 1258.23 ± 51.62 μmol∙min−1∙mg−1, 842.65 ± 41.53 μmol∙min−1∙mg−1, 499.38 ± 20.42 μmol∙min−1∙mg−1, and 681.41 ± 30.08 μmol∙min−1∙mg−1, respectively. The co-treatment of EG7A with the commercial cellulase increased the yield of reducing sugar by 155.77 % (filter paper) and 130.49 % (corn stover). Molecular docking assay showed the interaction energy of EG7A with cellotetraose at −10.50 kcal/mol, surpassing EG12A (−10.43 kcal/mol), EG12C (−10.28 kcal/mol), and EG5A (−9.00 kcal/mol). Root Mean Square Deviation (RMSD) and Solvent Accessible Surface Area (SASA) values revealed that the presence of cellotetraose stabilized the molecular dynamics simulation of the cellotetraose-protein complex over a 100 ns time scale. This study provides valuable insights for developing recombinant enzymes and biomass degradation technologies. [Display omitted] [ABSTRACT FROM AUTHOR]

Lignocellulosic biomass (LCB) is a prominent option for second-generation biofuels production. Cellulase hydrolyses cellulose, a component of LCB by attacking the β-1,4-glycosidic bonds, thus liberating mono, di, and oligosaccharides, which subsequently, can be converted to biofuel. In this study, a novel cellulase (Cel-3.1) of 1593 bp which encodes a 530 amino acid protein was identified from buffalo rumen metagenomic fosmid library, and functional expression was achieved through transformation into Escherichia coli. The molecular weight was estimated as 58 kDa on SDS-PAGE. Cel-3.1 belongs to glycosyl hydrolase family-5 (GH-5) and is predicted to have 14 α-helices and 15 β-strands. The optimal temperature and pH for Cel-3.1 were experimentally determined as 5.0 and 50 °C respectively. The synergistic effect of Ca2+ with K+ ions improved Cel-3.1 activity significantly (25%) and 1% Polyethylene Glycol (PEG-400), 1% β-mercaptoethanol enhanced the relative activity Cel-3.1 by 31.68%, 12.03% respectively. Further, the enzymatic (Cel-3.1) hydrolysis of pretreated rice straw and corncob released 13.41 ± 0.26 mg/mL and 15.04 ± 0.08 mg/mL reducing sugars respectively. High Performance Liquid Chromatography (HPLC), Scanning Electron Microscope (SEM), and Fourier Transformation Infrared spectroscopy (FTIR) analysis revealed the capability of Cel-3.1 for the breakdown and hydrolysis of both rice straw and corncob to generate various fermentable sugars. • Cel-3.1 was active at pH 5.0 and can with stand the changes up to pH 4.0 which make it tolerant to acidic environments. • It was thermally stable up to 45 °C and the optimum reaction temperature was 50 °C. • The synergistic effect of Ca2+ with K+ ions improved Cel-3.1 activity significantly (25%). • 1% Polyethylene Glycol (PEG-400) has enhanced the relative activity of Cel-3.1 by 31.68%. • 1% β-mercaptoethanol has enhanced the relative activity of Cel-3.1 by 12.03%. [ABSTRACT FROM AUTHOR]

Processive endoglucanases possess both endo- and exoglucanase activity, making them attractive discovery and engineering targets. Here, a processive endoglucanase EG5C-1 from Bacillus subtilis was employed as the starting point for enzyme engineering. Referring to the complex structure information of EG5C-1 and cellohexaose, the amino acid residues in the active site architecture were identified and subjected to alanine scanning mutagenesis. The residues were chosen for a saturation mutagenesis since their variants showed similar activities to EG5C-1. Variants D70Q and S235W showed increased activity towards the substrates CMC and Avicel, an increase was further enhanced in D70Q/S235W double mutant, which displayed a 2.1- and 1.7-fold improvement in the hydrolytic activity towards CMC and Avicel, respectively. In addition, kinetic measurements showed that double mutant had higher substrate affinity (K m) and a significantly higher catalytic efficiency (k cat /K m). The binding isotherms of wild-type EG5C-1 and double mutant D70Q/S235W suggested that the binding capability of EG5C-1 for the insoluble substrate was weaker than that of D70Q/S235W. Molecular dynamics simulations suggested that the collaborative substitutions of D70Q and S235W altered the hydrogen bonding network within the active site architecture and introduced new hydrogen bonds between the enzyme and cellohexaose, thus enhancing both substrate affinity and catalytic efficiency. • Processive endoglucanase EG5C-1 of Bacillus subtilis was mono-modular. • Substitutions of D70Q and S235W introduced new hydrogen bonds between enzyme and ligand. • Double mutant D70Q/S235W showed a higher binding capacity to insoluble substrate. • D70Q/S235W showed 2.1 and 1.7-fold higher catalytic activity towards CMC and Avicel. [ABSTRACT FROM AUTHOR]

The recently identified lytic polysaccharide monooxygenases (LPMOs) are important auxiliary proteins which contribute to lignocellulose biodegradation by oxidatively cleaving the glycosidic bonds in cellulose and other polysaccharides. The vast differences in terms of substrate specificity and regioselectivity within LPMOs provide us new possibilities to find promising candidates for the use in enzyme cocktails in biorefinery applications. In this study, a highly xyloglucan active family AA9 lytic polysaccharide monooxygenase Ep LPMO9A was identified from Eupenicillium parvum 4-14. Ep LPMO9A exhibited a mixed C1/C4 oxidative cleavage activity on cellulose and xyloglucan with a broad range of pH stability and good thermal stability at 40 °C. It showed a higher boosting effect on the enzymatic saccharification of complex lignocellulosic substrates associated with xyloglucan than on the lignocellulosic substrates without xyloglucan particularly in low commercial cellulase dosage cases. The oxidative cleavage of xyloglucan by Ep LPMO9A may facilitate to open up the sterical hindrance of cellulose by xyloglucan and thereby increase accessibility for cellulase to lignocellulosic substrates. The discovery of more and more hemicellulose-active LPMOs and their contribution to breaking down the barriers by oxidatively acting on hemicellulose may expand our knowledge for their functions of LPMOs in lignocellulose biodegradation. [ABSTRACT FROM AUTHOR]

The gene encoding Pcal_0842, annotated as a cellulase (accession no. ABO08268), was cloned and expressed in Escherichia coli. The gene product was produced in insoluble form in E. coli in high amounts even without addition of the inducer isopropyl β-D-1-thiogalactopyranoside. The recombinant protein was solubilized in 8 M urea and refolded by gradual removal of urea. The refolded protein exhibited both α-1,4- and β-1,4-glycosidic cleavage activities. The enzyme activity increased with the increase in temperature till 120 °C. Apart from very high optimal temperature, recombinant Pcal_0842 was extremely thermostable. There was no significant loss in activity even after heating for 100 h at 100 °C. The half-lives of Pcal_0842 were 6 and 2.5 h at 110 and 120 °C, respectively. To the best of our knowledge, Pcal_0842 is the most thermostable glycosidase characterized to date and this is the first report on cloning and characterization of an enzyme from archaea that displays both α-1,4- and β-1,4-glycosidic cleavage activities. [ABSTRACT FROM AUTHOR]

Cellulose, the most abundant feedstock of chemicals and energy is extracted from various agro-industrial wastes, such as sugarcane bagasse (SB). Pretreatment of SB with ionic liquids improves extraction of cellulose, yet the use of ionic liquid is hindered by its high cost. In this study, cellulose was extracted from SB pretreated with methyltrioctylammonium chloride under relatively mild conditions. The extracted cellulose from pretreated SB (PTB) and untreated SB (UTB) was characterized by scanning electron microscopy and FTIR. Fermentation of cellulose extracted from PTB by a thermophilic bacterium, Bacillus aestuarii UE25, yielded 245.16% higher titers of cellulase than cellulose extracted from UTB. The recyclability of the IL was assessed to make the pretreatment process cost effective and was monitored through TLC and FTIR. The results of this research demonstrated the potential of ionic liquid pretreated SB for cellulose extraction and for its subsequent utilization in thermostable cellulase production. Unlabelled Image • Cellulose was extracted from Methyltrioctylammonium chloride pretreated SB. • The IL could be recycled for five times without losing its efficiency. • Cellulose from SB enhanced the production of thermostable cellulase. • Cellulose from SB inhibited the production of other related enzymes. [ABSTRACT FROM AUTHOR]

In this work, magnetic Fe 2 O 3 /Fe 3 O 4 nanocomposites were prepared via a novel rapid combustion process. The silica was precipitated on the surface of Fe 2 O 3 /Fe 3 O 4 nanocomposites. The silica-coated magnetic nanocomposites were cross-linked with glutaraldehyde, on which cellulase was covalently immobilized. The morphology, composition, and property of the prepared nanomaterials were characterized by the scanning electron microscopy (SEM), the energy dispersive spectrometry (EDS), the X-ray diffraction (XRD), the vibrating sample magnetometer (VSM), and the Fourier transform infrared (FTIR) spectroscopy. The immobilization conditions were optimized by varying operating parameters and determined to be 0.05 mL of 0.5% cellulase solution for 2 h. The catalytic stabilities of the immobilized cellulase were evaluated. The results showed that the immobilized cellulases performed higher apparent activity at pH 4.5 and exhibited good thermal stability compared with their free counterparts. The Michaelis-Menten equation showed that K m and V max of free cellulase were 3.46 mol·L−1 and 0.53 mol·min−1, respectively. The immobilized cellulase had higher K m and V max (18.99 mol·L−1 and 0.59 mol·min−1). The retained activity of the immobilized cellulase maintained over 71% of the initial activity after being used for five cycles. • Hydroxy and aldehyde functionalized magnetic Fe 2 O 3 /Fe 3 O 4 nanocomposites were prepared for the immobilization of cellulase. • The morphology and properties of the prepared carrier were investigated, and immobilized conditions had been optimized. • The catalytic performances of the cellulase immobilized on magnetic Fe 2 O 3 /Fe 3 O 4 @SiO 2 -CHO nanocomposites were evaluated. [ABSTRACT FROM AUTHOR]

Pretreatment technology has attracted much attention as an effective method for the conversion of sugarcane bagasse into biochemicals. However, residual lignin-carbohydrate complexes (LCC) can negatively impact the subsequent enzymatic hydrolysis of bagasse. In this work, the changes in bagasse LCC after pretreatment with hot water and dilute acid were characterized by component analysis, 13C NMR and 1H-13C HSQC NMR to reveal the correlation between LCC structure and cellulase adsorption. A real-time dynamic model of LCC affecting adsorption of cellulase was constructed using a quartz crystal microbalance (QCM-D). The QCM-D results demonstrated that cellulase exhibited different adsorption characteristics on different LCCs. For example, the maximum adsorption capacities for cellulase onto the raw material LCC (RW-LCC), hot water pretreated LCC (LHW-LCC), and dilute acid pretreated LCC (AP-LCC) at 4 °C were 29.0 ng/cm2, 94.9 ng/cm2 and 129.8 ng/cm2, respectively. In addition, the adsorption rate constants for cellulase on RM-LCC, LHW-LCC and AP-LCC at 4 °C were 0.09, 0.14 and 0.19, respectively. [ABSTRACT FROM AUTHOR]

Cellulolytic enzymes have wide use in several industrial segments (e.g. biofuels, pulp and paper, food, and cosmetics). However, one of the challenges is their large-scale production with high specific activity to eliminate the dependence of the purchase of enzymatic cocktails produced by commercial parties. The aims of this study were (1) isolation, selection, and partial characterization of bacterial cellulases present in the intestinal tract of the sugarcane borer and (2) to identify cellulase-producing bacteria by analyzing the 16S rDNA gene. Cellulase production and purification assays resulted in similar electrophoretic profiles between four bacterial strains. These strains were identified as Klebsiella pneumoniae , Klebsiella sp., and Bacillus sp. K. pneumoniae was the main cellulase-producing microorganism. Our results show the possibility of finding cellulolytic microorganisms that inhabit the gut of herbivorous animals, especially those that are predators of important crops of economic value. Furthermore, K. pneumoniae cellulase is of medical importance. In hospitals, health professionals, hospital technicians, patients and visitors wear clothes containing cellulose. Thus, K. pneumoniae within hospitals can contaminate these clothes and be spread to the environment. In that case, it would be important for the hospital's chemical sterilization products to have at least one cellulase inhibitor. • Cellulolytic bacteria from the sugarcane borer were Klebsiella sp. and Bacillus sp. • K. pneumoniae was the main cellulase-producing microorganism. • This enzyme was able to hydrolyze sodium carboxymethylcellulose • K. pneumoniae cellulase is of medical importance. [ABSTRACT FROM AUTHOR]

Recently low-molecular lignin was reported to activate lytic polysaccharide monooxygenases (LPMOs) to oxidize cellulose. However, whether lignin formed in cell wall can play the role as electron donor for LPMOs is still largely unknown due to the complex ultrastructure of lignocellulosic biomass. In this study, we presented a new strategy to elucidate in-situ lignin function in LPMOs reaction. A lignocellulosic mimicking model was used as substrate, which was equipped with a polysaccharide template of self-assembled bacterial cellulose film and synthesized lignin. Remarkably, it has been demonstrated that lignin polymer deposited on cellulose can reduce LPMOs in-situ for cellulose oxidation and then boost cellulose hydrolysis, and the cellulose conversion ratio of the mimicked lignocellulosic film was increased by 26.0%. More importantly, lignin in-situ might exceed the well-known reductant of ascorbic acid to drive LPMOs for cellulase enzymatic hydrolysis with equivalent cellulose oxidation efficiency and extremely lower H 2 O 2 generation, avoiding the inactivation of enzymes. The maximum H 2 O 2 yield from lignin-driven LPMO reaction was 75.8% lower than that from ascorbic acid-driven reaction. Therefore, by using the lignocellulosic mimicking model, we have elucidated the function of in-situ lignin in boosting enzymatic hydrolysis. Such understanding could significantly promote current utilization of LPMOs in lignocellulosic biorefinery. • Lignocellulosic mimicking model was applied to elucidate LPMOs reaction. • Lignin polymer deposited on cellulose could activate LPMOs in-situ. • Lignin in-situ might exceed ascorbic acid to drive LPMOs to boost hydrolysis. [ABSTRACT FROM AUTHOR]

The present study was undertaken to establish a distinct relationship between blue crab chitosan (Cs) acetylation degree (AD) and molecular weight (Mw) and its structural features, thermal properties and bioactivity. Therefore, chitosans with different AD were prepared and Cellulase was used to produce Cs derivatives with decreasing Mw. Results clearly display a decrease of the ordered structure of Cs, with the increase of AD and the decrease of Mw. Thermal stability/degradation screening disclose a greater thermal resistance for Cs with lower AD and higher Mw. The anti-adhesive potential of Cs was, additionally, studied, as function of AD and Mw. The effectiveness of Cs in preventing biofilm adhesion was strongly influenced by its AD and Mw, with the lowest inhibition values for higher AD and lower Mw. Interestingly, the effectiveness of Cs in disrupting pre-formed biofilms increased with decreasing Mw. Moreover, Cs derivatives were found to be advantageously efficient in prolonging human blood clotting times, based on data of activated partial thromboplastin time, Quick time and thrombin time assays, typically for the intrinsic coagulation pathway. Accordingly, depending on the predicted application of Cs, either in food, biomedical and pharmaceutical industries, AD and Mw are critical traits to be inevitably reflected on. • Chitosan derivatives with diverse acetylation degree (AD) and molecular weight (Mw) • Greater thermal resistance for chitosan derivatives with lower AD and higher Mw • Lowest biofilm adhesion inhibition for higher AD and lower Mw chitosan derivatives • Disrupting pre-formed biofilms increase with decreasing chitosan derivatives Mw • Derivatives efficiency in prolonging clotting, typically for the intrinsic pathway [ABSTRACT FROM AUTHOR]

The manufacture of biofuels from cellulose is regarded as one of practicable strategies to meet increasing energy demand and alleviate environmental issues. Cellulases, which play an important role in the production of second-generation biofuels, are expected to be highly thermostable, halostable and organic solvent-stable to adapt to the harsh conditions in practical application. Here we cloned and characterized a novel cellulase (MaCel) from Mahella australiensis 50-1 BON, an anaerobic thermophile isolated from an oil reservoir. MaCel exhibited excellent thermostability, halostability as well as organic solvent stability, and could be efficiently produced in a yield of 1.7 × 106 U/L in 15 h with inexpensive culture medium. These results indicate that MaCel may be a suitable candidate for industrial applications, illustrating the potential benefits of enzymes from oil reservoir extremophiles in the manufacture of biofuels. Unlabelled Image [ABSTRACT FROM AUTHOR]

Effective management and the valorization of agro-industrial lignocellulosic feedstocks can only be realized if a versatile cellulase cocktail is developed that can release glucose at affordable cost irrespective of biomass type. In the present study the flexibility of using cellulase cocktail obtained from mutant UV-8 of Talaromyces verruculosus IIPC 324 in depolymerizing multiple agro-industrial lignocellulosic feedstocks was explored. Five different dilute acid pretreated biomasses were evaluated and cellulase loading was done at 25 mg protein/g cellulose content. After 72 h of hydrolysis at 55 °C and pH 4.5, corn cob and rice straw emerged as the easiest and toughest substrates with saccharification yield of 83.9 ± 1.17 and 35.5 ± 1.16% respectively from their cellulose fraction. Addition of PEG 6000 could retain >65% of all mono-component enzymes present in cellulase cocktail. Structural elucidation of biomasses gave an insight about key features responsible for variable recalcitrance in the different agro-industrial feedstock. Cellulose hydrolysis showed a significant negative correlation in the order of Cr I > S/G ratio > ash content. The chemical composition of lignin had a major impact on enzyme-lignin interactions. Higher H lignin content and lower S/G ratio promoted enzyme desorption, thereby increasing the likelihood of their recycling and reuse. [ABSTRACT FROM AUTHOR]