Commercial Cellulase Preparation Research Articles

Improving Enzymatic Saccharification of Peach Palm (Bactris gasipaes) Wastes via Biological Pretreatment with Pleurotus ostreatus.

The white-rot fungus Pleurotus ostreatus was used for biological pretreatment of peach palm (Bactris gasipaes) lignocellulosic wastes. Non-treated and treated B. gasipaes inner sheaths and peel were submitted to hydrolysis using a commercial cellulase preparation from T. reesei. The amounts of total reducing sugars and glucose obtained from the 30 d-pretreated inner sheaths were seven and five times higher, respectively, than those obtained from the inner sheaths without pretreatment. No such improvement was found, however, in the pretreated B. gasipaes peels. Scanning electronic microscopy of the lignocellulosic fibers was performed to verify the structural changes caused by the biological pretreatments. Upon the biological pretreatment, the lignocellulosic structures of the inner sheaths were substantially modified, making them less ordered. The main features of the modifications were the detachment of the fibers, cell wall collapse and, in several cases, the formation of pores in the cell wall surfaces. The peel lignocellulosic fibers showed more ordered fibrils and no modification was observed after pre-treatment. In conclusion, a seven-fold increase in the enzymatic saccharification of the Bactris gasipaes inner sheath was observed after pre-treatment, while no improvement in enzymatic saccharification was observed in the B. gasipaes peel.

Synergistic β-glucosidases for improving cellulases recyclability and biomass enzymatic saccharification in wheat straw

Cellulases required for the conversion of biomass to sugars contribute to a significant part of the final bioethanol cost in the overall lignocellulosic biorefinery. It has been proven that recycling cellulases for saccharification is a viable method to lower the cost of the enzymatic hydrolysis process. Any method for recycling enzymes must begin with a stable cellulase cocktail that can withstand operational recycling conditions. The majority of commercial cellulase preparations contain a small quantity of β-glucosidases (BG), which lowers overall recyclability and causes low hydrolysis yields. This study examines the use of synergistic β-glucosidases (AtuBG3II and PfuBG1 from Aspergillus tubingensis and Pyrococcus furiosus, respectively) to enhance the recyclability of cellulases during the hydrolysis process. It was able to recycle a significant amount of cellulases by recycling the insoluble solid fraction and liquid fraction following enzymatic hydrolysis of two-step treated wheat straw (ts-WS). In addition, the in-house cellulase cocktail (C-Mix) retained β-glucosidase activity and demonstrated glucose productivity of 18.9 g/L/h at 5% loading and enzyme efficiency (0.41 g of glucose per mg of protein) as AtuBG3II and PfuBG1 did not adsorb significantly to the biomass. This resulted in a 3.5-fold improvement in the recyclability and a 33% increase in the hydrolysis yield during the study. Therefore, all these results signify better cellulase recovery using two-step treated lignocellulose and stable cellulase cocktails, demonstrating great potential for enzymatic recycling.

Cellulases: From Lignocellulosic Biomass to Improved Production

Cellulases are enzymes that are attracting worldwide attention because of their ability to degrade cellulose in the lignocellulosic biomass and transform it into highly demanded bioethanol. The enzymatic hydrolysis of cellulose by cellulases into fermentable sugars is a crucial step in biofuel production, given the complex structure of lignocellulose. Due to cellulases’ unique ability to hydrolyze the very recaltricant nature of lignocellulosic biomass, the cellulase market demand is rapidly growing. Although cellulases have been used in industrial applications for decades, constant effort is being made in the field of enzyme innovation to develop cellulase mixtures/cocktails with improved performance. Given that the main producers of cellulases are of microbial origin, there is a constant need to isolate new microorganisms as potential producers of enzymes important for biofuel production. This review provides insight into current research on improving microbial cellulase production as well as the outlook for the cellulase market with commercial cellulase preparation involved in industrial bioethanol production.

Differential scanning calorimetry (DSC) as a tool for studying thermal properties of a crude cellulase cocktail

Differential scanning calorimetry (DSC) was used as an efficient and rapid tool in studying the conformational transitions between the folded and unfolded structures of cellulolytic enzymes. The thermal properties of two crude hydrolytic enzyme cocktails containing extracellular cellulases from Trichoderma longibrachiatum DIBAF-10 were analyzed and compared with three commercial cellulase preparations. Differences in the thermal behavior of fungal cellulases in the liquid phase, freeze-dried state, liquid formulations in sodium citrate buffer (pH 4.8), and contact with cellulose, carboxymethyl cellulose, and cellobiose were evaluated. DSC profiles of cellulases from the DIBAF-10 strain provided important thermodynamic information about the thermal stability of the included proteins. Crude enzyme cocktails underwent a reproducible and irreversible exothermic aggregation phenomenon at 52.45 ± 0.90 °C like commercial β-glucosidase. Freeze-dried and resuspended in a sodium citrate buffer, cellulases from T. longibrachiatum showed an endothermic peak dependent on buffer and enzyme concentration. In the enzyme-substrates systems, a shift of the same peak was recorded for all substrates tested. The thermal analysis of freeze-dried cellulase samples in the range of 20–150 °C gave information on the denaturation process. In conclusion, we demonstrated that DSC is a cost-effective tool for obtaining "conformational fingerprinting" of crude fungal cellulase preparations.Graphical abstract

Substrate-Dependent Cellulose Saccharification Efficiency and LPMO Activity of Cellic CTec2 and a Cellulolytic Secretome from Thermoascus aurantiacus and the Impact of H2O2-Producing Glucose Oxidase

Understanding and improving the efficiency of enzymatic saccharification of lignocellulosic biomass will promote the use of this renewable material. Here, we have studied several process parameters (reaction temperature, type of enzyme blend, type of substrate, type of reductant, and in situ supply of hydrogen peroxide) to better understand how saccharification could be optimized, focusing on the role of lytic polysaccharide monooxygenases (LPMOs). Comparison of a simple, LPMO-rich cellulolytic secretome from the thermophilic fungus Thermoascus aurantiacus with the commercial cellulase preparation Cellic CTec2 showed that saccharification of (lignin-poor) sulfite-pulped spruce at 60 °C with the secretome was as efficient as saccharification with Cellic CTec2 at 50 °C. Quantification of LPMO products showed that while LPMO activity contributed to saccharification efficiency, high levels of LPMO activity were not necessarily beneficial. Reactions with steam-exploded birch, rich in redox-active lignin, highlighted a strong impact of the feedstock on enzyme performance. In this case, the reaction with Cellic CTec2 at 50 °C was clearly most efficient. At 60 °C, enzyme inactivation became apparent for both enzyme blends, likely due to detrimental redox processes. Addition of H2O2-generating glucose oxidase to reactions with Cellic CTec2 at 50 °C led to strongly increased LPMO activity and, only for reactions with the lignin-poor substrate, improved saccharification yields. These results underpin the potential of the T. aurantiacus secretome for hydrolysis of lignin-poor substrates, and the usefulness of glucose oxidase for optimizing their saccharification. They also show that the efficiency of LPMO-containing cellulase preparations is highly dependent on the nature of the reductant and the substrate.

Enhancement of an efficient enzyme cocktail from Penicillium consortium on biodegradation of pretreated poplar

Lignocellulosic biomass is the most abundant feedstock in production of bio-based fuels and materials. In order to achieve high conversion efficiency, enzyme cocktail has been assessed by numerous studies as a powerful tool to allocate diverse specific activities in one matrix. This study reported an efficient cellulase cocktail produced by co-cultivation of several Penicillium strains with lytic polysaccharide monooxygenases (LPMOs) producing strains. The optimal inoculum proportion was screened with hydrolysis yield as the indicator and through ANOVA analysis. When blending the consortium-derived enzyme cocktail with commercial cellulase preparation, the highest hydrolysis yield reached 88.7%, equivalent to 32% increase compared to using commercial cellulase alone. The behind structure-effect relationship between the substrate and enzymatic degradation was illustrated. Comparative analysis of the enzyme components, activities and the strains transcriptome provided a comprehensive evaluation and insights of the proposed Penicillium consortium, indicating it as a potent cellulase cocktail producer for enhancing lignocellulose biodegradation.

Optimized Cellulase-Hydrolyzed Deoiled Coconut Cake Powder as Wheat Flour Substitute in Cookies.

Deoiled coconut cake powder (DCCP) was hydrolyzed to reduce the ratio of insoluble/soluble dietary fiber (RIS) by partially converting insoluble dietary fiber to soluble using Celluclast 1.5 L, a commercial cellulase preparation in citrate buffer medium. Firstly, the influence of citrate buffer amount, enzyme concentration, pH, and retention time on the enzymatic hydrolysis efficiency was investigated. Then, response surface methodology (RSM) was employed to optimize the process in which the insoluble and soluble dietary fiber contents were the responses. The results revealed that 10.3 g buffer/g of materials, 3.7 U/g of the materials, and 60 min of retention time were the optimal conditions for the enzymatic hydrolysis to obtain the insoluble and soluble contents of 68.21%db and 8.18%db, respectively. Finally, DCCP or hydrolyzed DCCP (HDCCP) was partially substituted for wheat flour at different replacement ratios in a cookie recipe at 0, 10, 20, 30, and 40%. The cookies with a 10% replacement ratio of hydrolyzed deoiled coconut cake powders had a lower RIS by more than two folds those of DCCP and had the same sensorial score as the control sample. This study proposed that Celluclast 1.5 L effectively reduced RIS by partially converting insoluble to soluble dietary fiber, improving the soluble dietary fiber content in fiber-enriched cookies.

Enhancing cellulases through synergistic β-glucosidases for intensifying cellulose hydrolysis

β-glucosidases (BGs) are key enzymes in cellulose hydrolysis, being the rate-limiting enzyme that is regulated by feedback inhibition from its own product—glucose. Due to which low hydrolysis yields are observed thus affecting overall ethanol yield per ton biomass. In this study, hydrolysis yields influence as a function of BG deficiency, supplementation, and its synergy in cellulase cocktails were investigated. Results show that commercial cellulase preparations were low in BG and produce hydrolysis yields below 60 % in 12 h. On blending commercial preparations with BGs (from fungal and bacterial sources), the degree of synergism was observed greater than 1, indicating that the components depicted synergistic behaviour in the mixture. A triple blend cellulase cocktail comprising commercial preparation, and two different BG preparations were optimized using the mixture design approach based on the synergism demonstrated. According to the findings, the in-house optimized cocktail (C-Mix) was able to hydrolyze six different two-stage treated LBM feedstocks and release more than 900 mg/g of glucose. The stability of the optimized cocktail was also enhanced two-fold due to strong synergistic interactions. Thus, a simple blending of BGs in the commercial preparations can be a sustainable approach for developing effective cocktails and intensifying cellulose hydrolysis.

Expression and production of thermophilic alginate lyases in Bacillus and direct application of culture supernatant for seaweed saccharification

Brown seaweeds are rich in carbohydrates and may be used as a source of fermentable sugars. Saccharification of the seaweed biomass can be carried out enzymatically by a combination of cellulases and alginate lyases. In this study, thermotolerant exo- and endo alginate lyases were cloned and expressed in Bacillus subtilis. The lyases were secreted to the culture supernatant and used directly together with a commercial cellulase preparation to saccharify Saccharina latissima biomass. The results showed that the strategy of using the culture supernatants directly as a source of alginate lyases worked very well, releasing glucose, mannitol, and uronic acids. The ratio between the exo- and endo-acting alginate lyases proved to be very important for saccharification yield, and under optimal reaction conditions the use of culture supernatants containing alginate lyases improved final glucose concentration by 73%, when compared to only applying cellulases. This direct use of culture supernatants as a source of alginate lyases shows that enzyme purification steps are not needed, saving seaweed processing costs and points to the possibility of a relatively simple on-site enzyme production for seaweed biorefining.

Effects of enzymatic treatment conditions on dietary fibre content of wheat bran and use of cellulase‐treated bran in cookie

SummaryA commercial cellulase preparation (Viscozyme Cassava C) was employed to process wheat bran to improve the ratio of insoluble to soluble dietary fibre. The effects of initial moisture content, enzyme dosage, and incubation time on the content of soluble and insoluble fibre during the enzymatic treatment were investigated. The appropriate conditions for the cellulase treatment were initial moisture content of 0.75 g water g−1 dry matter, enzyme dosage of 9 U g−1 dry matter and incubation time of 120 min under which the ratio of insoluble to soluble fibre of wheat bran was lowered by 42%. Untreated and cellulase‐treated wheat bran was incorporated into cookies at levels of 0%, 20%, 30%, 40% and 50%. Cookie with 50% cellulase‐treated bran showed 21% lower ratio of insoluble to soluble fibre as well as 14% lower hardness and 13% higher overall acceptability than sample with the same amount of untreated bran.

Efficient Constitutive Expression of Cellulolytic Enzymes in Penicillium oxalicum for Improved Efficiency of Lignocellulose Degradation.

Efficient cellulolytic enzyme production is important for the development of lignocellulose-degrading enzyme mixtures. However, purification of cellulases from their native hosts is time- and labor-consuming. In this study, a constitutive expression system was developed in Penicillium oxalicum for the secreted production of proteins. Using a constitutive polyubiquitin gene promoter and cultivating with glucose as the sole carbon source, nine cellulolytic enzymes of different origins with relatively high purity were produced within 48 h. When supplemented to a commercial cellulase preparation, cellobiohydrolase I from P. funiculosum and cellobiohydrolase II from Talaromyces verruculosus showed remarkable enhancing effects on the hydrolysis of steam-exploded corn stover. Additionally, a synergistic effect was observed for these two cellobiohydrolases during the hydrolysis. Taken together, the constitutive expression system provides a convenient tool for the production of cellulolytic enzymes, which is expected to be useful in the development of highly efficient lignocellulose-degrading enzyme mixtures.

Improving the fermentable sugar yields of wheat straw by high-temperature pre-hydrolysis with thermophilic enzymes of Malbranchea cinnamomea

BackgroundEnzymatic hydrolysis is a key step in the conversion of lignocellulosic polysaccharides to fermentable sugars for the production of biofuels and high-value chemicals. However, current enzyme preparations from mesophilic fungi are deficient in their thermostability and biomass-hydrolyzing efficiency at high temperatures. Thermophilic fungi represent promising sources of thermostable and highly active enzymes for improving the biomass-to-sugar conversion process. Here we present a comprehensive study on the lignocellulosic biomass-degrading ability and enzyme system of thermophilic fungus Malbranchea cinnamomea N12 and the application of its enzymes in the synergistic hydrolysis of lignocellulosic biomass.ResultsMalbranchea cinnamomea N12 was capable of utilizing untreated wheat straw to produce high levels of xylanases and efficiently degrading lignocellulose under thermophilic conditions. Temporal analysis of the wheat straw-induced secretome revealed that M. cinnamomea N12 successively degraded the lignocellulosic polysaccharides through sequential secretion of enzymes targeting xylan and cellulose. Xylanase-enriched cocktail from M. cinnamomea N12 was more active on native and alkali‑pretreated wheat straw than the commercial xylanases from Trichoderma reesei over temperatures ranging from 40 to 75 °C. Integration of M. cinnamomea N12 enzymes with the commercial cellulase preparation increased the glucose and xylose yields of alkali‑pretreated wheat straw by 32 and 166%, respectively, with pronounced effects at elevated temperature.ConclusionsThis study demonstrated the remarkable xylanase-producing ability and strategy of sequential lignocellulose breakdown of M. cinnamomea N12. A new process for the hydrolysis of lignocellulosic biomass was proposed, comprising thermophilic enzymolysis by enzymes of M. cinnamomea N12 followed with mesophilic enzymolysis by commercial cellulases. Developing M. cinnamomea N12 as platforms for thermophilic enzyme mixture production will provide new perspectives for improved conversion yields for current biomass saccharification schemes.

Integrated biorefinery approach to utilization of pulp and paper mill sludge for value-added products

The objective of this work was to examine opportunities for reducing the overall cost of lignocellulose hydrolysis to fermentable sugars. Primary sludge (PS), a negative cost lignocellulosic feedstock, was hydrolyzed sludge using a commercial cellulase preparation (Cellic® CTec2) in presence of non-ionic surfactants. Polyethylene glycol (PEG) 4000 facilitated the highest hydrolysis yield of 74.4% which represented a 2-fold increase over the control without surfactant. Response surface methodological analysis at four variables (hydrolysis time, solids, enzyme, and surfactant loadings) revealed that enzymatic hydrolysis was significantly enhanced by the interactive effect of all factors with solids and enzyme loadings as the most significant parameters (p < 0.05). Using ultrafiltration, 40% of the cellulase enzyme was recovered from the enzymatic hydrolysate and reused on fresh PS substrate. The hydrolysate fermentabilty was evaluated in production of 9.7 g/L bioethanol at 92% ethanol yield using Saccharomyces cerevisiae, and in production of 37.8% biolipids by the oleaginous yeast Cutaneotrichosporon oleaginosum at a carbon to nitrogen ratio of 40. The compression load of a sample prepared from the unhydrolyzed PS solids was 102% higher than Portland cement. The potential for valorization of the unhydrolyzed solids as reinforcement material warrants further investigations.

Biological pretreatment of rice straw with cellulase-free xylanolytic enzyme-producing Bacillus firmus K-1: Structural modification and biomass digestibility

Biological pretreatment using microorganisms or enzymes offers an eco-friendly process for biomass processing. Herein, the efficiency of pretreatment of rice straw with the cellulase-free xylanolytic enzyme-producing Bacillus firmus K-1 and its enzymes was assessed. After pretreatment with strain K-1 (BRS), the xylan content in rice straw reduced significantly (21% removal), thus increasing exposure of the cellulose crystal structure (Crystallinity index (CrI) = 40.2%) and creating biomass porosity. Subsequent treatment of BRS with the in-house xylanase preparation (BRS-E) slightly increased xylan removal (30% removal). The reduction of xylan thus led to larger pore size and increased crystallinity (CrI = 42.8%). Compared to untreated rice straw (24% glucan conversion), hydrolysis of BRS and BRS-E with the commercial cellulase preparation Accellerase 1500 at 100 g/L substrate load showed comparable glucose yield, giving about 74% glucan conversion. The results indicate that the removal of xylan can enhance accessibility of cellulose to cellulases, although the lignin content was not reduced (24% for BRS and 25% for BRS-E). This work demonstrates a new insight into the improvement of pretreatment efficiency using a xylan-degrading microorganism, which is an alternative to conventional lignin removal by fungal pretreatment.

Enzyme-Assisted Fucoidan Extraction from Brown Macroalgae Fucus distichus subsp. evanescens and Saccharina latissima.

Fucoidans from brown macroalgae (brown seaweeds) have different structures and many interesting bioactivities. Fucoidans are classically extracted from brown seaweeds by hot acidic extraction. Here, we report a new targeted enzyme-assisted methodology for fucoidan extraction from brown seaweeds. This enzyme-assisted extraction protocol involves a one-step combined use of a commercial cellulase preparation (Cellic®CTec2) and an alginate lyase from Sphingomonas sp. (SALy), reaction at pH 6.0, 40 °C, removal of non-fucoidan polysaccharides by Ca2+ precipitation, and ethanol-precipitation of crude fucoidan. The workability of this method is demonstrated for fucoidan extraction from Fucus distichus subsp. evanescens (basionym Fucus evanescens) and Saccharina latissima as compared with mild acidic extraction. The crude fucoidans resulting directly from the enzyme-assisted method contained considerable amounts of low molecular weight alginate, but this residual alginate was effectively removed by an additional ion-exchange chromatographic step to yield pure fucoidans (as confirmed by 1H NMR). The fucoidan yields that were obtained by the enzymatic method were comparable to the chemically extracted yields for both F. evanescens and S. latissima, but the molecular sizes of the fucoidans were significantly larger with enzyme-assisted extraction. The molecular weight distribution of the fucoidan fractions was 400 to 800 kDa for F. evanescens and 300 to 800 kDa for S. latissima, whereas the molecular weights of the corresponding chemically extracted fucoidans from these seaweeds were 10–100 kDa and 50–100 kDa, respectively. Enzyme-assisted extraction represents a new gentle strategy for fucoidan extraction and it provides new opportunities for obtaining high yields of native fucoidan structures from brown macroalgae.

Redesigning the Aspergillus nidulans xylanase regulatory pathway to enhance cellulase production with xylose as the carbon and inducer source

BackgroundBiomass contains cellulose (C6-sugars), hemicellulose (C5-sugars) and lignin. Biomass ranks amongst the most abundant hydrocarbon resources on earth. However, biomass is recalcitrant to enzymatic digestion by cellulases. Physicochemical pretreatment methods make cellulose accessible but partially destroy hemicellulose, producing a C5-sugar-rich liquor. Typically, digestion of pretreated LCB is performed with commercial cellulase preparations, but C5-sugars could in principle be used for “on site” production of cellulases by genetically engineered microorganism, thereby reducing costs.ResultsHere we report a succession of genetic interventions in Aspergillus nidulans that redesign the natural regulatory circuitry of cellulase genes in such a way that recombinant strains use C5-sugar liquors (xylose) to grow a vegetative tissue and simultaneously accumulate large amounts of cellulases. Overexpression of XlnR showed that under xylose-induction conditions only xylanase C was produced. XlnR overexpression strains were constructed that use the xynCp promoter to drive the production of cellobiohydrolases, endoglucanases and β-glucosidase. All five cellulases accumulated at high levels when grown on xylose. Production of cellulases in the presence of pretreated-biomass C5-sugar liquors was investigated, and cellulases accumulated to much higher enzyme titers than those obtained for traditional fungal cell factories with cellulase-inducing substrates.ConclusionsBy replacing expensive substrates with a cheap by-product carbon source, the use of C5-sugar liquors directly derived from LCB pretreatment processes not only reduces enzyme production costs, but also lowers operational costs by eliminating the need for off-site enzyme production, purification, concentration, transport and dilution.

Application of high-throughput screening for evaluating hydrolytic potential of cellulases

Heterogeneous nature of cellulose is defiant to enzymatic hydrolysis and individual methods for activity determination of cellulases are inefficient to define actual potency. Hence, three major enzyme activities (FPase, CMCase, CBase) along with protein concentrations were determined to define biomass hydrolyzing potential of cellulases. In present study, tenfold downscaled (modified) assays based on high-throughput screening (HTS) system were developed to increase the throughput and efficiency of enzyme component assays. HTS is a powerful tool which facilitates discovery of new or improved enzymes with appropriate biochemical properties for commercial biorefineries. Cellulase activity of four commercial cellulase preparations was determined and was found to have no significant difference at the 95% confidence level between the values obtained using standard (IUPAC) method and modified microplate method. This modification allowed us to characterize cellulases based on the relative ratios of their individual components and avoids ambiguity concerning potency based on the conventional FPase assay. Also, this work puts forth a simple, rapid, and complete cellulolytic enzyme assay method.

The cellulose binding region in Trichoderma reesei cellobiohydrolase I has a higher capacity in improving crystalline cellulose degradation than that of Penicillium oxalicum

Commercial cellulase preparations for lignocellulose bioconversion are mainly produced by the fungus Trichoderma reesei. The maximum cellulose conversion of T. reesei cellulase mixture was 15%–20% higher than that of Penicillium oxalicum in the hydrolysis of corncob residue and Avicel. Nevertheless, both preparations hydrolyzed more than 92% of cellulose in NaOH-mercerized Avicel. When added to Avicel hydrolysis residue that was less reactive to P. oxalicum cellulases, cellobiohydrolase I (CBH I) from T. reesei resulted in a higher cellulose conversion than its homologous proteins from P. oxalicum and Aspergillus niger at the same protein loadings. Further domain exchange experiment attributed the high hydrolytic efficiency of T. reesei CBH I to its inter-domain linker and cellulose-binding domain. The results in part explained the superior performance of T. reesei cellulases on the degradation of native crystalline cellulose, and highlighted the important role of cellulose-binding region in determining the degree of hydrolysis by cellulases.

Enhancing enzyme-aided production of fermentable sugars from poplar pulp in the presence of non-ionic surfactants.

Addition of surfactants to enzymatic hydrolysis has been reported to enhance the hydrolytic potential of enzymes in the bioconversion of lignocellulosic biomass to fermentable sugars. The objective of this investigation was to evaluate the effects of four non-ionic surfactants (PEG4000, PEG8000, TitronX-100, and Tween80) on the efficiency of enzymatic hydrolysis of steam-pretreated poplar using a commercial cellulase preparation (Cellic® CTec2). Statistical discriminant analysis at four variable factors (surfactant type, surfactant concentration, hydrolysis time, and substrate consistency) revealed that enzymatic hydrolysis was significantly enhanced in the presence of PEG4000, with 19.2% increase in glucose yield over control without surfactant, whereas ANOVA test indicated substrate consistency and hydrolysis time as the most significant factors (P < 0.05). Hydrolysis of poplar pulp at 5% w/w pulp consistency with CTec2 in presence of 1% w/w PEG4000 produced the highest glucose yield of 58.5% after 96h reaction time.

Lipopeptide produced from Bacillus sp. W112 improves the hydrolysis of lignocellulose by specifically reducing non-productive binding of cellulases with and without CBMs

BackgroundSurfactants have attracted increasing interest for their capability to improve the enzymatic hydrolysis of lignocellulosic biomass. Compared to chemical surfactants, biosurfactants have a broader prospect for industrial applications because they are more environmentally friendly and more effective in some researches. Commercial cellulase preparations are mainly composed of endoglucanases (EGs) and cellobiohydrolases (CBHs) that possess carbohydrate-binding modules (CBMs). However, the effects of lipopeptide-type biosurfactants on enzymatic saccharification of lignocellulose and adsorption behaviors of cellulases with CBMs remain unclear.ResultsIn this study, we found that Bacillus sp. W112 could produce a lipopeptide-type biosurfactant from untreated biomass, such as wheat bran and Jerusalem artichoke tuber. The lipopeptide could enhance the enzymatic hydrolysis of dilute acid pretreated Giant Juncao grass (DA-GJG) by fungal and bacterial enzymes. The enhancement increased over a range of temperatures from 30 to 50 °C. Lipopeptide was shown to be more effective in promoting DA-GJG saccharification than chemical surfactants at low dosages, with a best stimulatory degree of 20.8% at 2% loading of the substrates (w/w). Lipopeptide increased the thermostability of EG and CBH in commercial cellulase cocktails. Moreover, the dual effects of lipopeptide on the adsorption behaviors of cellulases were found. It specifically lowered the non-productive binding of cellulases to lignin and increased the binding of cellulases to cellulose. In addition, we investigated the influence of lipopeptide on the adsorption behaviors of CBHs with CBMs for the first time. Our results showed that lipopeptide reduced the adsorption of CBM-deleted CBH to DA-GJG to a greater extent than that of intact CBH while the non-productive binding of intact CBH to lignin was reduced more, indicating that lipopeptide decreased the binding of CBMs onto lignin but not their combination with cellulose.ConclusionsIn this study, we found that lipopeptide from Bacillus sp. W112 promoted the enzymatic hydrolysis of DA-GJG at relative low loadings. The stimulatory effect could be attributed to increasing the cellulase thermostability, reducing non-productive adsorption of cellulases with CBMs caused by lignin and enhancing the binding of cellulases to cellulose.