A high performance  Trichoderma reesei  strain that reveals the importance of xylanase III in cellulosic biomass conversion

Nakazawa, Hikaru; Kawai, Tetsushi; Ida, Noriko; Shida, Yosuke; Shioya, Kouki; Kobayashi, Yoshinori; Okada, Hirofumi; Tani, Shuji; Sumitani, Jun-ichi; Kawaguchi, Takashi; Morikawa, Yasushi; Ogasawara, Wataru

doi:10.1016/j.enzmictec.2015.08.019

Cited by 20 publications

(16 citation statements)

References 25 publications

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“…When treated with JN13H, the glucose and xylose yields were 43 ± 1 and 49 ± 0% (1.0 mg/g-biomass) and 69 ± 1 and 70 ± 2% (2.0 mg/g-biomass), respectively. It has been reported that JN13H, which contains TrXyn3 in the preparation, is suitable for use with alkaline-pretreated biomass such as sodium hydroxide-pretreated rice straw [ 29 ]. TrXyn3 belongs to glycoside hydrolase family 10 (GH10), while T. reeei XYN1 (TrXyn1) and XYN2 (TrXyn2), which are contained in the JN11H and JN13H, belong to GH11 [ 44 ], and it was reported that GH11 xylanases cleave in unsubstituted regions of the arabinoxylan backbone, whereas GH10 xylanases cleave in decorated regions [ 45 ].…”

Section: Resultsmentioning

confidence: 99%

“…Characterization of BGL1 from A. aculeatus (AaBGL1) was also reported [ 25 , 26 ], and it was found that this enzyme showed higher specific activity against cellobiose than TrBGL1 (AaBGL1 has an activity of 180 U/mg, while that of TrBGL1 is 19 U/mg) [ 27 ], and a higher Ki value for glucose than AnBGL1 (AaBGL1 has a Ki of 10 mM, while that of AnBGL1 is 3 mM) [ 26 , 28 ]. Therefore, Nakazawa et al constructed three AaBGL1-expressing T. reesei strains, X3AB1, C1AB1, and E1AB1 to enhance the performance of cellulase [ 27 , 29 ]. Strains X3AB1, C1AB1, and E1AB1 were derivatives of T. reesei strain PC-3-7 containing an aabg1 cDNA under the control of the xyn3 , cbh1 , and eg1 promoters, respectively.…”

Section: Introductionmentioning

confidence: 99%

“…X3AB1 was homologously recombined at the xyn3 locus, and therefore this strain did not harbor the xyn3 gene in the genome. Enzyme preparations named JN11H, JN12H, and JN13H were, respectively, produced from strains X3AB1, C1AB1, and E1AB1 grown on cellulose and xylan as a carbon source, and JN11H and JN13H showed almost the same saccharification efficiency against various substrates at one-fifth or less dosage of an enzyme preparation from PC-3-7 [ 29 , 30 ]. As a result of such studies, the dosage of enzymes for saccharification has been reduced.…”

Section: Introductionmentioning

confidence: 99%

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A novel GH10 xylanase from Penicillium sp. accelerates saccharification of alkaline-pretreated bagasse by an enzyme from recombinant Trichoderma reesei expressing Aspergillus β-glucosidase

Shibata

Suetsugu

Kakeshita

et al. 2017

Biotechnol Biofuels

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Background Trichoderma reesei is considered a candidate fungal enzyme producer for the economic saccharification of cellulosic biomass. However, performance of the saccharifying enzymes produced by T. reesei is insufficient. Therefore, many attempts have been made to improve its performance by heterologous protein expression. In this study, to increase the conversion efficiency of alkaline-pretreated bagasse to sugars, we conducted screening of biomass-degrading enzymes that showed synergistic effects with enzyme preparations produced by recombinant T. reesei.Results Penicillium sp. strain KSM-F532 produced the most effective enzyme to promote the saccharification of alkaline-pretreated bagasse. Biomass-degrading enzymes from strain KSM-F532 were fractionated and analyzed, and a xylanase, named PspXyn10, was identified. The amino acid sequence of PspXyn10 was determined by cDNA analysis: the enzyme shows a modular structure consisting of glycoside hydrolase family 10 (GH10) and carbohydrate-binding module family 1 (CBM1) domains. Purified PspXyn10 was prepared from the supernatant of a recombinant T. reesei strain. The molecular weight of PspXyn10 was estimated to be 55 kDa, and its optimal temperature and pH for xylanase activity were 75 °C and pH 4.5, respectively. More than 80% of the xylanase activity was maintained at 65 °C for 10 min. With beechwood xylan as the substrate, the enzyme had a K m of 2.2 mg/mL and a V max of 332 μmol/min/mg. PspXyn10ΔCBM, which lacked the CBM1 domain, was prepared by limited proteolysis. PspXyn10ΔCBM showed increased activity against soluble xylan, but decreased saccharification efficiency of alkaline-pretreated bagasse. This result indicated that the CBM1 domain of PspXyn10 contributes to the enhancement of the saccharification efficiency of alkaline-pretreated bagasse. A recombinant T. reesei strain, named X2PX10, was constructed from strain X3AB1. X3AB1 is an Aspergillus aculeatus β-glucosidase-expressing T. reesei PC-3-7. X2PX10 also expressed PspXyn10 under the control of the xyn2 promoter. An enzyme preparation from X2PX10 showed almost the same saccharification efficiency of alkaline-pretreated bagasse at half the enzyme dosage as that used for an enzyme preparation from X3AB1.ConclusionsOur results suggest that PspXyn10 promotes the saccharification of alkaline-pretreated bagasse more efficiently than TrXyn3, a GH10 family xylanase from T. reesei, and that the PspXyn10-expressing strain is suitable for enzyme production for biomass saccharification.Electronic supplementary materialThe online version of this article (10.1186/s13068-017-0970-2) contains supplementary material, which is available to authorized users.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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A novel GH10 xylanase from Penicillium sp. accelerates saccharification of alkaline-pretreated bagasse by an enzyme from recombinant Trichoderma reesei expressing Aspergillus β-glucosidase

Shibata

Suetsugu

Kakeshita

et al. 2017

Biotechnol Biofuels

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“…To attain the optimal saccharifying potential in T. reesei, we constructed a new strain E1AB1, in which aabg1 was expressed heterologously by means of the egl1 promoter, so that the endogenous XYNIII synthesis remained intact. 129) Due to the presence of wild-type xyn3 in T. reesei E1AB1, JN13, the enzyme preparations from this strain, was 20-30% more effective in the saccharification of NaOH-pretreated rice straw than enzyme extracts from X3AB1, and it also outperformed recent commercial cellulase preparations. These results demonstrate the importance of XYNIII in the conversion of NaOH-pretreated cellulosic biomass by T. reesei.…”

Section: Construction Of a Recombinant T Reesei Strain For Effimentioning

confidence: 93%

Deciphering the molecular mechanisms behind cellulase production in Trichoderma reesei, the hyper-cellulolytic filamentous fungus

Shida

Furukawa

Ogasawara

2016

Bioscience, Biotechnology, and Biochemistry

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The filamentous fungus Trichoderma reesei is a potent cellulase producer and the best-studied cellulolytic fungus. A lot of investigations not only on glycoside hydrolases produced by T. reesei, but also on the machinery controlling gene expression of these enzyme have made this fungus a model organism for cellulolytic fungi. We have investigated the T. reesei strain including mutants developed in Japan in detail to understand the molecular mechanisms that control the cellulase gene expression, the biochemical and morphological aspects that could favor this phenotype, and have attempted to generate novel strains that may be appropriate for industrial use. Subsequently, we developed recombinant strains by combination of these insights and the heterologous-efficient saccharifing enzymes. Resulting enzyme preparations were highly effective for saccharification of various biomass. In this review, we present some of the salient findings from the recent biochemical, morphological, and molecular analyses of this remarkable cellulase hyper-producing fungus.

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“…The lack of β-glucosidase leads to the accumulation of cellobiose during hydrolysis, which in turn slows down the activity of the other key cellulases such as cellobiohydrolases and endoglucanases. T. reesei strains have been engineered to overexpress native [ 27 , 28 ] and heterologous [ 29 – 33 ] β-glucosidases in several prior studies.…”

Section: Introductionmentioning

confidence: 99%

Development of a low-cost cellulase production process using Trichoderma reesei for Brazilian biorefineries

Ellilä

Fonseca²,

Uchima³

et al. 2017

Biotechnol Biofuels

184

101

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BackgroundDuring the past few years, the first industrial-scale cellulosic ethanol plants have been inaugurated. Although the performance of the commercial cellulase enzymes used in this process has greatly improved over the past decade, cellulases still represent a very significant operational cost. Depending on the region, transport of cellulases from a central production facility to a biorefinery may significantly add to enzyme cost. The aim of the present study was to develop a simple, cost-efficient cellulase production process that could be employed locally at a Brazilian sugarcane biorefinery.ResultsOur work focused on two main topics: growth medium formulation and strain improvement. We evaluated several Brazilian low-cost industrial residues for their potential in cellulase production. Among the solid residues evaluated, soybean hulls were found to display clearly the most desirable characteristics. We engineered a Trichoderma reesei strain to secrete cellulase in the presence of repressing sugars, enabling the use of sugarcane molasses as an additional carbon source. In addition, we added a heterologous β-glucosidase to improve the performance of the produced enzymes in hydrolysis. Finally, the addition of an invertase gene from Aspegillus niger into our strain allowed it to consume sucrose from sugarcane molasses directly. Preliminary cost analysis showed that the overall process can provide for very low-cost enzyme with good hydrolysis performance on industrially pre-treated sugarcane straw.ConclusionsIn this study, we showed that with relatively few genetic modifications and the right growth medium it is possible to produce considerable amounts of well-performing cellulase at very low cost in Brazil using T. reesei. With further enhancements and optimization, such a system could provide a viable alternative to delivered commercial cellulases.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-017-0717-0) contains supplementary material, which is available to authorized users.

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A high performance Trichoderma reesei strain that reveals the importance of xylanase III in cellulosic biomass conversion

Cited by 20 publications

References 25 publications

A novel GH10 xylanase from Penicillium sp. accelerates saccharification of alkaline-pretreated bagasse by an enzyme from recombinant Trichoderma reesei expressing Aspergillus β-glucosidase

A novel GH10 xylanase from Penicillium sp. accelerates saccharification of alkaline-pretreated bagasse by an enzyme from recombinant Trichoderma reesei expressing Aspergillus β-glucosidase

Deciphering the molecular mechanisms behind cellulase production in Trichoderma reesei, the hyper-cellulolytic filamentous fungus

Development of a low-cost cellulase production process using Trichoderma reesei for Brazilian biorefineries

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