Synthetic multi-component enzyme mixtures for deconstruction of lignocellulosic biomass

Banerjee, Goutami; Car, Suzana; Scott-Craig, John S.; Borrusch, Melissa S.; Bongers, Mareike; Walton, Jonathan D.

doi:10.1016/j.biortech.2010.07.028

Cited by 110 publications

(123 citation statements)

References 22 publications

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“…It has been shown that incorporating accessory enzymes such as hemicellulases and lytic polysaccharide monooxygenases in cellulase mixtures enhances the conversion to monosaccharides, thereby allowing reduced enzyme loading and crucially reducing process costs [74][75][76][77]. Although the use of xyloglucanactive enzymes as accessory enzymes has not yet been explored extensively, data indicate that endoxyloglucanases and a-xylosidase can synergistically increase the hydrolytic capacity of cellulases [78,79], perhaps by removing XyG that is intimately associated with cellulose microfibrils.…”

Section: Discussionmentioning

confidence: 99%

Functional and structural characterization of a potent GH74 endo‐xyloglucanase from the soil saprophyte Cellvibrio japonicus unravels the first step of xyloglucan degradation

et al. 2016

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The heteropolysaccharide xyloglucan (XyG) comprises up to one‐quarter of the total carbohydrate content of terrestrial plant cell walls and, as such, represents a significant reservoir in the global carbon cycle. The complex composition of XyG requires a consortium of backbone‐cleaving endo‐xyloglucanases and side‐chain cleaving exo‐glycosidases for complete saccharification. The biochemical basis for XyG utilization by the model Gram‐negative soil saprophytic bacterium Cellvibrio japonicus is incompletely understood, despite the recent characterization of associated side‐chain cleaving exo‐glycosidases. We present a detailed functional and structural characterization of a multimodular enzyme encoded by gene locus CJA_2477. The CJA_2477 gene product comprises an N‐terminal glycoside hydrolase family 74 (GH74) endo‐xyloglucanase module in train with two carbohydrate‐binding modules (CBMs) from families 10 and 2 (CBM10 and CBM2). The GH74 catalytic domain generates Glc4‐based xylogluco‐oligosaccharide (XyGO) substrates for downstream enzymes through an endo‐dissociative mode of action. X‐ray crystallography of the GH74 module, alone and in complex with XyGO products spanning the entire active site, revealed a broad substrate‐binding cleft specifically adapted to XyG recognition, which is composed of two seven‐bladed propeller domains characteristic of the GH74 family. The appended CBM10 and CBM2 members notably did not bind XyG, nor other soluble polysaccharides, and instead were specific cellulose‐binding modules. Taken together, these data shed light on the first step of xyloglucan utilization by C. japonicus and expand the repertoire of GHs and CBMs for selective biomass analysis and utilization. Database Structural data have been deposited in the RCSB protein database under the Protein Data Bank codes: http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5FKR, http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5FKS, http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5FKT and http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5FKQ.

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Section: Discussionmentioning

confidence: 99%

Functional and structural characterization of a potent GH74 endo‐xyloglucanase from the soil saprophyte Cellvibrio japonicus unravels the first step of xyloglucan degradation

et al. 2016

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“…They were then centrifuged at 3,000 ϫ g for 10 min, resuspended in assay buffer (20 mM Tris-acetate, pH 6.0, 1 mM CaCl 2 , 0.1% sodium azide), and recentrifuged, and the final cell pellet was resuspended in assay buffer. Lignocellulosic biomass was then added to the cell suspension such that there was a total of ϳ15 mg of cell-displayed cellulase enzymes per gram of biomass; 10-ml suspensions containing cells at an OD 600 of 2.5 were incubated with 60 mg of biomass at 37°C with shaking (22,42). In some instances, exogenous ␤-glucosidase (Sigma) was added to the cell-biomass mix (1 mg/g biomass), and the amount of cell suspension used was correspondingly adjusted to maintain a ratio of ϳ15 mg enzyme per g biomass.…”

Section: Methodsmentioning

confidence: 99%

Recombinant Bacillus subtilis That Grows on Untreated Plant Biomass

Anderson

Miller

Fierobe

et al. 2013

Appl Environ Microbiol

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Lignocellulosic biomass is a promising feedstock to produce biofuels and other valuable biocommodities. A major obstacle to its commercialization is the high cost of degrading biomass into fermentable sugars, which is typically achieved using cellulolytic enzymes from Trichoderma reesei. Here, we explore the use of microbes to break down biomass. Bacillus subtilis was engineered to display a multicellulase-containing minicellulosome. The complex contains a miniscaffoldin protein that is covalently attached to the cell wall and three noncovalently associated cellulase enzymes derived from Clostridium cellulolyticum (Cel48F, Cel9E, and Cel5A). The minicellulosome spontaneously assembles, thus increasing the practicality of the cells. The recombinant bacteria are highly cellulolytic and grew in minimal medium containing industrially relevant forms of biomass as the primary nutrient source (corn stover, hatched straw, and switch grass). Notably, growth did not require dilute acid pretreatment of the biomass and the cells achieved densities approaching those of cells cultured with glucose. An analysis of the sugars released from acid-pretreated corn stover indicates that the cells have stable cellulolytic activity that enables them to break down 62.3% ؎ 2.6% of the biomass. When supplemented with beta-glucosidase, the cells liberated 21% and 33% of the total available glucose and xylose in the biomass, respectively. As the cells display only three types of enzymes, increasing the number of displayed enzymes should lead to even more potent cellulolytic microbes. This work has important implications for the efficient conversion of lignocellulose to value-added biocommodities.

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“…To further increase specific activities, the production of nonessential enzymes should be reduced, and the relative ratio of critical enzymes to nonessential enzymes should be enhanced. This results in reduced enzyme cost (Banerjee et al 2010). Improving specific activities is one method to reduce enzyme cost.…”

Section: An Evaluation For Saccharification Efficiency Of S Commune mentioning

confidence: 99%

Optimization of Medium Components for β-glucosidase Production in Schizophyllum commune KUC9397 and Enzymatic Hydrolysis of Lignocellulosic Biomass

Lee¹,

Lee²,

Kim³

et al. 2014

BioResources

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Medium components for β-glucosidase (BGL) production in Schizophyllum commune KUC9397 were optimized using a central composite design and response surface methodology. From the various medium components tested, cellulose, soy peptone, and thiamine HCl were selected as the optimal carbon, nitrogen, and vitamin sources, respectively. The highest BGL production was obtained with 2.96% cellulose, 2.30% soy peptone, and 0.11% thiamine HCl. BGL production in the optimized medium was increased 7.2-fold compared to production in an unoptimized medium. Crude enzyme preparation from S. commune KUC9397 was used to saccharify pretreated lignocellulosic biomass. The crude enzyme preparations showed statistically equal saccharification rates as Cellobiase, a commercial BGL. This finding indicates that crude enzymes produced by S. commune KUC9397 have good potential for application in cellulosic biomass conversion systems in place of Cellobiase.

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Synthetic multi-component enzyme mixtures for deconstruction of lignocellulosic biomass

Cited by 110 publications

References 22 publications

Functional and structural characterization of a potent GH74 endo‐xyloglucanase from the soil saprophyte Cellvibrio japonicus unravels the first step of xyloglucan degradation

Functional and structural characterization of a potent GH74 endo‐xyloglucanase from the soil saprophyte Cellvibrio japonicus unravels the first step of xyloglucan degradation

Recombinant Bacillus subtilis That Grows on Untreated Plant Biomass

Optimization of Medium Components for β-glucosidase Production in Schizophyllum commune KUC9397 and Enzymatic Hydrolysis of Lignocellulosic Biomass

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