Efficient cellulose solubilization by a combined cellulosome-β-glucosidase system

Lamed, Raphael; Kenig, Rina; Morgenstern, Ely; Calzada, J. F.; Micheo, F. de; Bayer, Edward A.

doi:10.1007/bf02921525

Cited by 47 publications

(31 citation statements)

References 15 publications

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“…A 96-h reaction resulted in 10-fold enhancement of crystalline cellulose degradation. Lamed et al (22) achieved similar results using a purified β-glucosidase from the fungus Aspergillus niger combined with purified preparations of the C. thermocellum cellulosome. Nevertheless, neither of these approaches address the fact that the cellulosome is bound to the insoluble substrate; thus, cellobiose accumulates in close proximity to the cellulosomecellulose interface and only a fraction of soluble β-glucosidase would be available to convert the inhibitory cellobiose to the noninhibitory glucose.…”

Section: Discussionmentioning

confidence: 81%

“…In contrast to natural environments where the cellobiose is removed from the medium by microbial assimilation, efficient cellulolysis by cell-free enzymatic systems would benefit by the removal of cellobiose (22,33). In this context, Kadam and Demain (21) used a combined system of cloned β-glucosidase (BglB) from C. thermocellum with the crude cellulase system from the same strain.…”

Section: Discussionmentioning

confidence: 99%

“…Previous works have shown that addition of the cellobiosedegrading enzyme β-glucosidase can enhance the rate and degree of solubilization of crystalline cellulose by the C. thermocellum cellulosome (21,22). It does so by converting cellobiose to two molecules of noninhibitory glucose; however, in the process of crystalline cellulose degradation, the cell-free cellulosome binds to the insoluble cellulosic substrate and, therefore, only a fraction of the soluble β-glucosidase can be involved directly in digestion of cellobiose that accumulates in the immediate environment of the substrate-attached cellulosome.…”

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confidence: 99%

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Enhanced cellulose degradation by targeted integration of a cohesin-fused β-glucosidase into the Clostridium thermocellum cellulosome

Gefen

Anbar

Morag³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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The conversion of recalcitrant plant-derived cellulosic biomass into biofuels is dependent on highly efficient cellulase systems that produce near-quantitative levels of soluble saccharides. Similar to other fungal and bacterial cellulase systems, the multienzyme cellulosome system of the anaerobic, cellulolytic bacterium Clostridium thermocellum is strongly inhibited by the major end product cellobiose. Cellobiose-induced inhibition can be relieved via its cleavage to noninhibitory glucose by the addition of exogenous noncellulosomal enzyme β-glucosidase; however, because the cellulosome is adsorbed to the insoluble substrate only a fraction of β-glucosidase would be available to the cellulosome. Towards this end, we designed a chimeric cohesin-fused β-glucosidase (BglA-CohII) that binds directly to the cellulosome through an unoccupied dockerin module of its major scaffoldin subunit. The β-glucosidase activity is thus focused at the immediate site of cellobiose production by the cellulosomal enzymes. BglA-CohII was shown to retain cellobiase activity and was readily incorporated into the native cellulosome complex. Surprisingly, it was found that the native C. thermocellum cellulosome exists as a homooligomer and the high-affinity interaction of BglA-CohII with the scaffoldin moiety appears to dissociate the oligomeric state of the cellulosome. Complexation of the cellulosome and BglA-CohII resulted in higher overall degradation of microcrystalline cellulose and pretreated switchgrass compared to the native cellulosome alone or in combination with wild-type BglA in solution. These results demonstrate the effect of enzyme targeting and its potential for enhanced degradation of cellulosic biomass.

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Section: Discussionmentioning

confidence: 81%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Enhanced cellulose degradation by targeted integration of a cohesin-fused β-glucosidase into the Clostridium thermocellum cellulosome

Gefen

Anbar

Morag³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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111

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“…A high-efficiency system chooses the appropriate promoter to initiate the transcription of downstream genes. [8][9][10] An inducible promoter can be controlled by inducers, but this increases the complexity of the process in practice. Therefore, many researchers have chosen constitutive promoters for exogenous gene expression.…”

Section: Introductionmentioning

confidence: 99%

Simultaneous saccharification and fermentation of corncobs with genetically modifiedSaccharomyces cerevisiaeand characterization of their microstructure during hydrolysis

et al. 2016

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(2016) Simultaneous saccharification and fermentation of corncobs with genetically modified Saccharomycescerevisiae and characterization of their microstructure during hydrolysis, Bioengineered, 7:3, 198-204, DOI: 10.1080/21655979.2016 ABSTRACTCellulose is an abundant natural polysaccharide that is universally distributed. It can be extracted from corncobs, which are inexpensive, easily accessible, renewable, and environmentally friendly. A common strategy for effectively utilizing cellulose is efficient heterogeneous expression of cellulase genes in Saccharomyces cerevisiae. However, the improvement of cellulose utilization is a relevant issue. Based on our previous findings, we constructed an integrated secretion expression vector, pHBM368-pgk, containing a constitutive promoter sequence. Three genetically modified S. cerevisiae strains containing heterologous b-glucosidase, exoglucanase, and endoglucanase genes were constructed. The results of a 1-L bioreactor fermentation process revealed that the mixed recombinant S. cerevisiae could efficiently carry out simultaneous saccharification and fermentation (SSF) by using corncobs as the sole carbon source. The ethanol concentration reached 6.37 g/L after 96 hours of fermentation, which was about 3 times higher than that produced by genetically modified S. cerevisiae with the inducible promoter sequence. To investigate the microstructure characteristics of hydrolyzed corncobs during the fermentation process, corncob residues were detected by using a scanning electron microscope. This study provides a feasible method to improve the effect of SSF using corncobs as the sole carbon source.

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“…Apesar de não atuarem diretamente na molécula de celulose e por isso não serem consideradas celulases verdadeiras, as β-glicosidases, ou celobiases, são indispensáveis no processo de conversão enzimática da celulose em glicose, pois hidrolisam moléculas de celobiose e celodextrinas liberadas pelas celobiohidrolases (LYND et al, 2002). Além disso, seu efeito sinérgico no sistema celulolítico está relacionado à diminuição da forte inibição das celulases, especialmente celobiohidrolases, causada pelo acúmulo de celobiose (LAMED et al, 1991 (HORN et al, 2012;VAAJE-KOLSTAD et al, 2010). Estudos recentes têm mostrado que a AA9 potencializa a ação das enzimas hidrolíticas, principalmente nas regiões cristalinas, resultando em rendimentos de hidrólise de celulose significativamente mais elevados (HARRIS et al, 2010;HORN et al, 2012;HU et al, 2014).…”

Section: Parede Celular Da Cana-de-açúcarunclassified

Efeito da lignina de bagaços de cana-de-açúcar pré-tratados na hidrólise enzimática da celulose

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Efficient cellulose solubilization by a combined cellulosome-β-glucosidase system

Cited by 47 publications

References 15 publications

Enhanced cellulose degradation by targeted integration of a cohesin-fused β-glucosidase into the Clostridium thermocellum cellulosome

Enhanced cellulose degradation by targeted integration of a cohesin-fused β-glucosidase into the Clostridium thermocellum cellulosome

Simultaneous saccharification and fermentation of corncobs with genetically modifiedSaccharomyces cerevisiaeand characterization of their microstructure during hydrolysis

Efeito da lignina de bagaços de cana-de-açúcar pré-tratados na hidrólise enzimática da celulose

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